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Sample records for transgenic tobacco overexpressing

  1. Overexpression of CsANR increased flavan-3-ols and decreased anthocyanins in transgenic tobacco.

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    Kumar, Vinay; Yadav, Sudesh Kumar

    2013-06-01

    Anthocyanins and flavan-3-ols are distributed widely in plants and synthesized by a common biosynthetic pathway. Anthocyanin reductase (ANR) represents branching-point enzyme of this pathway converting anthocyanidins to flavan-3-ols. Since tea contains highest amount of flavonoids, a cDNA encoding anthocyanin reductase from tea (CsANR) was overexpressed in transgenic tobacco to check the influence on anthocyanin and flavan-3-ols. The transgenic tobacco was confirmed by genomic PCR and expression of transgene was analyzed through semiquantitative PCR. Interestingly flowers of transgenic tobacco were light pink/white in color instead of dark pink in wild tobacco, documenting the decrease in anthocyanins content. Upon measurement, flower anthocyanin content was found to be lesser. While flavan-3-ols (epicatechin and epigallocatechin) contents were increased in leaf tissue of transgenic lines. The expressions of other endogenous flavonoid biosynthetic pathway genes in different floral parts (sepal, petal, stamen, and carpel) of CsANR overexpressing tobacco as well as wild tobacco were analyzed. The transcript levels of PAL and CHI genes were downregulated, while transcript levels of F3H, FLS, CHS, ANR1, and ANR2 genes were upregulated in all floral parts of CsANR transgenic plants compared to wild tobacco. The expressions of DFR and ANS genes were also spatially modulated in different floral parts due to overexpression of CsANR. Thus, CsANR overexpression increased flavan-3-ols and decreased anthocyanin content by modulating the expressions of various flavonoid biosynthetic pathway genes in flower of tobacco. These changes might be responsible for the observed pollen tube in the pollens of CsANR overexpressing transgenic tobacco when they were still in the anther before pollination.

  2. Overexpression of monoubiquitin improves photosynthesis in transgenic tobacco plants following high temperature stress.

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    Tian, Fengxia; Gong, Jiangfeng; Zhang, Jin; Feng, Yanan; Wang, Guokun; Guo, Qifang; Wang, Wei

    2014-09-01

    The ubiquitin/26S proteasome system (Ub/26S) is implicated in abiotic stress responses in plants. In this paper, transgenic tobacco plants overexpressing Ta-Ub2 from wheat were used to study the functions of Ub in the improvement of photosynthesis under high temperature (45°C) stress. We observed higher levels of Ub conjugates in transgenic plants under high temperature stress conditions compared to wild type (WT) as a result of the constitutive overexpression of Ta-Ub2, suggesting increased protein degradation by the 26S proteasome system under high temperature stress. Overexpressing Ub increased the photosynthetic rate (Pn) of transgenic tobacco plants, consistent with the improved ATPase activity in the thylakoid membrane and enhanced efficiency of PSII photochemistry. The higher D1 protein levels following high temperature stress in transgenic plants than WT were also observed. These findings imply that Ub may be involved in tolerance of photosynthesis to high temperature stress in plants. Compared with WT, the transgenic plants showed lower protein carbonylation and malondialdehyde (MDA) levels, less reactive oxygen species (ROS) accumulation, but higher antioxidant enzyme activity under high temperature stress. These findings suggest that the improved antioxidant capacity of transgenic plants may be one of the most important mechanisms underlying Ub-regulated high temperature tolerance. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  3. Over-expression of SlJA2 decreased heat tolerance of transgenic tobacco plants via salicylic acid pathway.

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    Liu, Zhong-Ming; Yue, Meng-Meng; Yang, Dong-Yue; Zhu, Shao-Bo; Ma, Na-Na; Meng, Qing-Wei

    2017-04-01

    Over-expression of SlJA2 decreased the accumulation of SA, which resulted in significant physiological and gene expression changes in transgenic tobacco plants, leading to the decreased heat tolerance of transgenic tobacco. NAC family, the largest transcription factors in plants, responses to different environmental stimuli. Here, we isolated a typical NAC transcription factor (SlJA2) from tomato and got transgenic tobacco with SlJA2 over-expression. Expression of SlJA2 was induced by heat stress (42 °C), chilling stress (4 °C), drought stress, osmotic stress, abscisic acid, and salicylic acid. Over-expression of SlJA2 decreased the accumulation of salicylic acid by regulating expression of salicylic acid degradation gene under heat stress. Compared to WT plants, stomatal apertures and water loss increased in transgenic plants, and the damage of photosynthetic apparatus and chlorophyll breakdown were more serious in transgenic plants under heat stress. Meanwhile, more H 2 O 2 and O 2 ·- were accumulated transgenic plants and proline synthesis was restricted, which resulted in more serious oxidative damage compared to WT. qRT-PCR analysis showed that over-expression of SlJA2 could down-regulate genes involved in reactive oxygen species scavenging, proline biosynthesis, and response to heat stress. All the above results indicated that SlJA2 may be a negative regulator responded to plant's heat tolerance. Thus, this study provides new insight into roles of NAC family member in plant response to abiotic stress.

  4. PDH45 overexpressing transgenic tobacco and rice plants provide salinity stress tolerance via less sodium accumulation.

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    Nath, Manoj; Garg, Bharti; Sahoo, Ranjan Kumar; Tuteja, Narendra

    2015-01-01

    Salinity stress negatively affects the crop productivity worldwide, including that of rice. Coping with these losses is a major concern for all countries. The pea DNA helicase, PDH45 is a unique member of helicase family involved in the salinity stress tolerance. However, the exact mechanism of the PDH45 in salinity stress tolerance is yet to be established. Therefore, the present study was conducted to investigate the mechanism of PDH45-mediated salinity stress tolerance in transgenic tobacco and rice lines along with wild type (WT) plants using CoroNa Green dye based sodium localization in root and shoot sections. The results showed that under salinity stress root and shoot of PDH45 overexpressing transgenic tobacco and rice accumulated less sodium (Na(+)) as compared to their respective WT. The present study also reports salinity tolerant (FL478) and salinity susceptible (Pusa-44) varieties of rice accumulated lowest and highest Na(+) level, respectively. All the varieties and transgenic lines of rice accumulate differential Na(+) ions in root and shoot. However, roots accumulate high Na(+) as compared to the shoots in both tobacco and rice transgenic lines suggesting that the Na(+) transport in shoot is somehow inhibited. It is proposed that the PDH45 is probably involved in the deposition of apoplastic hydrophobic barriers and consequently inhibit Na(+) transport to shoot and therefore confers salinity stress tolerance to PDH45 overexpressing transgenic lines. This study concludes that tobacco (dicot) and rice (monocot) transgenic plants probably share common salinity tolerance mechanism mediated by PDH45 gene.

  5. Overexpression of a tea flavanone 3-hydroxylase gene confers tolerance to salt stress and Alternaria solani in transgenic tobacco.

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    Mahajan, Monika; Yadav, Sudesh Kumar

    2014-08-01

    Flavan-3-ols are the major flavonoids present in tea (Camellia sinensis) leaves. These are known to have antioxidant and free radical scavenging properties in vitro. Flavanone 3-hydroxylase is considered to be an important enzyme of flavonoid pathway leading to accumulation of flavan-3-ols in tea. Expression analysis revealed the upregulation in transcript levels of C. sinensis flavanone 3-hydroxylase (CsF3H) encoding gene under salt stress. In this study, the biotechnological potential of CsF3H was evaluated by gene overexpression in tobacco (Nicotiana tabacum cv. Xanthi). Overexpression of CsF3H cDNA increased the content of flavan-3-ols in tobacco and conferred tolerance to salt stress and fungus Alternaria solani infection. Transgenic tobaccos were observed for increase in primary root length, number of lateral roots, chlorophyll content, antioxidant enzyme expression and their activities. Also, they showed lesser malondialdehyde content and electrolyte leakage compared to control tobacco plants. Further, transgenic plants produced higher degree of pectin methyl esterification via decreasing pectin methyl esterase (PME) activity in roots and leaves under unstressed and salt stressed conditions. The effect of flavan-3-ols on pectin methyl esterification under salt stressed conditions was further validated through in vitro experiments in which non-transgenic (wild) tobacco seedlings were exposed to salt stress in presence of flavan-3-ols, epicatechin and epigallocatechin. The in vitro exposed seedlings showed similar trend of increase in pectin methyl esterification through decreasing PME activity as observed in CsF3H transgenic lines. Taken together, overexpression of CsF3H provided tolerance to salt stress and fungus A. solani infection to transgenic tobacco through improved antioxidant system and enhanced pectin methyl esterification.

  6. Overexpression of the Maize Sulfite Oxidase Increases Sulfate and GSH Levels and Enhances Drought Tolerance in Transgenic Tobacco

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    Zongliang Xia

    2018-03-01

    Full Text Available Sulfite oxidase (SO plays a pivotal role in sulfite metabolism. In our previous study, sulfite-oxidizing function of the SO from Zea mays (ZmSO was characterized. To date, the knowledge of ZmSO’s involvement in abiotic stress response is scarce. In this study, we aimed to investigate the role of ZmSO in drought stress. The transcript levels of ZmSO were relatively high in leaves and immature embryos of maize plants, and were up-regulated markedly by PEG-induced water stress. Overexpression of ZmSO improved drought tolerance in tobacco. ZmSO-overexpressing transgenic plants showed higher sulfate and glutathione (GSH levels but lower hydrogen peroxide (H2O2 and malondialdehyde (MDA contents under drought stress, indicating that ZmSO confers drought tolerance by enhancing GSH-dependent antioxidant system that scavenged ROS and reduced membrane injury. In addition, the transgenic plants exhibited more increased stomatal response than the wild-type (WT to water deficit. Interestingly, application of exogenous GSH effectively alleviated growth inhibition in both WT and transgenic plants under drought conditions. qPCR analysis revealed that the expression of several sulfur metabolism-related genes was significantly elevated in the ZmSO-overexpressing lines. Taken together, these results imply that ZmSO confers enhanced drought tolerance in transgenic tobacco plants possibly through affecting stomatal regulation, GSH-dependent antioxidant system, and sulfur metabolism-related gene expression. ZmSO could be exploited for developing drought-tolerant maize varieties in molecular breeding.

  7. Overexpression of a Plasma Membrane-Localized SbSRP-Like Protein Enhances Salinity and Osmotic Stress Tolerance in Transgenic Tobacco

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    Avinash Mishra

    2017-04-01

    Full Text Available An obligate halophyte, Salicornia brachiata grows in salt marshes and is considered to be a potential resource of salt- and drought-responsive genes. It is important to develop an understanding of the mechanisms behind enhanced salt tolerance. To increase this understanding, a novel SbSRP gene was cloned, characterized, over-expressed, and functionally validated in the model plant Nicotiana tabacum. The genome of the halophyte S. brachiata contains two homologs of an intronless SbSRP gene of 1,262 bp in length that encodes for a stress-related protein. An in vivo localization study confirmed that SbSRP is localized on the plasma membrane. Transgenic tobacco plants (T1 that constitutively over-express the SbSRP gene showed improved salinity and osmotic stress tolerance. In comparison to Wild Type (WT and Vector Control (VC plants, transgenic lines showed elevated relative water and chlorophyll content, lower malondialdehyde content, lower electrolyte leakage and higher accumulation of proline, free amino acids, sugars, polyphenols, and starch under abiotic stress treatments. Furthermore, a lower build-up of H2O2 content and superoxide-radicals was found in transgenic lines compared to WT and VC plants under stress conditions. Transcript expression of Nt-APX (ascorbate peroxidase, Nt-CAT (catalase, Nt-SOD (superoxide dismutase, Nt-DREB (dehydration responsive element binding factor, and Nt-AP2 (apetala2 genes was higher in transgenic lines under stress compared to WT and VC plants. The results suggested that overexpression of membrane-localized SbSRP mitigates salt and osmotic stress in the transgenic tobacco plant. It was hypothesized that SbSRP can be a transporter protein to transmit the environmental stimuli downward through the plasma membrane. However, a detailed study is required to ascertain its exact role in the abiotic stress tolerance mechanism. Overall, SbSRP is a potential candidate to be used for engineering salt and osmotic

  8. Overexpression of the wheat aquaporin gene, TaAQP7, enhances drought tolerance in transgenic tobacco.

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    Shiyi Zhou

    Full Text Available Aquaporin (AQP proteins have been shown to transport water and other small molecules through biological membranes, which is crucial for plants to combat stress caused by drought. However, the precise role of AQPs in drought stress response is not completely understood in plants. In this study, a PIP2 subgroup gene AQP, designated as TaAQP7, was cloned and characterized from wheat. Expression of TaAQP7-GFP fusion protein revealed its localization in the plasma membrane. TaAQP7 exhibited high water channel activity in Xenopus laevis oocytes and TaAQP7 transcript was induced by dehydration, and treatments with polyethylene glycol (PEG, abscisic acid (ABA and H(2O(2. Further, TaAQP7 was upregulated after PEG treatment and was blocked by inhibitors of ABA biosynthesis, implying that ABA signaling was involved in the upregulation of TaAQP7 after PEG treatment. Overexpression of TaAQP7 increased drought tolerance in tobacco. The transgenic tobacco lines had lower levels of malondialdehyde (MDA and H(2O(2, and less ion leakage (IL, but higher relative water content (RWC and superoxide dismutase (SOD and catalase (CAT activities when compared with the wild type (WT under drought stress. Taken together, our results show that TaAQP7 confers drought stress tolerance in transgenic tobacco by increasing the ability to retain water, reduce ROS accumulation and membrane damage, and enhance the activities of antioxidants.

  9. Overexpression of a specific soybean GmGSTU4 isoenzyme improves diphenyl ether and chloroacetanilide herbicide tolerance of transgenic tobacco plants.

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    Benekos, Kostantinos; Kissoudis, Christos; Nianiou-Obeidat, Irini; Labrou, Nikolaos; Madesis, Panagiotis; Kalamaki, Mary; Makris, Antonis; Tsaftaris, Athanasios

    2010-10-01

    Plant glutathione transferases (GSTs) superfamily consists of multifunctional enzymes and forms a major part of the plants herbicide detoxification enzyme network. The tau class GST isoenzyme GmGSTU4 from soybean, exhibits catalytic activity towards the diphenyl ether herbicide fluorodifen and is active as glutathione-dependent peroxidase (GPOX). Transgenic tobacco plants of Basmas cultivar were generated via Agrobacterium transformation. The aim was to evaluate in planta, GmGSTU4's role in detoxifying the diphenyl ether herbicides fluorodifen and oxyfluorfen and the chloroacetanilides alachlor and metolachlor. Transgenic tobacco plants were verified by PCR and Southern blot hybridization and expression of GmGSTU4 was determined by RT-PCR. Leaf extracts from transgenic plants showed moderate increase in GST activity towards CDNB and a significant increase towards fluorodifen and alachlor, and at the same time an increased GPOX activity towards cumene hydroperoxide. GmGSTU4 overexpressing plants when treated with 200 μM fluorodifen or oxyfluorfen exhibited reduced relative electrolyte leakage compared to wild type plants. Moreover all GmGSTU4 overexpressing lines exhibited significantly increased tolerance towards alachlor when grown in vitro at 7.5 mg/L alachlor compared to wild type plants. No significant increased tolerance was observed to metolachlor. These results confirm the contribution of this particular GmGSTU4 isoenzyme from soybean in the detoxification of fluorodifen and alachlor, and provide the basis towards the development of transgenic plants with improved phytoremediation capabilities for future use in environmental cleanup of herbicides. Copyright © 2010 Elsevier B.V. All rights reserved.

  10. Enhanced cadmium accumulation and tolerance in transgenic tobacco overexpressing rice metal tolerance protein gene OsMTP1 is promising for phytoremediation.

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    Das, Natasha; Bhattacharya, Surajit; Maiti, Mrinal K

    2016-08-01

    One of the most grievous heavy metal pollutants in the environment is cadmium (Cd), which is not only responsible for the crop yield loss owing to its phytotoxicity, but also for the human health hazards as the toxic elements usually accumulate in the consumable parts of crop plants. In the present study, we aimed to isolate and functionally characterize the OsMTP1 gene from indica rice (Oryza sativa L. cv. IR64) to study its potential application for efficient phytoremediation of Cd. The 1257 bp coding DNA sequence (CDS) of OsMTP1 encodes a ∼46 kDa protein belonging to the cation diffusion facilitator (CDF) or metal tolerance/transport protein (MTP) family. The OsMTP1 transcript in rice plant was found to respond during external Cd stress. Heterologous expression of OsMTP1 in tobacco resulted in the reduction of Cd stress-induced phytotoxic effects, including growth inhibition, lipid peroxidation, and cell death. Compared to untransformed control, the transgenic tobacco plants showed enhanced vacuolar thiol content, indicating vacuolar localization of the sequestered Cd. The transgenic tobacco plants exhibited significantly higher biomass growth (2.2-2.8-folds) and hyperaccumulation of Cd (1.96-2.22-folds) compared to untransformed control under Cd exposure. The transgenic plants also showed moderate tolerance and accumulation of arsenic (As) upon exogenous As stress, signifying broad substrate specificity of OsMTP1. Together, findings of our research suggest that the transgenic tobacco plants overexpressing OsMTP1 with its hyperaccumulating activity and increased growth rate could be useful for future phytoremediation applications to clean up the Cd-contaminated soil. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  11. Ectopic expression of wheat expansin gene TaEXPA2 improved the salt tolerance of transgenic tobacco by regulating Na+ /K+ and antioxidant competence.

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    Chen, Yanhui; Han, Yangyang; Kong, Xiangzhu; Kang, Hanhan; Ren, Yuanqing; Wang, Wei

    2017-02-01

    High salinity is one of the most serious environmental stresses that limit crop growth. Expansins are cell wall proteins that regulate plant development and abiotic stress tolerance by mediating cell wall expansion. We studied the function of a wheat expansin gene, TaEXPA2, in salt stress tolerance by overexpressing it in tobacco. Overexpression of TaEXPA2 enhanced the salt stress tolerance of transgenic tobacco plants as indicated by the presence of higher germination rates, longer root length, more lateral roots, higher survival rates and more green leaves under salt stress than in the wild type (WT). Further, when leaf disks of WT plants were incubated in cell wall protein extracts from the transgenic tobacco plants, their chlorophyll content was higher under salt stress, and this improvement from TaEXPA2 overexpression in transgenic tobacco was inhibited by TaEXPA2 protein antibody. The water status of transgenic tobacco plants was improved, perhaps by the accumulation of osmolytes such as proline and soluble sugar. TaEXPA2-overexpressing tobacco lines exhibited lower Na + but higher K + accumulation than WT plants. Antioxidant competence increased in the transgenic plants because of the increased activity of antioxidant enzymes. TaEXPA2 protein abundance in wheat was induced by NaCl, and ABA signaling was involved. Gene expression regulation was involved in the enhanced salt stress tolerance of the TaEXPA2 transgenic plants. Our results suggest that TaEXPA2 overexpression confers salt stress tolerance on the transgenic plants, and this is associated with improved water status, Na + /K + homeostasis, and antioxidant competence. ABA signaling participates in TaEXPA2-regulated salt stress tolerance. © 2016 Scandinavian Plant Physiology Society.

  12. Over-expression of ascorbate oxidase in the apoplast of transgenic tobacco results in altered ascorbate and glutathione redox states and increased sensitivity to ozone

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    Sanmartin, Maite; Drogoudi, Pavlina D.; Lyons, Tom

    2003-01-01

    overexpressing plants exposed to 100 nmol mol-1 ozone for 7 h day-1 exhibited a substantial increase in foliar injury, and a greater pollutant-induced reduction in both the light-saturated rate of CO2 assimilation and the maximum in vivo rate of ribulose-1,5-bisphosphate carboxylase/oxygenase carboxylation......Transgenic tobacco (Nicotiana tabacum L. cv. Xanthi) plants expressing cucumber ascorbate oxidase (EC.1.10.3.3) were used to examine the role of extracellular ascorbic acid in mediating tolerance to the ubiquitous air pollutant, ozone (O3). Three homozygous transgenic lines, chosen on the basis...

  13. Overexpression of a Triticum aestivum Calreticulin gene (TaCRT1 Improves Salinity Tolerance in Tobacco.

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    Yang Xiang

    Full Text Available Calreticulin (CRT is a highly conserved and abundant multifunctional protein that is encoded by a small gene family and is often associated with abiotic/biotic stress responses in plants. However, the roles played by this protein in salt stress responses in wheat (Triticum aestivum remain obscure. In this study, three TaCRT genes were identified in wheat and named TaCRT1, TaCRT2 and TaCRT3-1 based on their sequence characteristics and their high homology to other known CRT genes. Quantitative real-time PCR expression data revealed that these three genes exhibit different expression patterns in different tissues and are strongly induced under salt stress in wheat. The calcium-binding properties of the purified recombinant TaCRT1 protein were determined using a PIPES/Arsenazo III analysis. TaCRT1 gene overexpression in Nicotiana tabacum decreased salt stress damage in transgenic tobacco plants. Physiological measurements indicated that transgenic tobacco plants showed higher activities of superoxide dismutase (SOD, peroxidase (POD and catalase (CAT than non-transgenic tobacco under normal growth conditions. Interestingly, overexpression of the entire TaCRT1 gene or of partial TaCRT1 segments resulted in significantly higher tolerance to salt stress in transgenic plants compared with their WT counterparts, thus revealing the essential role of the C-domain of TaCRT1 in countering salt stress in plants.

  14. Dehydrins Impart Protection against Oxidative Stress in Transgenic Tobacco Plants.

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    Halder, Tanmoy; Upadhyaya, Gouranga; Basak, Chandra; Das, Arup; Chakraborty, Chandrima; Ray, Sudipta

    2018-01-01

    Environmental stresses generate reactive oxygen species (ROS) which might be detrimental to the plants when produced in an uncontrolled way. However, the plants ameliorate such stresses by synthesizing antioxidants and enzymes responsible for the dismutation of ROS. Additionally, the dehydrins were also able to protect the inactivation of the enzyme lactate dehydrogenase against hydroxyl radicals (OH ⋅ ) generated during Fenton's reaction. SbDhn1 and SbDhn2 overexpressing transgenic tobacco plants were able to protect against oxidative damage. Transgenic tobacco lines showed better photosynthetic efficiency along with high chlorophyll content, soluble sugar and proline. However, the malonyl dialdehyde (MDA) content was significantly lower in transgenic lines. Experimental evidence demonstrates the protective effect of dehydrins on electron transport chain in isolated chloroplast upon methyl viologen (MV) treatment. The transgenic tobacco plants showed significantly lower superoxide radical generation () upon MV treatment. The accumulation of the H 2 O 2 was also lower in the transgenic plants. Furthermore, in the transgenic plants the expression of ROS scavenging enzymes was higher compared to non-transformed (NT) or vector transformed (VT) plants. Taken together these data, during oxidative stress dehydrins function by scavenging the () directly and also by rendering protection to the enzymes responsible for the dismutation of () thereby significantly reducing the amount of hydrogen peroxides formed. Increase in proline content along with other antioxidants might also play a significant role in stress amelioration. Dehydrins thus function co-operatively with other protective mechanisms under oxidative stress conditions rendering protection in stress environment.

  15. Overexpression of MfPIP2-7 from Medicago falcata promotes cold tolerance and growth under NO3 (-) deficiency in transgenic tobacco plants.

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    Zhuo, Chunliu; Wang, Ting; Guo, Zhenfei; Lu, Shaoyun

    2016-06-14

    Plasma membrane intrinsic proteins (PIPs), which belong to aquaporins (AQPs) superfamily, are subdivided into two groups, PIP1 and PIP2, based on sequence similarity. Several PIP2s function as water channels, while PIP1s have low or no water channel activity, but have a role in water permeability through interacting with PIP2. A cold responsive PIP2 named as MfPIP2-7 was isolated from Medicago falcata (hereafter falcata), a forage legume with great cold tolerance, and transgenic tobacco plants overexpressing MfPIP2-7 were analyzed in tolerance to multiple stresses including freezing, chilling, and nitrate reduction in this study. MfPIP2-7 transcript was induced by 4 to 12 h of cold treatment and 2 h of abscisic acid (ABA) treatment. Pretreatment with inhibitor of ABA synthesis blocked the cold induced MfPIP2-7 transcript, indicating that ABA was involved in cold induced transcription of MfPIP2-7 in falcata. Overexpression of MfPIP2-7 resulted in enhanced tolerance to freezing, chilling and NO3 (-) deficiency in transgenic tobacco (Nicotiana tabacum L.) plants as compared with the wild type. Moreover, MfPIP2-7 was demonstrated to facilitate H2O2 diffusion in yeast. Higher transcript levels of several stress responsive genes, such as NtERD10B, NtERD10C, NtDREB1, and 2, and nitrate reductase (NR) encoding genes (NtNIA1, and NtNIA2) were observed in transgenic plants as compared with the wild type with dependence upon H2O2. In addition, NR activity was increased in transgenic plants, which led to alterations in free amino acid components and concentrations. The results suggest that MfPIP2-7 plays an important role in plant tolerance to freezing, chilling, and NO3 (-) deficiency by promoted H2O2 diffusion that in turn up-regulates expression of NIAs and multiple stress responsive genes.

  16. An overexpression of chalcone reductase of Pueraria montana var. lobata alters biosynthesis of anthocyanin and 5'-deoxyflavonoids in transgenic tobacco.

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    Joung, Jae-youl; Kasthuri, G Mangai; Park, Ji-young; Kang, Won-jin; Kim, Hyun-soon; Yoon, Bong-sik; Joung, Hyouk; Jeon, Jae-heung

    2003-03-28

    We isolated the chalcone reductase (pl-chr) gene of Pueraria montana var. lobata by using a PCR strategy from cDNA pools of storage roots. A high level of expression of RNA was found in both stems and roots. The genomic Southern blot result suggests that pl-chr exists as a member of a small gene family. By introducing a pl-chr gene under the control of the 35S CaMV promoter into the pink-flowering Xanthi line of Nicotiana tabacum, the flower color was changed from pink to white-to-pink. The contents of anthocyanin in the flowers of the transgenic lines were dramatically decreased by 40%, but the total UV absorption compounds remained unchanged. The production of liquiritigenin in pl-chr overexpressed transgenic tobacco lines was confirmed by HPLC and MS analysis. The introduction of pl-chr gene provides a method to redirect the flavonoid pathway into 5'-deoxyflavonoid production in non-legume crops, in order to manipulate the phenylpropanoid pathway for isoflavonoid production.

  17. Overexpression of a New Chitinase Gene EuCHIT2 Enhances Resistance to Erysiphe cichoracearum DC. in Tobacco Plants

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    Xuan Dong

    2017-11-01

    Full Text Available In this study, we cloned a new chitinase gene, EuCHIT2, from Eucommia ulmoides Oliver (E. ulmoides using rapid amplification of cDNA ends (RACE technology and constructed an overexpression vector, pSH-35S-EuCHIT2, to introduce it into tobacco (Nicotiana tabacum cv. Xanthi. Resistance to Erysiphe cichoracearum de Candolle (E.cichoracearum DC and molecular mechanisms in the transgenic tobacco were determined by drop inoculation, spore counting, determination of physicochemical indicators, and analysis of gene expression. The chitinase activity and resistance to E. cichoracearum DC were significantly higher in the transgenic tobacco than in wild-type tobacco (p < 0.05. The activities of peroxidase (POD and catalase (CAT, after inoculation with E. cichoracearum DC, were higher in the transgenic tobacco than in the wild-type. Conversely, the malondialdehyde (MDA content was significantly lower in the transgenic tobacco than the wild-type before and after inoculation. In addition, our study also indicated that the resistance to E. cichoracearum DC might involve the salicylic acid (SA and jasmonic acid (JA pathways, because the expression levels of pathogenesis-related gene 1 (PR-1a and coronatine-insensitive 1 (COI1 were significantly increased and decreased, respectively, after inoculation with E. cichoracearum DC. The present study supports the notion that PR-1a and POD participate in resistance to E. cichoracearum DC in the transgenic tobacco plants.

  18. A CBL-Interacting Protein Kinase TaCIPK2 Confers Drought Tolerance in Transgenic Tobacco Plants through Regulating the Stomatal Movement.

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    Wang, Yan; Sun, Tao; Li, Tingting; Wang, Meng; Yang, Guangxiao; He, Guangyuan

    2016-01-01

    In plants, the CBL-CIPK signaling pathways play key roles in the response to abiotic stresses. However, functional studies of CIPKs in the important staple crop wheat are very rare. In this study, we identified a CIPK gene from wheat, designated TaCIPK2. Expression analysis results showed that TaCIPK2 could be up-regulated in wheat leaves by polyethylene glycol, abscisic acid and H2O2 treatments. Subcellular localization analyses revealed that TaCIPK2 was present in whole wheat epidermal cells. A yeast two-hybrid assay indicated that TaCIPK2 interacted with TaCBL1, 2, 3 and 4 in vitro. Transgenic tobacco plants over-expressing TaCIPK2 exhibited increased drought tolerance, indicated by a larger proportion of green cotyledons and higher survival rates under the osmotic and drought stress conditions compared with control plants. Additionally, physiological index analyses revealed that the transgenic tobacco plants had lower water loss rates and ion leakage, accumulated less malondialdehyde and H2O2, and had higher catalase and superoxide dismutase activities than the control plants. The transgenic plants also exhibited faster stomatal closure following exposure to osmotic stress conditions. The seed germination rates and stomatal aperture of TaCIPK2-overexpressing tobacco plants decreased after exogenous abscisic acid treatment was applied, implying that the transgenic tobacco plants were more sensitive to exogenous abscisic acid than the control plants. Our results indicate that TaCIPK2 plays a positive regulatory role in drought stress responses in transgenic tobacco plants.

  19. Co-expression of peppermint geranyl diphosphate synthase small subunit enhances monoterpene production in transgenic tobacco plants.

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    Yin, Jun-Lin; Wong, Woon-Seng; Jang, In-Cheol; Chua, Nam-Hai

    2017-02-01

    Monoterpenes are important for plant survival and useful to humans. In addition to their function in plant defense, monoterpenes are also used as flavors, fragrances and medicines. Several metabolic engineering strategies have been explored to produce monoterpene in tobacco but only trace amounts of monoterpenes have been detected. We investigated the effects of Solanum lycopersicum 1-deoxy-d-xylulose-5-phosphate synthase (SlDXS), Arabidopsis thaliana geranyl diphosphate synthase 1 (AtGPS) and Mentha × piperita geranyl diphosphate synthase small subunit (MpGPS.SSU) on production of monoterpene and geranylgeranyl diphosphate (GGPP) diversities, and plant morphology by transient expression in Nicotiana benthamiana and overexpression in transgenic Nicotiana tabacum. We showed that MpGPS.SSU could enhance the production of various monoterpenes such as (-)-limonene, (-)-linalool, (-)-α-pinene/β-pinene or myrcene, in transgenic tobacco by elevating geranyl diphosphate synthase (GPS) activity. In addition, overexpression of MpGPS.SSU in tobacco caused early flowering phenotype and increased shoot branching by elevating contents of GA 3 and cytokinins due to upregulated transcript levels of several plastidic 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway genes, geranylgeranyl diphosphate synthases 3 (GGPPS3) and GGPPS4. Our method would allow the identification of new monoterpene synthase genes using transient expression in N. benthamiana and the improvement of monoterpene production in transgenic tobacco plants. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

  20. Heterologous production of a ginsenoside saponin (compound K) and its precursors in transgenic tobacco impairs the vegetative and reproductive growth.

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    Gwak, Yu Shin; Han, Jung Yeon; Adhikari, Prakash Babu; Ahn, Chang Ho; Choi, Yong Eui

    2017-06-01

    Production of compound K (a ginsenoside saponin) and its precursors in transgenic tobacco resulted in stunted growth and seed set failure, which may be caused by strong autotoxicity of heterologously produced phytochemicals against the tobacco itself. Panax ginseng roots contain various saponins (ginsenosides), which are major bioactive compounds. A monoglucosylated saponin, compound K (20-O-(β-D-glucopyranosyl)-20(S)-protopanaxadiol), has high medicinal and cosmetic values but is present in undetectable amounts in naturally grown ginseng roots. The production of compound K (CK) requires complicated deglycosylation of ginsenosides using physicochemical and/or enzymatic degradation. In this work, we report the production of CK in transgenic tobacco by co-overexpressing three genes (PgDDS, CYP716A47 and UGT71A28) isolated from P. ginseng. Introduction and expression of the transgenes in tobacco lines were confirmed by genomic PCR and RT-PCR. All the lines of transgenic tobacco produced CK including its precursors, protopanaxadiol and dammarenediol-II (DD). The concentrations of CK in the leaves ranged from 1.55 to 2.64 µg/g dry weight, depending on the transgenic line. Interestingly, production of CK in tobacco brought stunted plant growth and gave rise to seed set failure. This seed set failure was caused by both long-styled flowers and abnormal pollen development in transgenic tobacco. Both CK and DD treatments highly suppressed in vitro germination and tube growth in wild-type pollens. Based on these results, metabolic engineering for CK production in transgenic tobacco was successfully achieved, but the production of CK and its precursors in tobacco severely affects vegetative and reproductive growth due to the cytotoxicity of phytochemicals that are heterologously produced in transgenic tobacco.

  1. Overexpression of the Wheat Expansin Gene TaEXPA2 Improved Seed Production and Drought Tolerance in Transgenic Tobacco Plants.

    Science.gov (United States)

    Chen, Yanhui; Han, Yangyang; Zhang, Meng; Zhou, Shan; Kong, Xiangzhu; Wang, Wei

    2016-01-01

    Expansins are cell wall proteins that are grouped into two main families, α-expansins and β-expansins, and they are implicated in the control of cell extension via the disruption of hydrogen bonds between cellulose and matrix glucans. TaEXPA2 is an α-expansin gene identified in wheat. Based on putative cis-regulatory elements in the TaEXPA2 promoter sequence and the expression pattern induced when polyethylene glycol (PEG) is used to mimic water stress, we hypothesized that TaEXPA2 is involved in plant drought tolerance and plant development. Through transient expression of 35S::TaEXPA2-GFP in onion epidermal cells, TaEXPA2 was localized to the cell wall. Constitutive expression of TaEXPA2 in tobacco improved seed production by increasing capsule number, not seed size, without having any effect on plant growth patterns. The transgenic tobacco exhibited a significantly greater tolerance to water-deficiency stress than did wild-type (WT) plants. We found that under drought stress, the transgenic plants maintained a better water status. The accumulated content of osmotic adjustment substances, such as proline, in TaEXPA2 transgenic plants was greater than that in WT plants. Transgenic plants also displayed greater antioxidative competence as indicated by their lower malondialdehyde (MDA) content, relative electrical conductivity, and reactive oxygen species (ROS) accumulation than did WT plants. This result suggests that the transgenic plants suffer less damage from ROS under drought conditions. The activities of some antioxidant enzymes as well as expression levels of several genes encoding key antioxidant enzymes were higher in the transgenic plants than in the WT plants under drought stress. Collectively, our results suggest that ectopic expression of the wheat expansin gene TaEXPA2 improves seed production and drought tolerance in transgenic tobacco plants.

  2. Changes in cell wall properties coincide with overexpression of extensin fusion proteins in suspension cultured tobacco cells.

    Science.gov (United States)

    Tan, Li; Pu, Yunqiao; Pattathil, Sivakumar; Avci, Utku; Qian, Jin; Arter, Allison; Chen, Liwei; Hahn, Michael G; Ragauskas, Arthur J; Kieliszewski, Marcia J

    2014-01-01

    Extensins are one subfamily of the cell wall hydroxyproline-rich glycoproteins, containing characteristic SerHyp4 glycosylation motifs and intermolecular cross-linking motifs such as the TyrXaaTyr sequence. Extensins are believed to form a cross-linked network in the plant cell wall through the tyrosine-derivatives isodityrosine, pulcherosine, and di-isodityrosine. Overexpression of three synthetic genes encoding different elastin-arabinogalactan protein-extensin hybrids in tobacco suspension cultured cells yielded novel cross-linking glycoproteins that shared features of the extensins, arabinogalactan proteins and elastin. The cell wall properties of the three transgenic cell lines were all changed, but in different ways. One transgenic cell line showed decreased cellulose crystallinity and increased wall xyloglucan content; the second transgenic cell line contained dramatically increased hydration capacity and notably increased cell wall biomass, increased di-isodityrosine, and increased protein content; the third transgenic cell line displayed wall phenotypes similar to wild type cells, except changed xyloglucan epitope extractability. These data indicate that overexpression of modified extensins may be a route to engineer plants for bioenergy and biomaterial production.

  3. Overexpression of snapdragon Delila (Del) gene in tobacco enhances anthocyanin accumulation and abiotic stress tolerance

    OpenAIRE

    Naing, Aung Htay; Park, Kyeung Il; Ai, Trinh Ngoc; Chung, Mi Young; Han, Jeung Sul; Kang, Young-Wha; Lim, Ki Byung; Kim, Chang Kil

    2017-01-01

    Background Rosea1 (Ros1) and Delila (Del) co-expression controls anthocyanin accumulation in snapdragon flowers, while their overexpression in tomato strongly induces anthocyanin accumulation. However, little data exist on how Del expression alone influences anthocyanin accumulation. Results In tobacco (Nicotiana tabacum ?Xanthi?), Del expression enhanced leaf and flower anthocyanin production through regulating NtCHS, NtCHI, NtF3H, NtDFR, and NtANS transcript levels. Transgenic lines display...

  4. Genetically pyramiding protease-inhibitor genes for dual broad-spectrum resistance against insect and phytopathogens in transgenic tobacco.

    Science.gov (United States)

    Senthilkumar, Rajendran; Cheng, Chiu-Ping; Yeh, Kai-Wun

    2010-01-01

    Protease inhibitors provide a promising means of engineering plant resistance against attack by insects and pathogens. Sporamin (trypsin inhibitor) from sweet potato and CeCPI (phytocystatin) from taro were stacked in a binary vector, using pMSPOA (a modified sporamin promoter) to drive both genes. Transgenic tobacco lines of T0 and T1 generation with varied inhibitory activity against trypsin and papain showed resistance to both insects and phytopathogens. Larvae of Helicoverpa armigera that ingested tobacco leaves either died or showed delayed growth and development relative to control larvae. Transgenic tobacco-overexpressing the stacked genes also exhibited strong resistance against bacterial soft rot disease caused by Erwinia carotovora and damping-off disease caused by Pythium aphanidermatum. Thus, stacking protease-inhibitor genes, driven by the wound and pathogen responsive pMSPOA promoter, is an effective strategy for engineering crops to resistance against insects and phytopathogens.

  5. Overexpression of a cytosolic abiotic stress responsive universal stress protein (SbUSP mitigates salt and osmotic stress in transgenic tobacco plants

    Directory of Open Access Journals (Sweden)

    Pushpika eUdawat

    2016-04-01

    Full Text Available The Universal Stress Protein (USP is a ubiquitous protein and plays an indispensable role in plant abiotic stress tolerance. The genome of Salicornia brachiata contains two homologues of intron less SbUSP gene which encodes for salt and osmotic responsive universal stress protein. In vivo localization reveals that SbUSP is a membrane bound cytosolic protein. The role of the gene was functionally validated by developing transgenic tobacco and compared with control (wild type and vector control plants under different abiotic stress condition. Transgenic lines (T1 exhibited higher chlorophyll, relative water, proline, total sugar, reducing sugar, free amino acids, polyphenol contents, osmotic potential, membrane stability and lower electrolyte leakage and lipid peroxidation (malondialdehyde content under stress treatments than control (WT and VC plants. Lower accumulation of H2O2 and O2- radicals was also detected in transgenic lines compared to control plants under stress conditions. Present study confers that overexpression of the SbUSP gene enhances plant growth, alleviates ROS buildup, maintains ion homeostasis and improves the physiological status of the plant under salt and osmotic stresses. Principal component analysis (PCA exhibited a statistical distinction of plant response to salinity stress, and a significant response was observed for transgenic lines under stress, which provides stress endurance to the plant. A possible signaling role is proposed that some downstream genes may get activated by abiotic stress responsive cytosolic SbUSP, which leads to the protection of cell from oxidative damages. The study unveils that ectopic expression of the gene mitigates salt or osmotic stress by scavenging ROS and modulating the physiological process of the plant.

  6. Overexpression of a Cytosolic Abiotic Stress Responsive Universal Stress Protein (SbUSP) Mitigates Salt and Osmotic Stress in Transgenic Tobacco Plants

    Science.gov (United States)

    Udawat, Pushpika; Jha, Rajesh K.; Sinha, Dinkar; Mishra, Avinash; Jha, Bhavanath

    2016-01-01

    The universal stress protein (USP) is a ubiquitous protein and plays an indispensable role in plant abiotic stress tolerance. The genome of Salicornia brachiata contains two homologs of intron less SbUSP gene which encodes for salt and osmotic responsive USP. In vivo localization reveals that SbUSP is a membrane bound cytosolic protein. The role of the gene was functionally validated by developing transgenic tobacco and compared with control [wild-type (WT) and vector control (VC)] plants under different abiotic stress condition. Transgenic lines (T1) exhibited higher chlorophyll, relative water, proline, total sugar, reducing sugar, free amino acids, polyphenol contents, osmotic potential, membrane stability, and lower electrolyte leakage and lipid peroxidation (malondialdehyde content) under stress treatments than control (WT and VC) plants. Lower accumulation of H2O2 and O2− radicals was also detected in transgenic lines compared to control plants under stress conditions. Present study confers that overexpression of the SbUSP gene enhances plant growth, alleviates ROS buildup, maintains ion homeostasis and improves the physiological status of the plant under salt and osmotic stresses. Principal component analysis exhibited a statistical distinction of plant response to salinity stress, and a significant response was observed for transgenic lines under stress, which provides stress endurance to the plant. A possible signaling role is proposed that some downstream genes may get activated by abiotic stress responsive cytosolic SbUSP, which leads to the protection of cell from oxidative damages. The study unveils that ectopic expression of the gene mitigates salt or osmotic stress by scavenging ROS and modulating the physiological process of the plant. PMID:27148338

  7. Over-expression of a Rab family GTPase from phreatophyte Prosopis juliflora confers tolerance to salt stress on transgenic tobacco.

    Science.gov (United States)

    George, Suja; Parida, Ajay

    2011-03-01

    Plant growth and productivity are adversely affected by various abiotic and biotic stress factors. In our previous study, we used Prosopis juliflora, an abiotic stress tolerant tree species of Fabaceae, as a model plant system for isolating genes functioning in abiotic stress tolerance. Here we report the isolation and characterization of a Rab family GTPase from P. juliflora (Pj Rab7) and the ability of this gene to confer salt stress tolerance in transgenic tobacco. Northern analysis for Pj Rab7 in P. juliflora leaf tissue revealed up-regulation of this gene under salt stress under the concentrations and time points analyzed. Pj Rab7 transgenic tobacco lines survived better under conditions of 150 mM NaCl stress compared to control un-transformed plants. Pj Rab7 transgenic plants were found to accumulate more sodium than control plants during salt stress. The results of our studies could be used as a starting point for generation of crop plants tolerant to abiotic stress.

  8. Phenotypic Changes in Transgenic Tobacco Plants Overexpressing Vacuole-Targeted Thermotoga maritima BglB Related to Elevated Levels of Liberated Hormones

    Science.gov (United States)

    Nguyen, Quynh Anh; Lee, Dae-Seok; Jung, Jakyun; Bae, Hyeun-Jong

    2015-01-01

    The hyperthermostable β-glucosidase BglB of Thermotoga maritima was modified by adding a short C-terminal tetrapeptide (AFVY, which transports phaseolin to the vacuole, to its C-terminal sequence). The modified β-glucosidase BglB was transformed into tobacco (Nicotiana tabacum L.) plants. We observed a range of significant phenotypic changes in the transgenic plants compared to the wild-type (WT) plants. The transgenic plants had faster stem growth, earlier flowering, enhanced root systems development, an increased biomass biosynthesis rate, and higher salt stress tolerance in young plants compared to WT. In addition, programed cell death was enhanced in mature plants. Furthermore, the C-terminal AFVY tetrapeptide efficiently sorted T. maritima BglB into the vacuole, which was maintained in an active form and could perform its glycoside hydrolysis function on hormone conjugates, leading to elevated hormone [abscisic acid (ABA), indole 3-acetic acid (IAA), and cytokinin] levels that likely contributed to the phenotypic changes in the transgenic plants. The elevation of cytokinin led to upregulation of the transcription factor WUSCHELL, a homeodomain factor that regulates the development, division, and reproduction of stem cells in the shoot apical meristems. Elevation of IAA led to enhanced root development, and the elevation of ABA contributed to enhanced tolerance to salt stress and programed cell death. These results suggest that overexpressing vacuole-targeted T. maritima BglB may have several advantages for molecular farming technology to improve multiple targets, including enhanced production of the β-glucosidase BglB, increased biomass, and shortened developmental stages, that could play pivotal roles in bioenergy and biofuel production. PMID:26618153

  9. Overexpression of host plant urease in transgenic silkworms.

    Science.gov (United States)

    Jiang, Liang; Huang, Chunlin; Sun, Qiang; Guo, Huizhen; Peng, Zhengwen; Dang, Yinghui; Liu, Weiqiang; Xing, Dongxu; Xu, Guowen; Zhao, Ping; Xia, Qingyou

    2015-06-01

    Bombyx mori and mulberry constitute a model of insect-host plant interactions. Urease hydrolyzes urea to ammonia and is important for the nitrogen metabolism of silkworms because ammonia is assimilated into silk protein. Silkworms do not synthesize urease and acquire it from mulberry leaves. We synthesized the artificial DNA sequence ureas using the codon bias of B. mori to encode the signal peptide and mulberry urease protein. A transgenic vector that overexpresses ure-as under control of the silkworm midgut-specific P2 promoter was constructed. Transgenic silkworms were created via embryo microinjection. RT-PCR results showed that urease was expressed during the larval stage and qPCR revealed the expression only in the midgut of transgenic lines. Urea concentration in the midgut and hemolymph of transgenic silkworms was significantly lower than in a nontransgenic line when silkworms were fed an artificial diet. Analysis of the daily body weight and food conversion efficiency of the fourth and fifth instar larvae and economic characteristics indicated no differences between transgenic silkworms and the nontransgenic line. These results suggested that overexpression of host plant urease promoted nitrogen metabolism in silkworms.

  10. A spontaneous dominant-negative mutation within a 35S::AtMYB90 transgene inhibits flower pigment production in tobacco.

    Science.gov (United States)

    Velten, Jeff; Cakir, Cahid; Cazzonelli, Christopher I

    2010-03-29

    In part due to the ease of visual detection of phenotypic changes, anthocyanin pigment production has long been the target of genetic and molecular research in plants. Specific members of the large family of plant myb transcription factors have been found to play critical roles in regulating expression of anthocyanin biosynthetic genes and these genes continue to serve as important tools in dissecting the molecular mechanisms of plant gene regulation. A spontaneous mutation within the coding region of an Arabidopsis 35S::AtMYB90 transgene converted the activator of plant-wide anthocyanin production to a dominant-negative allele (PG-1) that inhibits normal pigment production within tobacco petals. Sequence analysis identified a single base change that created a premature nonsense codon, truncating the encoded myb protein. The resulting mutant protein lacks 78 amino acids from the wild type C-terminus and was confirmed as the source of the white-flower phenotype. A putative tobacco homolog of AtMYB90 (NtAN2) was isolated and found to be expressed in flower petals but not leaves of all tobacco plants tested. Using transgenic tobacco constitutively expressing the NtAN2 gene confirmed the NtAN2 protein as the likely target of PG-1-based inhibition of tobacco pigment production. Messenger RNA and anthocyanin analysis of PG-1Sh transgenic lines (and PG-1Sh x purple 35S::NtAN2 seedlings) support a model in which the mutant myb transgene product acts as a competitive inhibitor of the native tobacco NtAN2 protein. This finding is important to researchers in the field of plant transcription factor analysis, representing a potential outcome for experiments analyzing in vivo protein function in test transgenic systems that over-express or mutate plant transcription factors.

  11. A Wheat R2R3-type MYB Transcription Factor TaODORANT1 Positively Regulates Drought and Salt Stress Responses in Transgenic Tobacco Plants

    Directory of Open Access Journals (Sweden)

    Qiuhui Wei

    2017-08-01

    Full Text Available MYB transcription factors play important roles in plant responses to biotic and abiotic stress. In this study, TaODORANT1, a R2R3-MYB gene, was cloned from wheat (Triticum aestivum L.. TaODORANT1 was localized in the nucleus and functioned as a transcriptional activator. TaODORANT1 was up-regulated in wheat under PEG6000, NaCl, ABA, and H2O2 treatments. TaODORANT1-overexpressing transgenic tobacco plants exhibited higher relative water content and lower water loss rate under drought stress, as well as lower Na+ accumulation in leaves under salt stress. The transgenic plants showed higher CAT activity but lower ion leakage, H2O2 and malondialdehyde contents under drought and salt stresses. Besides, the transgenic plants also exhibited higher SOD activity under drought stress. Our results also revealed that TaODORANT1 overexpression up-regulated the expression of several ROS- and stress-related genes in response to both drought and salt stresses, thus enhancing transgenic tobacco plants tolerance. Our studies demonstrate that TaODORANT1 positively regulates plant tolerance to drought and salt stresses.

  12. Development of a transgenic tobacco plant for phytoremediation of methylmercury pollution.

    Science.gov (United States)

    Nagata, Takeshi; Morita, Hirofumi; Akizawa, Toshifumi; Pan-Hou, Hidemitsu

    2010-06-01

    To develop the potential of plant for phytoremediation of methylmercury pollution, a genetically engineered tobacco plant that coexpresses organomercurial lyase (MerB) with the ppk-specified polyphosphate (polyP) and merT-encoding mercury transporter was constructed by integrating a bacterial merB gene into ppk/merT-transgenic tobacco. A large number of independent transgenic tobaccos was obtained, in some of which the merB gene was stably integrated in the plant genome and substantially translated to the expected MerB enzyme in the transgenic tobacco. The ppk/merT/merB-transgenic tobacco callus showed more resistance to methylmercury (CH3Hg+) and accumulated more mercury from CH3Hg+-containing medium than the ppk/merT-transgenic and wild-type progenitors. These results suggest that the MerB enzyme encoded by merB degraded the incorporated CH3Hg+ to Hg2+, which then accumulated as a less toxic Hg-polyP complex in the tobacco cells. Phytoremediation of CH3Hg+ and Hg2+ in the environment with this engineered ppk/merT/merB-transgenic plant, which prevents the release mercury vapor (Hg0) into the atmosphere in addition to generating potentially recyclable mercury-rich plant residues, is believed to be more acceptable to the public than other competing technologies, including phytovolatilization.

  13. Co-expression of NCED and ALO improves vitamin C level and tolerance to drought and chilling in transgenic tobacco and stylo plants.

    Science.gov (United States)

    Bao, Gegen; Zhuo, Chunliu; Qian, Chunmei; Xiao, Ting; Guo, Zhenfei; Lu, Shaoyun

    2016-01-01

    Abscisic acid (ABA) regulates plant adaptive responses to various environmental stresses, while L-ascorbic acid (AsA) that is also named vitamin C is an important antioxidant and involves in plant stress tolerance and the immune system in domestic animals. Transgenic tobacco (Nicotiana tabacum L.) and stylo [Stylosanthes guianensis (Aublet) Swartz], a forage legume, plants co-expressing stylo 9-cis-epoxycarotenoid dioxygenase (SgNCED1) and yeast D-arabinono-1,4-lactone oxidase (ALO) genes were generated in this study, and tolerance to drought and chilling was analysed in comparison with transgenic tobacco overexpressing SgNCED1 or ALO and the wild-type plants. Compared to the SgNCED1 or ALO transgenic plants, in which only ABA or AsA levels were increased, both ABA and AsA levels were increased in transgenic tobacco and stylo plants co-expressing SgNCED1 and ALO genes. Compared to the wild type, an enhanced drought tolerance was observed in SgNCED1 transgenic tobacco plants with induced expression of drought-responsive genes, but not in ALO plants, while an enhanced chilling tolerance was observed in ALO transgenic tobaccos with induced expression of cold-responsive genes, but not in SgNCED1 plants. Co-expression of SgNCED1 and ALO genes resulted in elevated tolerance to both drought and chilling in transgenic tobacco and stylo plants with induced expression of both drought and cold-responsive genes. Our result suggests that co-expression of SgNCED1 and ALO genes is an effective way for use in forage plant improvement for increased tolerance to drought and chilling and nutrition quality. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  14. Simultaneous Expression of PDH45 with EPSPS Gene Improves Salinity and Herbicide Tolerance in Transgenic Tobacco Plants.

    Science.gov (United States)

    Garg, Bharti; Gill, Sarvajeet S; Biswas, Dipul K; Sahoo, Ranjan K; Kunchge, Nandkumar S; Tuteja, Renu; Tuteja, Narendra

    2017-01-01

    To cope with the problem of salinity- and weed-induced crop losses, a multi-stress tolerant trait is need of the hour but a combinatorial view of such traits is not yet explored. The overexpression of PDH45 (pea DNA helicase 45) and EPSPS (5-enoylpruvyl shikimate-3-phosphate synthase) genes have been reported to impart salinity and herbicide tolerance. Further, the understanding of mechanism and pathways utilized by PDH45 and EPSPS for salinity and herbicide tolerance will help to improve the crops of economical importance. In the present study, we have performed a comparative analysis of salinity and herbicide tolerance to check the biochemical parameters and antioxidant status of tobacco transgenic plants. Collectively, the results showed that PDH45 overexpressing transgenic lines display efficient tolerance to salinity stress, while PDH45+EPSPS transgenics showed tolerance to both the salinity and herbicide as compared to the control [wild type (WT) and vector control (VC)] plants. The activities of the components of enzymatic antioxidant machinery were observed to be higher in the transgenic plants indicating the presence of an efficient antioxidant defense system which helps to cope with the stress-induced oxidative-damages. Photosynthetic parameters also showed significant increase in PDH45 and PDH45+EPSPS overexpressing transgenic plants in comparison to WT, VC and EPSPS transgenic plants under salinity stress. Furthermore, PDH45 and PDH45+EPSPS synergistically modulate the jasmonic acid and salicylic acid mediated signaling pathways for combating salinity stress. The findings of our study suggest that pyramiding of the PDH45 gene with EPSPS gene renders host plants tolerant to salinity and herbicide by enhancing the antioxidant machinery thus photosynthesis.

  15. Effect of ATP sulfurylase overexpression in bright yellow 2 tobacco cells: regulation of ATP sulfurylase and SO4(-2) transport activities

    International Nuclear Information System (INIS)

    Hatzfeld, Y.; Cathala, N.; Grignon, C.; Davidian, J.C.

    1998-01-01

    To determine if the ATP sulfurylase reaction is a regulatory step for the SO4(2-)-assimilation pathway in plants, an Arabidopsis thaliana ATP sulfurylase cDNA, APS2, was fused to the 355 promoter of the cauliflower mosaic virus and introduced by Agrobacterium tumefaciens-mediated transformation into isolated Bright Yellow 2 tobacco (Nicotiana tabacum) cells. The ATP sulfurylase activity in transgenic cells was 8-fold that in control cells, and was correlated with the expression of a specific polypeptide revealed by western analysis using an anti-ATP sulfurylase antibody. The molecular mass of this polypeptide agreed with that for the overexpressed mature protein. ATP sulfurylase overexpression had no effect on [35S]SO4(2-) influx or ATP sulfurylase activity regulation by S availability, except that ATP sulfurylase activity variations in response to S starvation in transgenic cells were 8 times higher than in the wild type. There were also no differences in cell growth or sensitivity to SeO4(2-) (a toxic SO4(2-) analog) between transgenic and wild-type cells. We propose that in Bright Yellow 2 tobacco cells, the ATP sulfurylase derepression by S deficiency may involve a posttranscriptional mechanism, and that the ATP sulfurylase abundance is not limiting for cell metabolism

  16. Transgenic overexpression of BAFF regulates the expression of ...

    Indian Academy of Sciences (India)

    To investigate whether transgenic overexpression of the zebrafish BAFF leads to ... and BAFF proteins were expressed separately and confirmed in HeLa cells. ... body homogenate of zebrafish and demonstrated a significant increase in ...

  17. Expression of a defence-related intercellular barley peroxidase in transgenic tobacco

    DEFF Research Database (Denmark)

    Kristensen, B.K.; Brandt, J.; Bojsen, K.

    1997-01-01

    genetically, phenotypically and biochemically. The T-DNA was steadily inherited through three generations. The barley peroxidase is expressed and sorted to the intercellular space in the transgenic tobacco plants. The peroxidase can be extracted from the intercellular space in two molecular forms from both...... barley and transgenic tobacco. The tobacco expressed forms are indistinguishable from the barley expressed forms as determined by analytical isoelectric focusing (pI 8.5) and Western-blotting. Staining for N-glycosylation showed that one form only was glycosylated. The N-terminus of purified Prx8 from...... transgenic tobacco was blocked by pyroglutamate, after the removal of which, N-terminal sequencing verified the transit signal-peptide cleavage site deduced from the cDNA sequence. Phenotype comparisons show that the constitutive expression of Prx8 lead to growth retardation. However, an infection assay...

  18. Accumulation of nickel in transgenic tobacco

    Science.gov (United States)

    Sidik, Nik Marzuki; Othman, Noor Farhan

    2013-11-01

    The accumulation of heavy metal Ni in the roots and leaves of four T1 transgenic lines of tobacco (T(1)20E, T(1)24C, T(1)18B1 and T(1)20B) expressing eiMT1 from E.indica was assessed. The aim of the study was to investigate the level of Ni accumulation in the leaves and roots of each transgenic lines and to evaluate the eligibility of the plants to be classified as a phytoremediation agent. All of the transgenic lines showed different ability in accumulating different metals and has translocation factor (TF) less than 1 (TFtransgenic lines, transgenic line T(1)24C showed the highest accumulation of Ni (251.9 ± 0.014 mg/kg) and the lowest TF value (TFT(1)24C=0.0875) at 60 ppm Ni.

  19. Regulation of galactolipid biosynthesis by overexpression of the rice MGD gene contributes to enhanced aluminum tolerance in tobacco

    Directory of Open Access Journals (Sweden)

    Meijuan eZhang

    2016-03-01

    Full Text Available Membrane lipid alterations affect Al tolerance in plants, but little is known about the regulation of membrane lipid metabolism in response to Al stress. Transgenic tobacco (Nicotiana tabacum overexpressing rice monogalactosyldiacylglycerol (MGDG synthase (OsMGD gene and wild-type tobacco plants were exposed to AlCl3, and the impact of Al toxicity on root growth, Al accumulation, plasma membrane integrity, lipid peroxidation and membrane lipid composition were investigated. Compared with the wild type, the transgenic plants exhibited rapid regrowth of roots after removal of Al and less damage to membrane integrity and lipid peroxidation under Al stress, meanwhile, the Al accumulation showed no difference between wild-type and transgenic plants. Lipid analysis showed that Al treatment dramatically decreased the content of MGDG and the ratio of MGDG to digalactosyldiacylglycerol (DGDG in wild-type plants, while it was unchanged in transgenic plants. The stable of MGDG level and the ratio of MGDG/DGDG contribute to maintain the membrane stability and permeability. Moreover, Al caused a significant increase in phospholipids in wild-type plants, resulting in a high proportion of phospholipids and low proportion of galactolipids, but these proportions were unaffected in transgenic plants. The high proportion of phospholipids could contribute to a higher rate of Al3+ binding in the membrane and thereby leads to more membrane perturbation and damage. These results show that the regulation of galactolipid biosynthesis could play an important role in maintaining membrane structure and function under Al stress.

  20. Characterisation of the nociceptive phenotype of suppressible galanin overexpressing transgenic mice

    Directory of Open Access Journals (Sweden)

    Wynick David

    2010-10-01

    Full Text Available Abstract The neuropeptide galanin is widely expressed in both the central and peripheral nervous systems and is involved in many diverse biological functions. There is a substantial data set that demonstrates galanin is upregulated after injury in the DRG, spinal cord and in many brain regions where it plays a predominantly antinociceptive role in addition to being neuroprotective and pro-regenerative. To further characterise the role of galanin following nerve injury, a novel transgenic line was created using the binary transgenic tet-off system, to overexpress galanin in galaninergic tissue in a suppressible manner. The double transgenic mice express significantly more galanin in the DRG one week after sciatic nerve section (axotomy compared to WT mice and this overexpression is suppressible upon administration of doxycycline. Phenotypic analysis revealed markedly attenuated allodynia when galanin is overexpressed and an increase in allodynia following galanin suppression. This novel transgenic line demonstrates that whether galanin expression is increased at the time of nerve injury or only after allodynia is established, the neuropeptide is able to reduce neuropathic pain behaviour. These new findings imply that administration of a galanin agonist to patients with established allodynia would be an effective treatment for neuropathic pain.

  1. Potential of MuS1 Transgenic Tobacco for Phytoremediation of the Urban Soils Contaminated with Cadmium

    Science.gov (United States)

    Kim, K. H.; Kim, Y. N.; Kim, S. H.

    2010-05-01

    Urban soils are prone to contamination by trace elements such as Cd, Cu, Pb and Zn. Phytoremediation is one of the attractive remediation methods for soils contaminated with trace elements due to its non-destructive and environmentally-friendly characteristic. Scientists have tried to find hyper-accumulator plants in nature or to develop transgenic plant through genetic engineering. This study was carried out to identify a potential of MuS1 transgenic tobacco for phytoremediation of the urban soils contaminated with Cd. MuS1 is known as a multiple stress related gene with several lines. The previous study using RT-PCR showed that the expression of MuS1 gene in tobacco plant induced tolerance to Cd stress. For this study, MuS1 transgenic tobacco and wild-type tobacco (control) were cultivated in a hydroponic system treated with Cd (0, 50, 100 and 200μM Cd) for 3 weeks. At harvest, both tobacco and nutrient solution were collected and were analyzed for Cd. Effect of Cd treatment on morphological change of the tobacco leaves was also observed by variable-pressure scanning electron microscopy (VP-SEM). The tolerance of MuS1 transgenic tobacco to Cd stress was better than that of wild-type tobacco at all Cd levels. Especially, wild-type tobacco showed chlorosis and withering with 200μM Cd treatment, whereas MuS1 transgenic tobacco gradually recovered from Cd damage. Wild-type tobacco accumulated more Cd (4.65mg per plant) than MuS1 transgenic tobacco (2.37mg per plant) with 200μM Cd treatment. Cd translocation rate from root to leaves was 81.8 % for wild-type tobacco compared to 37.1 % for MuS1 transgenic tobacco. Result of VP-SEM showed that the number of trichome in the leaves for wild-type tobacco increased in comparison with that for untreated samples after 3 weeks, while that for MuS1 transgenic tobacco was not changed by Cd treatment. Results showed that the mechanism of the recovery of the MuS1 tobacco plant was not by high level of Cd uptake and accumulation

  2. Brain phenotype of transgenic mice overexpressing cystathionine β-synthase.

    Directory of Open Access Journals (Sweden)

    Vinciane Régnier

    Full Text Available The cystathionine β-synthase (CBS gene, located on human chromosome 21q22.3, is a good candidate for playing a role in the Down Syndrome (DS cognitive profile: it is overexpressed in the brain of individuals with DS, and it encodes a key enzyme of sulfur-containing amino acid (SAA metabolism, a pathway important for several brain physiological processes.Here, we have studied the neural consequences of CBS overexpression in a transgenic mouse line (60.4P102D1 expressing the human CBS gene under the control of its endogenous regulatory regions. These mice displayed a ∼2-fold increase in total CBS proteins in different brain areas and a ∼1.3-fold increase in CBS activity in the cerebellum and the hippocampus. No major disturbance of SAA metabolism was observed, and the transgenic mice showed normal behavior in the rotarod and passive avoidance tests. However, we found that hippocampal synaptic plasticity is facilitated in the 60.4P102D1 line.We demonstrate that CBS overexpression has functional consequences on hippocampal neuronal networks. These results shed new light on the function of the CBS gene, and raise the interesting possibility that CBS overexpression might have an advantageous effect on some cognitive functions in DS.

  3. Brain phenotype of transgenic mice overexpressing cystathionine β-synthase.

    Science.gov (United States)

    Régnier, Vinciane; Billard, Jean-Marie; Gupta, Sapna; Potier, Brigitte; Woerner, Stéphanie; Paly, Evelyne; Ledru, Aurélie; David, Sabrina; Luilier, Sabrina; Bizot, Jean-Charles; Vacano, Guido; Kraus, Jan P; Patterson, David; Kruger, Warren D; Delabar, Jean M; London, Jaqueline

    2012-01-01

    The cystathionine β-synthase (CBS) gene, located on human chromosome 21q22.3, is a good candidate for playing a role in the Down Syndrome (DS) cognitive profile: it is overexpressed in the brain of individuals with DS, and it encodes a key enzyme of sulfur-containing amino acid (SAA) metabolism, a pathway important for several brain physiological processes. Here, we have studied the neural consequences of CBS overexpression in a transgenic mouse line (60.4P102D1) expressing the human CBS gene under the control of its endogenous regulatory regions. These mice displayed a ∼2-fold increase in total CBS proteins in different brain areas and a ∼1.3-fold increase in CBS activity in the cerebellum and the hippocampus. No major disturbance of SAA metabolism was observed, and the transgenic mice showed normal behavior in the rotarod and passive avoidance tests. However, we found that hippocampal synaptic plasticity is facilitated in the 60.4P102D1 line. We demonstrate that CBS overexpression has functional consequences on hippocampal neuronal networks. These results shed new light on the function of the CBS gene, and raise the interesting possibility that CBS overexpression might have an advantageous effect on some cognitive functions in DS.

  4. RNAi-mediated resistance to SMV and BYMV in transgenic tobacco

    Directory of Open Access Journals (Sweden)

    Lo Thi Mai Thu

    2016-09-01

    Full Text Available Soybean mosaic virus (SMV and bean yellow mosaic virus (BYMV are two typical types of viruses that cause mosaic in soybean plants. Multiple viral infections at the same site can lead to 66% to 80% yield reduction. We have aimed to improve SMV and BYMV resistance in Vietnamese soybeans using gene transfer techniques under the mechanism of RNAi. In this study, we present newly generated transgenic tobacco plants carrying RNAi [CPi (SMV-BYMV] resistance to the two types of viruses; 73.08% of transgenic tobacco lines proved to be fully resistant to SMV and BYMV. In addition, the number of virus copies in transgenic tobacco plants was reduced on average by more than 51% compared to the control plants (wild type. This promising result shows the potential of transerring the CPi (SMV-BYMV structure in soybean to increase resistance of soybean to SMV and BYMV and advance the aims of antiviral soybean breeding in Vietnam.

  5. Split-Cre complementation restores combination activity on transgene excision in hair roots of transgenic tobacco.

    Directory of Open Access Journals (Sweden)

    Mengling Wen

    Full Text Available The Cre/loxP system is increasingly exploited for genetic manipulation of DNA in vitro and in vivo. It was previously reported that inactive ''split-Cre'' fragments could restore Cre activity in transgenic mice when overlapping co-expression was controlled by two different promoters. In this study, we analyzed recombination activities of split-Cre proteins, and found that no recombinase activity was detected in the in vitro recombination reaction in which only the N-terminal domain (NCre of split-Cre protein was expressed, whereas recombination activity was obtained when the C-terminal (CCre or both NCre and CCre fragments were supplied. We have also determined the recombination efficiency of split-Cre proteins which were co-expressed in hair roots of transgenic tobacco. No Cre recombination event was observed in hair roots of transgenic tobacco when the NCre or CCre genes were expressed alone. In contrast, an efficient recombination event was found in transgenic hairy roots co-expressing both inactive split-Cre genes. Moreover, the restored recombination efficiency of split-Cre proteins fused with the nuclear localization sequence (NLS was higher than that of intact Cre in transgenic lines. Thus, DNA recombination mediated by split-Cre proteins provides an alternative method for spatial and temporal regulation of gene expression in transgenic plants.

  6. Hepatic steatosis in transgenic mice overexpressing human histone deacetylase 1

    International Nuclear Information System (INIS)

    Wang, Ai-Guo; Seo, Sang-Beom; Moon, Hyung-Bae; Shin, Hye-Jun; Kim, Dong Hoon; Kim, Jin-Man; Lee, Tae-Hoon; Kwon, Ho Jeong; Yu, Dae-Yeul; Lee, Dong-Seok

    2005-01-01

    It is generally thought that histone deacetylases (HDACs) play important roles in the transcriptional regulation of genes. However, little information is available concerning the specific functions of individual HDACs in disease states. In this study, two transgenic mice lines were established which harbored the human HDAC1 gene. Overexpressed HDAC1 was detected in the nuclei of transgenic liver cells, and HDAC1 enzymatic activity was significantly higher in the transgenic mice than in control littermates. The HDAC1 transgenic mice exhibited a high incidence of hepatic steatosis and nuclear pleomorphism. Molecular studies showed that HDAC1 may contribute to nuclear pleomorphism through the p53/p21 signaling pathway

  7. TaCIPK29, a CBL-interacting protein kinase gene from wheat, confers salt stress tolerance in transgenic tobacco.

    Directory of Open Access Journals (Sweden)

    Xiaomin Deng

    Full Text Available Calcineurin B-like protein-interacting protein kinases (CIPKs have been found to be responsive to abiotic stress. However, their precise functions and the related molecular mechanisms in abiotic stress tolerance are not completely understood, especially in wheat. In the present study, TaCIPK29 was identified as a new member of CIPK gene family in wheat. TaCIPK29 transcript increased after NaCl, cold, methyl viologen (MV, abscisic acid (ABA and ethylene treatments. Over-expression of TaCIPK29 in tobacco resulted in increased salt tolerance, which was demonstrated by higher germination rates, longer root lengths and better growth status of transgenic tobacco plants compared to controls when both were treated with salt stress. Physiological measurements indicated that transgenic tobacco seedlings retained high K(+/Na(+ ratios and Ca(2+ content by up-regulating some transporter genes expression and also possessed lower H2O2 levels and reduced membrane injury by increasing the expression and activities of catalase (CAT and peroxidase (POD under salt stress. Moreover, transgenic lines conferred tolerance to oxidative stress by increasing the activity and expression of CAT. Finally, TaCIPK29 was located throughout cells and it preferentially interacted with TaCBL2, TaCBL3, NtCBL2, NtCBL3 and NtCAT1. Taken together, our results showed that TaCIPK29 functions as a positive factor under salt stress and is involved in regulating cations and reactive oxygen species (ROS homeostasis.

  8. Pronounced Phenotypic Changes in Transgenic Tobacco Plants Overexpressing Sucrose Synthase May Reveal a Novel Sugar Signaling Pathway

    Science.gov (United States)

    Nguyen, Quynh Anh; Luan, Sheng; Wi, Seung G.; Bae, Hanhong; Lee, Dae-Seok; Bae, Hyeun-Jong

    2016-01-01

    Soluble sugars not only serve as nutrients, but also act as signals for plant growth and development, but how sugar signals are perceived and translated into physiological responses in plants remains unclear. We manipulated sugar levels in transgenic plants by overexpressing sucrose synthase (SuSy), which is a key enzyme believed to have reversible sucrose synthesis and sucrose degradation functions. The ectopically expressed SuSy protein exhibited sucrose-degrading activity, which may change the flux of sucrose demand from photosynthetic to non-photosynthetic cells, and trigger an unknown sucrose signaling pathway that lead to increased sucrose content in the transgenic plants. An experiment on the transition from heterotrophic to autotrophic growth demonstrated the existence of a novel sucrose signaling pathway, which stimulated photosynthesis, and enhanced photosynthetic synthesis of sucrose, which was the direct cause or the sucrose increase. In addition, a light/dark time treatment experiment, using different day length ranges for photosynthesis/respiration showed the carbohydrate pattern within a 24-h day and consolidated the role of sucrose signaling pathway as a way to maintain sucrose demand, and indicated the relationships between increased sucrose and upregulation of genes controlling development of the shoot apical meristem (SAM). As a result, transgenic plants featured a higher biomass and a shorter time required to switch to reproduction compared to those of control plants, indicating altered phylotaxis and more rapid advancement of developmental stages in the transgenic plants. PMID:26793204

  9. Transgenic tobacco overexpressing Brassica juncea HMG-CoA synthase 1 shows increased plant growth, pod size and seed yield.

    Directory of Open Access Journals (Sweden)

    Pan Liao

    Full Text Available Seeds are very important not only in the life cycle of the plant but they represent food sources for man and animals. We report herein a mutant of 3-hydroxy-3-methylglutaryl-coenzyme A synthase (HMGS, the second enzyme in the mevalonate (MVA pathway that can improve seed yield when overexpressed in a phylogenetically distant species. In Brassica juncea, the characterisation of four isogenes encoding HMGS has been previously reported. Enzyme kinetics on recombinant wild-type (wt and mutant BjHMGS1 had revealed that S359A displayed a 10-fold higher enzyme activity. The overexpression of wt and mutant (S359A BjHMGS1 in Arabidopsis had up-regulated several genes in sterol biosynthesis, increasing sterol content. To quickly assess the effects of BjHMGS1 overexpression in a phylogenetically more distant species beyond the Brassicaceae, wt and mutant (S359A BjHMGS1 were expressed in tobacco (Nicotiana tabacum L. cv. Xanthi of the family Solanaceae. New observations on tobacco OEs not previously reported for Arabidopsis OEs included: (i phenotypic changes in enhanced plant growth, pod size and seed yield (more significant in OE-S359A than OE-wtBjHMGS1 in comparison to vector-transformed tobacco, (ii higher NtSQS expression and sterol content in OE-S359A than OE-wtBjHMGS1 corresponding to greater increase in growth and seed yield, and (iii induction of NtIPPI2 and NtGGPPS2 and downregulation of NtIPPI1, NtGGPPS1, NtGGPPS3 and NtGGPPS4. Resembling Arabidopsis HMGS-OEs, tobacco HMGS-OEs displayed an enhanced expression of NtHMGR1, NtSMT1-2, NtSMT2-1, NtSMT2-2 and NtCYP85A1. Overall, increased growth, pod size and seed yield in tobacco HMGS-OEs were attributed to the up-regulation of native NtHMGR1, NtIPPI2, NtSQS, NtSMT1-2, NtSMT2-1, NtSMT2-2 and NtCYP85A1. Hence, S359A has potential in agriculture not only in improving phytosterol content but also seed yield, which may be desirable in food crops. This work further demonstrates HMGS function in plant

  10. Overexpression of a Medicago truncatula stress-associated protein gene (MtSAP1) leads to nitric oxide accumulation and confers osmotic and salt stress tolerance in transgenic tobacco.

    Science.gov (United States)

    Charrier, Aurélie; Planchet, Elisabeth; Cerveau, Delphine; Gimeno-Gilles, Christine; Verdu, Isabelle; Limami, Anis M; Lelièvre, Eric

    2012-08-01

    The impact of Medicago truncatula stress-associated protein gene (MtSAP1) overexpression has been investigated in Nicotiana tabacum transgenic seedlings. Under optimal conditions, transgenic lines overexpressing MtSAP1 revealed better plant development and higher chlorophyll content as compared to wild type seedlings. Interestingly, transgenic lines showed a stronger accumulation of nitric oxide (NO), a signaling molecule involved in growth and development processes. This NO production seemed to be partially nitrate reductase dependent. Due to the fact that NO has been also reported to play a role in tolerance acquisition of plants to abiotic stresses, the responses of MtSAP1 overexpressors to osmotic and salt stress have been studied. Compared to the wild type, transgenic lines were less affected in their growth and development. Moreover, NO content in MtSAP1 overexpressors was always higher than that detected in wild seedlings under stress conditions. It seems that this better tolerance induced by MtSAP1 overexpression could be associated with this higher NO production that would enable seedlings to reach a high protection level to prepare them to cope with abiotic stresses.

  11. Bone turnover is altered in transgenic rats overexpressing the P2Y2 purinergic receptor

    DEFF Research Database (Denmark)

    Ellegaard, Maria; Agca, Cansu; Petersen, Solveig

    2017-01-01

    overexpression on bone status and bone cell function using a transgenic rat. Three-month-old female transgenic Sprague Dawley rats overexpressing P2Y2R (P2Y2R-Tg) showed higher bone strength of the femoral neck. Histomorphometry showed increase in resorptive surfaces and reduction in mineralizing surfaces. Both...

  12. Protection of the photosynthetic apparatus from extreme dehydration and oxidative stress in seedlings of transgenic tobacco.

    Directory of Open Access Journals (Sweden)

    Concepción Almoguera

    Full Text Available A genetic program that in sunflower seeds is activated by Heat Shock transcription Factor A9 (HaHSFA9 has been analyzed in transgenic tobacco seedlings. The ectopic overexpression of the HSFA9 program protected photosynthetic membranes, which resisted extreme dehydration and oxidative stress conditions. In contrast, heat acclimation of seedlings induced thermotolerance but not resistance to the harsh stress conditions employed. The HSFA9 program was found to include the expression of plastidial small Heat Shock Proteins that accumulate only at lower abundance in heat-stressed vegetative organs. Photosystem II (PSII maximum quantum yield was higher for transgenic seedlings than for non-transgenic seedlings, after either stress treatment. Furthermore, protection of both PSII and Photosystem I (PSI membrane protein complexes was observed in the transgenic seedlings, leading to their survival after the stress treatments. It was also shown that the plastidial D1 protein, a labile component of the PSII reaction center, and the PSI core protein PsaB were shielded from oxidative damage and degradation. We infer that natural expression of the HSFA9 program during embryogenesis may protect seed pro-plastids from developmental desiccation.

  13. RNA-seq analysis of unintended effects in transgenic wheat overexpressing the transcription factor GmDREB1

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    Qiyan Jiang

    2017-06-01

    Full Text Available The engineering of plants with enhanced tolerance to abiotic stresses typically involves complex multigene networks and may therefore have a greater potential to introduce unintended effects than the genetic modification for simple monogenic traits. For this reason, it is essential to study the unintended effects in transgenic plants engineered for stress tolerance. We selected drought- and salt-tolerant transgenic wheat overexpressing the transcription factor, GmDREB1, to investigate unintended pleiotropic effects using RNA-seq analysis. We compared the transcriptome alteration of transgenic plants with that of wild-type plants subjected to salt stress as a control. We found that GmDREB1 overexpression had a minimal impact on gene expression under normal conditions. GmDREB1 overexpression resulted in transcriptional reprogramming of the salt response, but many of the genes with differential expression are known to mitigate salt stress and contribute incrementally to the enhanced stress tolerance of transgenic wheat. GmDREB1 overexpression did not activate unintended gene networks with respect to gene expression in the roots of transgenic wheat. This work is important for establishing a method of detecting unintended effects of genetic engineering and the safety of such traits with the development of marketable transgenic crops in the near future.

  14. Overexpression of snapdragon Delila (Del) gene in tobacco enhances anthocyanin accumulation and abiotic stress tolerance.

    Science.gov (United States)

    Naing, Aung Htay; Park, Kyeung Il; Ai, Trinh Ngoc; Chung, Mi Young; Han, Jeung Sul; Kang, Young-Wha; Lim, Ki Byung; Kim, Chang Kil

    2017-03-23

    Rosea1 (Ros1) and Delila (Del) co-expression controls anthocyanin accumulation in snapdragon flowers, while their overexpression in tomato strongly induces anthocyanin accumulation. However, little data exist on how Del expression alone influences anthocyanin accumulation. In tobacco (Nicotiana tabacum 'Xanthi'), Del expression enhanced leaf and flower anthocyanin production through regulating NtCHS, NtCHI, NtF3H, NtDFR, and NtANS transcript levels. Transgenic lines displayed different anthocyanin colors (e.g., pale red: T 0 -P, red: T 0 -R, and strong red: T 0 -S), resulting from varying levels of biosynthetic gene transcripts. Under salt stress, the T 2 generation had higher total polyphenol content, radical (DPPH, ABTS) scavenging activities, antioxidant-related gene expression, as well as overall greater salt and drought tolerance than wild type (WT). We propose that Del overexpression elevates transcript levels of anthocyanin biosynthetic and antioxidant-related genes, leading to enhanced anthocyanin production and antioxidant activity. The resultant increase of anthocyanin and antioxidant activity improves abiotic stress tolerance.

  15. The SbSOS1 gene from the extreme halophyte Salicornia brachiata enhances Na(+) loading in xylem and confers salt tolerance in transgenic tobacco.

    Science.gov (United States)

    Yadav, Narendra Singh; Shukla, Pushp Sheel; Jha, Anupama; Agarwal, Pradeep K; Jha, Bhavanath

    2012-10-11

    Soil salinity adversely affects plant growth and development and disturbs intracellular ion homeostasis resulting cellular toxicity. The Salt Overly Sensitive 1 (SOS1) gene encodes a plasma membrane Na(+)/H(+) antiporter that plays an important role in imparting salt stress tolerance to plants. Here, we report the cloning and characterisation of the SbSOS1 gene from Salicornia brachiata, an extreme halophyte. The SbSOS1 gene is 3774 bp long and encodes a protein of 1159 amino acids. SbSOS1 exhibited a greater level of constitutive expression in roots than in shoots and was further increased by salt stress. Overexpressing the S. brachiata SbSOS1 gene in tobacco conferred high salt tolerance, promoted seed germination and increased root length, shoot length, leaf area, fresh weight, dry weight, relative water content (RWC), chlorophyll, K(+)/Na(+) ratio, membrane stability index, soluble sugar, proline and amino acid content relative to wild type (WT) plants. Transgenic plants exhibited reductions in electrolyte leakage, reactive oxygen species (ROS) and MDA content in response to salt stress, which probably occurred because of reduced cytosolic Na(+) content and oxidative damage. At higher salt stress, transgenic tobacco plants exhibited reduced Na(+) content in root and leaf and higher concentrations in stem and xylem sap relative to WT, which suggests a role of SbSOS1 in Na(+) loading to xylem from root and leaf tissues. Transgenic lines also showed increased K(+) and Ca(2+) content in root tissue compared to WT, which reflect that SbSOS1 indirectly affects the other transporters activity. Overexpression of SbSOS1 in tobacco conferred a high degree of salt tolerance, enhanced plant growth and altered physiological and biochemical parameters in response to salt stress. In addition to Na(+) efflux outside the plasma membrane, SbSOS1 also helps to maintain variable Na(+) content in different organs and also affect the other transporters activity indirectly. These

  16. Production of transgenic pigs over-expressing the antiviral gene Mx1

    Directory of Open Access Journals (Sweden)

    Quanmei Yan

    2014-01-01

    Full Text Available The myxovirus resistance gene (Mx1 has a broad spectrum of antiviral activities. It is therefore an interesting candidate gene to improve disease resistance in farm animals. In this study, we report the use of somatic cell nuclear transfer (SCNT to produce transgenic pigs over-expressing the Mx1 gene. These transgenic pigs express approximately 15–25 times more Mx1 mRNA than non-transgenic pigs, and the protein level of Mx1 was also markedly enhanced. We challenged fibroblast cells isolated from the ear skin of transgenic and control pigs with influenza A virus and classical swine fever virus (CFSV. Indirect immunofluorescence assay (IFA revealed a profound decrease of influenza A proliferation in Mx1 transgenic cells. Growth kinetics showed an approximately 10-fold reduction of viral copies in the transgenic cells compared to non-transgenic controls. Additionally, we found that the Mx1 transgenic cells were more resistant to CSFV infection in comparison to non-transgenic cells. These results demonstrate that the Mx1 transgene can protect against viral infection in cells of transgenic pigs and indicate that the Mx1 transgene can be harnessed to develop disease-resistant pigs.

  17. Ectopic Expression of JcWRKY Confers Enhanced Resistance in Transgenic Tobacco Against Macrophomina phaseolina.

    Science.gov (United States)

    Agarwal, Parinita; Patel, Khantika; Agarwal, Pradeep K

    2018-04-01

    Plants possess an innate immune system comprising of a complex network of closely regulated defense responses involving differential gene expression mediated by transcription factors (TFs). The WRKYs comprise of an important plant-specific TF family, which is involved in regulation of biotic and abiotic defenses. The overexpression of JcWRKY resulted in improved resistance in transgenic tobacco against Macrophomina phaseolina. The production of reactive oxygen species (ROS) and its detoxification through antioxidative system in the transgenics facilitates defense against Macrophomina. The enhanced catalase activity on Macrophomina infection limits the spread of infection. The transcript expression of antioxidative enzymes gene (CAT and SOD) and salicylic acid (SA) biosynthetic gene ICS1 showed upregulation during Macrophomina infection and combinatorial stress. The enhanced transcript of pathogenesis-related genes PR-1 indicates the accumulation of SA during different stresses. The PR-2 and PR-5 highlight the activation of defense responses comprising of activation of hydrolytic cleavage of glucanases and thaumatin-like proteins causing disruption of fungal cells. The ROS homeostasis in coordination with signaling molecules regulate the defense responses and inhibit fungal growth.

  18. Overexpression of rice serotonin N-acetyltransferase 1 in transgenic rice plants confers resistance to cadmium and senescence and increases grain yield.

    Science.gov (United States)

    Lee, Kyungjin; Back, Kyoungwhan

    2017-04-01

    While ectopic overexpression of serotonin N-acetyltransferase (SNAT) in plants has been accomplished using animal SNAT genes, ectopic overexpression of plant SNAT genes in plants has not been investigated. Because the plant SNAT protein differs from that of animals in its subcellular localization and enzyme kinetics, its ectopic overexpression in plants would be expected to give outcomes distinct from those observed from overexpression of animal SNAT genes in transgenic plants. Consistent with our expectations, we found that transgenic rice plants overexpressing rice (Oryza sativa) SNAT1 (OsSNAT1) did not show enhanced seedling growth like that observed in ovine SNAT-overexpressing transgenic rice plants, although both types of plants exhibited increased melatonin levels. OsSNAT1-overexpressing rice plants did show significant resistance to cadmium and senescence stresses relative to wild-type controls. In contrast to tomato, melatonin synthesis in rice seedlings was not induced by selenium and OsSNAT1 transgenic rice plants did not show tolerance to selenium. T 2 homozygous OsSNAT1 transgenic rice plants exhibited increased grain yield due to increased panicle number per plant under paddy field conditions. These benefits conferred by ectopic overexpression of OsSNAT1 had not been observed in transgenic rice plants overexpressing ovine SNAT, suggesting that plant SNAT functions differently from animal SNAT in plants. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  19. Overexpression of the IbMYB1 gene in an orange-fleshed sweet potato cultivar produces a dual-pigmented transgenic sweet potato with improved antioxidant activity.

    Science.gov (United States)

    Park, Sung-Chul; Kim, Yun-Hee; Kim, Sun Ha; Jeong, Yu Jeong; Kim, Cha Young; Lee, Joon Seol; Bae, Ji-Yeong; Ahn, Mi-Jeong; Jeong, Jae Cheol; Lee, Haeng-Soon; Kwak, Sang-Soo

    2015-04-01

    The R2R3-type protein IbMYB1 is a key regulator of anthocyanin biosynthesis in the storage roots of sweet potato [Ipomoea batatas (L.) Lam]. Previously, we demonstrated that IbMYB1 expression stimulated anthocyanin pigmentation in tobacco leaves and Arabidopsis. Here, we generated dual-pigmented transgenic sweet potato plants that accumulated high levels of both anthocyanins and carotenoids in a single sweet potato storage root. An orange-fleshed cultivar with high carotenoid levels was transformed with the IbMYB1 gene under the control of either the storage root-specific sporamin 1 (SPO1) promoter or the oxidative stress-inducible peroxidase anionic 2 (SWPA2) promoter. The SPO1-MYB transgenic lines exhibited higher anthocyanin levels in storage roots than empty vector control (EV) or SWPA2-MYB plants, but carotenoid content was unchanged. SWPA2-MYB transgenic lines exhibited higher levels of both anthocyanin and carotenoids than EV plants. Analysis of hydrolyzed anthocyanin extracts indicated that cyanidin and peonidin predominated in both overexpression lines. Quantitative reverse transcription-polymerase chain reaction analysis demonstrated that IbMYB1 expression in both IbMYB1 transgenic lines strongly induced the upregulation of several genes in the anthocyanin biosynthetic pathway, whereas the expression of carotenoid biosynthetic pathway genes varied between transgenic lines. Increased anthocyanin levels in transgenic plants also promoted the elevation of proanthocyanidin and total phenolic levels in fresh storage roots. Consequently, all IbMYB1 transgenic plants displayed much higher antioxidant activities than EV plants. In field cultivations, storage root yields varied between the transgenic lines. Taken together, our results indicate that overexpression of IbMYB1 is a highly promising strategy for the generation of transgenic plants with enhanced antioxidant capacity. © 2014 Scandinavian Plant Physiology Society.

  20. Overexpression of a Grapevine Sucrose Transporter (VvSUC27 in Tobacco Improves Plant Growth Rate in the Presence of Sucrose In vitro

    Directory of Open Access Journals (Sweden)

    Yumeng Cai

    2017-06-01

    Full Text Available The import of sugar from source leaves and it further accumulation in grape berries are considerably high during ripening, and this process is mediated via sucrose transporters. In this study, a grape sucrose transporter (SUT gene, VvSUC27, located at the plasma membrane, was transferred to tobacco (Nicotiana tabacum. The transformants were more sensitive to sucrose and showed more rapid development, especially roots, when cultured on MS agar medium containing sucrose, considering that the shoot/root dry weight ratio was only half that of the control. Moreover, all transformed plants exhibited light-colored leaves throughout their development, which indicated chlorosis and an associated reduction in photosynthesis. The total sugar content in the roots and stems of transformants was higher than that in control plants. No significant difference was observed in the leaves between the transformants and control plants. The levels of growth-promoting hormones were increased, and those of stress-mediating hormones were reduced in transgenic tobacco plants. The qRT-PCR analysis revealed that the expression of VvSUC27 was 1,000 times higher than that of the autologous tobacco sucrose transporter, which suggested that the markedly increased growth rate of transformants was because of the heterogeneously expressed gene. The transgenic tobacco plants showed resistance to abiotic stresses. Strikingly, the overexpression of VvSUC27 leaded to the up regulation of most reactive oxygen species scavengers and abscisic acid-related genes that might enable transgenic plants to overcome abiotic stress. Taken together, these results revealed an important role of VvSUC27 in plant growth and response to abiotic stresses, especially in the presence of sucrose in vitro.

  1. Cadmium resistance in tobacco plants expressing the MuSI gene.

    Science.gov (United States)

    Kim, Young-Nam; Kim, Ji-Seoung; Seo, Sang-Gyu; Lee, Youngwoo; Baek, Seung-Woo; Kim, Il-Sup; Yoon, Ho-Sung; Kim, Kwon-Rae; Kim, Sun-Hyung; Kim, Kye-Hoon

    2011-10-01

    MuSI, a gene that corresponds to a domain that contains the rubber elongation factor (REF), is highly homologous to many stress-related proteins in plants. Since MuSI is up-regulated in the roots of plants treated with cadmium or copper, the involvement of MuSI in cadmium tolerance was investigated in this study. Escherichia coli cells overexpressing MuSI were more resistant to Cd than wild-type cells transfected with vector alone. MuSI transgenic plants were also more resistant to Cd. MuSI transgenic tobacco plants absorbed less Cd than wild-type plants. Cd translocation from roots to shoots was reduced in the transgenic plants, thereby avoiding Cd toxicity. The number of short trichomes in the leaves of wild-type tobacco plants was increased by Cd treatment, while this was unchanged in MuSI transgenic tobacco. These results suggest that MuSI transgenic tobacco plants have enhanced tolerance to Cd via reduced Cd uptake and/or increased Cd immobilization in the roots, resulting in less Cd translocation to the shoots.

  2. Expression of chimeric HCV peptide in transgenic tobacco plants ...

    African Journals Online (AJOL)

    Expression of chimeric HCV peptide in transgenic tobacco plants infected with recombinant alfalfa mosaic virus for development of a plant-derived vaccine against HCV. AK El Attar, AM Shamloul, AA Shalaby, BY Riad, A Saad, HM Mazyad, JM Keith ...

  3. Overexpression of cotton RAV1 gene in Arabidopsis confers transgenic plants high salinity and drought sensitivity.

    Science.gov (United States)

    Li, Xiao-Jie; Li, Mo; Zhou, Ying; Hu, Shan; Hu, Rong; Chen, Yun; Li, Xue-Bao

    2015-01-01

    RAV (related to ABI3/VP1) protein containing an AP2 domain in the N-terminal region and a B3 domain in the C-terminal region, which belongs to AP2 transcription factor family, is unique in higher plants. In this study, a gene (GhRAV1) encoding a RAV protein of 357 amino acids was identified in cotton (Gossypium hirsutum). Transient expression analysis of the eGFP:GhRAV1 fusion genes in tobacco (Nicotiana tabacum) epidermal cells revealed that GhRAV1 protein was localized in the cell nucleus. Quantitative RT-PCR analysis indicated that expression of GhRAV1 in cotton is induced by abscisic acid (ABA), NaCl and polyethylene glycol (PEG). Overexpression of GhRAV1 in Arabidopsis resulted in plant sensitive to ABA, NaCl and PEG. With abscisic acid (ABA) treatment, seed germination and green seedling rates of the GhRAV1 transgenic plants were remarkably lower than those of wild type. In the presence of NaCl, the seed germination and seedling growth of the GhRAV1 transgenic lines were inhibited greater than those of wild type. And chlorophyll content and maximum photochemical efficiency of the transgenic plants were significantly lower than those of wild type. Under drought stress, the GhRAV1 transgenic plants displayed more severe wilting than wild type. Furthermore, expressions of the stress-related genes were altered in the GhRAV1 transgenic Arabidopsis plants under high salinity and drought stresses. Collectively, our data suggested that GhRAV1 may be involved in response to high salinity and drought stresses through regulating expressions of the stress-related genes during cotton development.

  4. Transcriptome response signatures associated with the overexpression of a mitochondrial uncoupling protein (AtUCP1 in tobacco.

    Directory of Open Access Journals (Sweden)

    Alessandra Vasconcellos Nunes Laitz

    Full Text Available Mitochondrial inner membrane uncoupling proteins (UCP dissipate the proton electrochemical gradient established by the respiratory chain, thus affecting the yield of ATP synthesis. UCP overexpression in plants has been correlated with oxidative stress tolerance, improved photosynthetic efficiency and increased mitochondrial biogenesis. This study reports the main transcriptomic responses associated with the overexpression of an UCP (AtUCP1 in tobacco seedlings. Compared to wild-type (WT, AtUCP1 transgenic seedlings showed unaltered ATP levels and higher accumulation of serine. By using RNA-sequencing, a total of 816 differentially expressed genes between the investigated overexpressor lines and the untransformed WT control were identified. Among them, 239 were up-regulated and 577 were down-regulated. As a general response to AtUCP1 overexpression, noticeable changes in the expression of genes involved in energy metabolism and redox homeostasis were detected. A substantial set of differentially expressed genes code for products targeted to the chloroplast and mainly involved in photosynthesis. The overall results demonstrate that the alterations in mitochondrial function provoked by AtUCP1 overexpression require important transcriptomic adjustments to maintain cell homeostasis. Moreover, the occurrence of an important cross-talk between chloroplast and mitochondria, which culminates in the transcriptional regulation of several genes involved in different pathways, was evidenced.

  5. Stress-inducible expression of an F-box gene TaFBA1 from wheat enhanced the drought tolerance in transgenic tobacco plants without impacting growth and development

    Directory of Open Access Journals (Sweden)

    Xiangzhu Kong

    2016-09-01

    Full Text Available E3 ligase plays an important role in the response to many environment stresses in plants. In our previous study, constitutive overexpression of an F-box protein gene TaFBA1 driven by 35S promoter improved the drought tolerance in transgenic tobacco plants, but the growth and development in transgenic plants was altered in normal conditions. In this study, we used stress-inducible promoter RD29A instead of 35S promoter, as a results, the stress-inducible transgenic tobacco plants exhibit a similar phenotype with WT plants. However, the drought tolerance of the transgenic plants with stress-inducible expressed TaFBA1 was enhanced. The improved drought tolerance of transgenic plants was indicated by their higher seed germination rate and survival rate, greater biomass and photosynthesis than those of WT under water stress, which may be related to their greater water retention capability and osmotic adjustment. Moreover, the transgenic plants accumulated less reactive oxygen species (ROS, kept lower MDA content and membrane leakage under water stress, which may be related to their higher levels of antioxidant enzyme activity and upregulated gene expression of some antioxidant enzymes. These results suggest that stress induced expression of TaFBA1 confers drought tolerance via the improved water retention and antioxidative compete abilibty. Meanwhile, this stress-inducible expression strategy by RD29A promoter can minimize the unexpectable effects by 35S constitutive promoter on phenotypes of the transgenic plants.

  6. Stress-Inducible Expression of an F-box Gene TaFBA1 from Wheat Enhanced the Drought Tolerance in Transgenic Tobacco Plants without Impacting Growth and Development.

    Science.gov (United States)

    Kong, Xiangzhu; Zhou, Shumei; Yin, Suhong; Zhao, Zhongxian; Han, Yangyang; Wang, Wei

    2016-01-01

    E3 ligase plays an important role in the response to many environment stresses in plants. In our previous study, constitutive overexpression of an F-box protein gene TaFBA1 driven by 35S promoter improved the drought tolerance in transgenic tobacco plants, but the growth and development in transgenic plants was altered in normal conditions. In this study, we used stress-inducible promoter RD29A instead of 35S promoter, as a results, the stress-inducible transgenic tobacco plants exhibit a similar phenotype with wild type (WT) plants. However, the drought tolerance of the transgenic plants with stress-inducible expressed TaFBA1 was enhanced. The improved drought tolerance of transgenic plants was indicated by their higher seed germination rate and survival rate, greater biomass and photosynthesis than those of WT under water stress, which may be related to their greater water retention capability and osmotic adjustment. Moreover, the transgenic plants accumulated less reactive oxygen species, kept lower MDA content and membrane leakage under water stress, which may be related to their higher levels of antioxidant enzyme activity and upregulated gene expression of some antioxidant enzymes. These results suggest that stress induced expression of TaFBA1 confers drought tolerance via the improved water retention and antioxidative compete ability. Meanwhile, this stress-inducible expression strategy by RD29A promoter can minimize the unexpectable effects by 35S constitutive promoter on phenotypes of the transgenic plants.

  7. Overexpression of EcbHLH57 Transcription Factor from Eleusine coracana L. in Tobacco Confers Tolerance to Salt, Oxidative and Drought Stress.

    Directory of Open Access Journals (Sweden)

    K C Babitha

    Full Text Available Basic helix-loop-helix (bHLH transcription factors constitute one of the largest families in plants and are known to be involved in various developmental processes and stress tolerance. We report the characterization of a stress responsive bHLH transcription factor from stress adapted species finger millet which is homologous to OsbHLH57 and designated as EcbHLH57. The full length sequence of EcbHLH57 consisted of 256 amino acids with a conserved bHLH domain followed by leucine repeats. In finger millet, EcbHLH57 transcripts were induced by ABA, NaCl, PEG, methyl viologen (MV treatments and drought stress. Overexpression of EcbHLH57 in tobacco significantly increased the tolerance to salinity and drought stress with improved root growth. Transgenic plants showed higher photosynthetic rate and stomatal conductance under drought stress that resulted in higher biomass. Under long-term salinity stress, the transgenic plants accumulated higher seed weight/pod and pod number. The transgenic plants were also tolerant to oxidative stress and showed less accumulation of H202 and MDA levels. The overexpression of EcbHLH57 enhanced the expression of stress responsive genes such as LEA14, rd29A, rd29B, SOD, APX, ADH1, HSP70 and also PP2C and hence improved tolerance to diverse stresses.

  8. The SbSOS1 gene from the extreme halophyte Salicornia brachiata enhances Na+ loading in xylem and confers salt tolerance in transgenic tobacco

    Directory of Open Access Journals (Sweden)

    Yadav Narendra

    2012-10-01

    Full Text Available Abstract Background Soil salinity adversely affects plant growth and development and disturbs intracellular ion homeostasis resulting cellular toxicity. The Salt Overly Sensitive 1 (SOS1 gene encodes a plasma membrane Na+/H+ antiporter that plays an important role in imparting salt stress tolerance to plants. Here, we report the cloning and characterisation of the SbSOS1 gene from Salicornia brachiata, an extreme halophyte. Results The SbSOS1 gene is 3774 bp long and encodes a protein of 1159 amino acids. SbSOS1 exhibited a greater level of constitutive expression in roots than in shoots and was further increased by salt stress. Overexpressing the S. brachiata SbSOS1 gene in tobacco conferred high salt tolerance, promoted seed germination and increased root length, shoot length, leaf area, fresh weight, dry weight, relative water content (RWC, chlorophyll, K+/Na+ ratio, membrane stability index, soluble sugar, proline and amino acid content relative to wild type (WT plants. Transgenic plants exhibited reductions in electrolyte leakage, reactive oxygen species (ROS and MDA content in response to salt stress, which probably occurred because of reduced cytosolic Na+ content and oxidative damage. At higher salt stress, transgenic tobacco plants exhibited reduced Na+ content in root and leaf and higher concentrations in stem and xylem sap relative to WT, which suggests a role of SbSOS1 in Na+ loading to xylem from root and leaf tissues. Transgenic lines also showed increased K+ and Ca2+ content in root tissue compared to WT, which reflect that SbSOS1 indirectly affects the other transporters activity. Conclusions Overexpression of SbSOS1 in tobacco conferred a high degree of salt tolerance, enhanced plant growth and altered physiological and biochemical parameters in response to salt stress. In addition to Na+ efflux outside the plasma membrane, SbSOS1 also helps to maintain variable Na+ content in different organs and also affect the other

  9. The SbSOS1 gene from the extreme halophyte Salicornia brachiata enhances Na+ loading in xylem and confers salt tolerance in transgenic tobacco

    Science.gov (United States)

    2012-01-01

    Background Soil salinity adversely affects plant growth and development and disturbs intracellular ion homeostasis resulting cellular toxicity. The Salt Overly Sensitive 1 (SOS1) gene encodes a plasma membrane Na+/H+ antiporter that plays an important role in imparting salt stress tolerance to plants. Here, we report the cloning and characterisation of the SbSOS1 gene from Salicornia brachiata, an extreme halophyte. Results The SbSOS1 gene is 3774 bp long and encodes a protein of 1159 amino acids. SbSOS1 exhibited a greater level of constitutive expression in roots than in shoots and was further increased by salt stress. Overexpressing the S. brachiata SbSOS1 gene in tobacco conferred high salt tolerance, promoted seed germination and increased root length, shoot length, leaf area, fresh weight, dry weight, relative water content (RWC), chlorophyll, K+/Na+ ratio, membrane stability index, soluble sugar, proline and amino acid content relative to wild type (WT) plants. Transgenic plants exhibited reductions in electrolyte leakage, reactive oxygen species (ROS) and MDA content in response to salt stress, which probably occurred because of reduced cytosolic Na+ content and oxidative damage. At higher salt stress, transgenic tobacco plants exhibited reduced Na+ content in root and leaf and higher concentrations in stem and xylem sap relative to WT, which suggests a role of SbSOS1 in Na+ loading to xylem from root and leaf tissues. Transgenic lines also showed increased K+ and Ca2+ content in root tissue compared to WT, which reflect that SbSOS1 indirectly affects the other transporters activity. Conclusions Overexpression of SbSOS1 in tobacco conferred a high degree of salt tolerance, enhanced plant growth and altered physiological and biochemical parameters in response to salt stress. In addition to Na+ efflux outside the plasma membrane, SbSOS1 also helps to maintain variable Na+ content in different organs and also affect the other transporters activity indirectly

  10. Overexpressing Exogenous 5-Enolpyruvylshikimate-3-Phosphate Synthase (EPSPS Genes Increases Fecundity and Auxin Content of Transgenic Arabidopsis Plants

    Directory of Open Access Journals (Sweden)

    Jia Fang

    2018-02-01

    Full Text Available Transgenic glyphosate-tolerant plants overproducing EPSPS (5-enolpyruvylshikimate-3-phosphate synthase may exhibit enhanced fitness in glyphosate-free environments. If so, introgression of transgenes overexpressing EPSPS into wild relative species may lead to increased competitiveness of crop-wild hybrids, resulting in unpredicted environmental impact. Assessing fitness effects of transgenes overexpressing EPSPS in a model plant species can help address this question, while elucidating how overproducing EPSPS affects the fitness-related traits of plants. We produced segregating T2 and T3Arabidopsis thaliana lineages with or without a transgene overexpressing EPSPS isolated from rice or Agrobacterium (CP4. For each of the three transgenes, we compared glyphosate tolerance, some fitness-related traits, and auxin (indole-3-acetic acid content in transgene-present, transgene-absent, empty vector (EV, and parental lineages in a common-garden experiment. We detected substantially increased glyphosate tolerance in T2 plants of transgene-present lineages that overproduced EPSPS. We also documented significant increases in fecundity, which was associated with increased auxin content in T3 transgene-present lineages containing rice EPSPS genes, compared with their segregating transgene-absent lineages, EV, and parental controls. Our results from Arabidopsis with nine transgenic events provide a strong support to the hypothesis that transgenic plants overproducing EPSPS can benefit from a fecundity advantage in glyphosate-free environments. Stimulated biosynthesis of auxin, an important plant growth hormone, by overproducing EPSPS may play a role in enhanced fecundity of the transgenic Arabidopsis plants. The obtained knowledge is useful for assessing environmental impact caused by introgression of transgenes overproducing EPSPS from any GE crop into populations of its wild relatives.

  11. Overexpressing Exogenous 5-Enolpyruvylshikimate-3-Phosphate Synthase (EPSPS) Genes Increases Fecundity and Auxin Content of Transgenic Arabidopsis Plants.

    Science.gov (United States)

    Fang, Jia; Nan, Peng; Gu, Zongying; Ge, Xiaochun; Feng, Yu-Qi; Lu, Bao-Rong

    2018-01-01

    Transgenic glyphosate-tolerant plants overproducing EPSPS (5-enolpyruvylshikimate-3-phosphate synthase) may exhibit enhanced fitness in glyphosate-free environments. If so, introgression of transgenes overexpressing EPSPS into wild relative species may lead to increased competitiveness of crop-wild hybrids, resulting in unpredicted environmental impact. Assessing fitness effects of transgenes overexpressing EPSPS in a model plant species can help address this question, while elucidating how overproducing EPSPS affects the fitness-related traits of plants. We produced segregating T 2 and T 3 Arabidopsis thaliana lineages with or without a transgene overexpressing EPSPS isolated from rice or Agrobacterium ( CP4 ). For each of the three transgenes, we compared glyphosate tolerance, some fitness-related traits, and auxin (indole-3-acetic acid) content in transgene-present, transgene-absent, empty vector (EV), and parental lineages in a common-garden experiment. We detected substantially increased glyphosate tolerance in T 2 plants of transgene-present lineages that overproduced EPSPS. We also documented significant increases in fecundity, which was associated with increased auxin content in T 3 transgene-present lineages containing rice EPSPS genes, compared with their segregating transgene-absent lineages, EV, and parental controls. Our results from Arabidopsis with nine transgenic events provide a strong support to the hypothesis that transgenic plants overproducing EPSPS can benefit from a fecundity advantage in glyphosate-free environments. Stimulated biosynthesis of auxin, an important plant growth hormone, by overproducing EPSPS may play a role in enhanced fecundity of the transgenic Arabidopsis plants. The obtained knowledge is useful for assessing environmental impact caused by introgression of transgenes overproducing EPSPS from any GE crop into populations of its wild relatives.

  12. Overexpression of SoCYP85A1, a Spinach Cytochrome p450 Gene in Transgenic Tobacco Enhances Root Development and Drought Stress Tolerance

    Directory of Open Access Journals (Sweden)

    Fangmeng Duan

    2017-11-01

    Full Text Available Brassinosteroids (BRs play an essential role in plant growth, development, and responses to diverse abiotic stresses. However, previous studies mainly analyzed how exogenous BRs influenced plant physiological reactions to drought stress, therefore, genetic evidences for the endogenous BRs-mediated regulation of plant responses still remain elusive. In this study, a key BRs biosynthetic gene, SoCYP85A1 was cloned from Spinacia oleracea, which has a complete open reading frame of 1,392 bp encoding a 464 amino acid peptide and shares high sequence similarities with CYP85A1 from other plants. The expression of SoCYP85A1 which was higher in leaf compared with root and stem, was induced by treatments of PEG6000, abscisic acid (ABA, low temperature and high salt. Increases in both SoCYP85A1 transcripts and endogenous BRs in transgenic tobacco which resulted in longer primary root and more lateral roots enhanced drought tolerance compared with wild types. The transgenic tobacco accumulated much lower levels of reactive oxygen species and malondialdehyde (MDA than wild types did, accompanied by significantly higher content of proline and notably enhanced activities of antioxidant enzymes. Besides, transcriptional expressions of six stress-responsive genes were regulated to higher levels in transgenic lines under drought stress. Taken together, our results demonstrated that SoCYP85A1 involves in response to drought stress by promoting root development, scavenging ROS, and regulating expressions of stress-responsive genes.

  13. The SbASR-1 gene cloned from an extreme halophyte Salicornia brachiata enhances salt tolerance in transgenic tobacco.

    Science.gov (United States)

    Jha, Bhavanath; Lal, Sanjay; Tiwari, Vivekanand; Yadav, Sweta Kumari; Agarwal, Pradeep K

    2012-12-01

    Salinity severely affects plant growth and development. Plants evolved various mechanisms to cope up stress both at molecular and cellular levels. Halophytes have developed better mechanism to alleviate the salt stress than glycophytes, and therefore, it is advantageous to study the role of different genes from halophytes. Salicornia brachiata is an extreme halophyte, which grows luxuriantly in the salty marshes in the coastal areas. Earlier, we have isolated SbASR-1 (abscisic acid stress ripening-1) gene from S. brachiata using cDNA subtractive hybridisation library. ASR-1 genes are abscisic acid (ABA) responsive, whose expression level increases under abiotic stresses, injury, during fruit ripening and in pollen grains. The SbASR-1 transcript showed up-regulation under salt stress conditions. The SbASR-1 protein contains 202 amino acids of 21.01-kDa molecular mass and has 79 amino acid long signatures of ABA/WDS gene family. It has a maximum identity (73 %) with Solanum chilense ASR-1 protein. The SbASR-1 has a large number of disorder-promoting amino acids, which make it an intrinsically disordered protein. The SbASR-1 gene was over-expressed under CaMV 35S promoter in tobacco plant to study its physiological functions under salt stress. T(0) transgenic tobacco seeds showed better germination and seedling growth as compared to wild type (Wt) in a salt stress condition. In the leaf tissues of transgenic lines, Na(+) and proline contents were significantly lower, as compared to Wt plant, under salt treatment, suggesting that transgenic plants are better adapted to salt stress.

  14. A dominant-negative mutation within AtMYB90 blocks flower pigment production in transgenic tobacco.

    Science.gov (United States)

    During de novo shoot induction in cultured transgenic tobacco callus a spontaneous mutation within the coding region of a AtMYB90 transgene produced a plant line in which the original transgene-induced over-pigmented phenotype (dark red/purple from anthocyanin overproduction in most tissues) was los...

  15. Overexpression AtNHX1 confers salt-tolerance of transgenic tall ...

    African Journals Online (AJOL)

    Saline soil is a serious problem worldwide, and it is necessary to improve the salt tolerance of plants so as to avoid the progressive deterioration of saline soil. Here we report that over-expression of AtNHX1 improves salt tolerance in transgenic tall fescue. The AtNHX1 gene driven with CaMV35S promoter was constructed ...

  16. Enhanced whitefly resistance in transgenic tobacco plants expressing double stranded RNA of v-ATPase A gene.

    Science.gov (United States)

    Thakur, Nidhi; Upadhyay, Santosh Kumar; Verma, Praveen C; Chandrashekar, Krishnappa; Tuli, Rakesh; Singh, Pradhyumna K

    2014-01-01

    Expression of double strand RNA (dsRNA) designed against important insect genes in transgenic plants have been shown to give protection against pests through RNA interference (RNAi), thus opening the way for a new generation of insect-resistant crops. We have earlier compared the efficacy of dsRNAs/siRNAs, against a number of target genes, for interference in growth of whitefly (Bemisia tabaci) upon oral feeding. The v-ATPase subunit A (v-ATPaseA) coding gene was identified as a crucial target. We now report the effectiveness of transgenic tobacco plants expressing siRNA to silence v-ATPaseA gene expression for the control of whitefly infestation. Transgenic tobacco lines were developed for the expression of long dsRNA precursor to make siRNA and knock down the v-ATPaseA mRNA in whitefly. Molecular analysis and insecticidal properties of the transgenic plants established the formation of siRNA targeting the whitefly v-ATPaseA, in the leaves. The transcript level of v-ATPaseA in whiteflies was reduced up to 62% after feeding on the transgenic plants. Heavy infestation of whiteflies on the control plants caused significant loss of sugar content which led to the drooping of leaves. The transgenic plants did not show drooping effect. Host plant derived pest resistance was achieved against whiteflies by genetic transformation of tobacco which generated siRNA against the whitefly v-ATPaseA gene. Transgenic tobacco lines expressing dsRNA of v-ATPaseA, delivered sufficient siRNA to whiteflies feeding on them, mounting a significant silencing response, leading to their mortality. The transcript level of the target gene was reduced in whiteflies feeding on transgenic plants. The strategy can be taken up for genetic engineering of plants to control whiteflies in field crops.

  17. Enhanced whitefly resistance in transgenic tobacco plants expressing double stranded RNA of v-ATPase A gene.

    Directory of Open Access Journals (Sweden)

    Nidhi Thakur

    Full Text Available BACKGROUND: Expression of double strand RNA (dsRNA designed against important insect genes in transgenic plants have been shown to give protection against pests through RNA interference (RNAi, thus opening the way for a new generation of insect-resistant crops. We have earlier compared the efficacy of dsRNAs/siRNAs, against a number of target genes, for interference in growth of whitefly (Bemisia tabaci upon oral feeding. The v-ATPase subunit A (v-ATPaseA coding gene was identified as a crucial target. We now report the effectiveness of transgenic tobacco plants expressing siRNA to silence v-ATPaseA gene expression for the control of whitefly infestation. METHODOLOGY/PRINCIPAL FINDINGS: Transgenic tobacco lines were developed for the expression of long dsRNA precursor to make siRNA and knock down the v-ATPaseA mRNA in whitefly. Molecular analysis and insecticidal properties of the transgenic plants established the formation of siRNA targeting the whitefly v-ATPaseA, in the leaves. The transcript level of v-ATPaseA in whiteflies was reduced up to 62% after feeding on the transgenic plants. Heavy infestation of whiteflies on the control plants caused significant loss of sugar content which led to the drooping of leaves. The transgenic plants did not show drooping effect. CONCLUSIONS/SIGNIFICANCE: Host plant derived pest resistance was achieved against whiteflies by genetic transformation of tobacco which generated siRNA against the whitefly v-ATPaseA gene. Transgenic tobacco lines expressing dsRNA of v-ATPaseA, delivered sufficient siRNA to whiteflies feeding on them, mounting a significant silencing response, leading to their mortality. The transcript level of the target gene was reduced in whiteflies feeding on transgenic plants. The strategy can be taken up for genetic engineering of plants to control whiteflies in field crops.

  18. Transgenic plants over-expressing insect-specific microRNA acquire insecticidal activity against Helicoverpa armigera: an alternative to Bt-toxin technology.

    Science.gov (United States)

    Agrawal, Aditi; Rajamani, Vijayalakshmi; Reddy, Vanga Siva; Mukherjee, Sunil Kumar; Bhatnagar, Raj K

    2015-10-01

    The success of Bt transgenics in controlling predation of crops has been tempered by sporadic emergence of resistance in targeted insect larvae. Such emerging threats have prompted the search for novel insecticidal molecules that are specific and could be expressed through plants. We have resorted to small RNA-based technology for an investigative search and focused our attention to an insect-specific miRNA that interferes with the insect molting process resulting in the death of the larvae. In this study, we report the designing of a vector that produces artificial microRNA (amiR), namely amiR-24, which targets the chitinase gene of Helicoverpa armigera. This vector was used as transgene in tobacco. Northern blot and real-time analysis revealed the high level expression of amiR-24 in transgenic tobacco plants. Larvae feeding on the transgenic plants ceased to molt further and eventually died. Our results demonstrate that transgenic tobacco plants can express amiR-24 insectice specific to H. armigera.

  19. A cold-induced myo-inositol transporter-like gene confers tolerance to multiple abiotic stresses in transgenic tobacco plants.

    Science.gov (United States)

    Sambe, Mame Abdou Nahr; He, Xueying; Tu, Qinghua; Guo, Zhenfei

    2015-03-01

    A full length cDNA encoding a myo-inositol transporter-like protein, named as MfINT-like, was cloned from Medicago sativa subsp. falcata (herein falcata), a species with greater cold tolerance than alfalfa (M. sativa subsp. sativa). MfINT-like is located on plasma membranes. MfINT-like transcript was induced 2-4 h after exogenous myo-inositol treatment, 24-96 h with cold, and 96 h by salinity. Given that myo-inositol accumulates higher in falcata after 24 h of cold treatment, myo-inositol is proposed to be involved in cold-induced expression of MfINT-like. Higher levels of myo-inositol was observed in leaves of transgenic tobacco plants overexpressing MfINT-like than the wild-type but not in the roots of plants grown on myo-inositol containing medium, suggesting that transgenic plants had higher myo-inositol transport activity than the wild-type. Transgenic plants survived better to freezing temperature, and had lower ion leakage and higher maximal photochemical efficiency of photosystem II (Fv /Fm ) after chilling treatment. In addition, greater plant fresh weight was observed in transgenic plants as compared with the wild-type when plants were grown under drought or salinity stress. The results suggest that MfINT-like mediated transport of myo-inositol is associated with plant tolerance to abiotic stresses. © 2014 Scandinavian Plant Physiology Society.

  20. Trichostatin A suppresses lung adenocarcinoma development in Grg1 overexpressing transgenic mice

    International Nuclear Information System (INIS)

    Liu, Ju; Li, Yan; Dong, Fengyun; Li, Liqun; Masuda, Takahiro; Allen, Thaddeus D.; Lobe, Corrinne G.

    2015-01-01

    Trichostatin A (TSA) is a histone deacetylase inhibitor and a potential therapeutic for various malignancies. The in vivo effect of TSA, however, has not been investigated in a transgenic lung cancer model. Previously, we generated transgenic mice with overexpression of Groucho-related-gene 1 (Grg1) and these mice all developed mucinous lung adenocarcinoma. Grg1 is a transcriptional co-repressor protein, the function of which is thought to depend on HDAC activity. However, functions outside the nucleus have also been proposed. We tested the supposition that Grg1-induced tumorigenesis is HDAC-dependent by assaying the therapeutic effect of TSA in the Grg1 transgenic mouse model. We found that TSA significantly inhibited lung tumorigenesis in Grg1 transgenic mice (p < 0.01). TSA did not affect overall Grg1 protein levels, but instead reduced ErbB1 and ErbB2 expression, which are upregulated by Grg1 in the absence of TSA. We confirmed this effect in A549 cells. Furthermore, lapatinib, an inhibitor of both ErbB1 and ErbB2, effectively masked the effect of TSA on the inhibition of A549 cell proliferation and migration, suggesting TSA does work, at least in part, by downregulating ErbB receptors. We additionally found that TSA reduced the expression of VEGF and VEGFR2, but not basic FGF and FGFR1. Our findings indicate that TSA effectively inhibits Grg1-induced lung tumorigenesis through the down-regulation of ErbB1 and ErbB2, as well as reduced VEGF signaling. This suggests TSA and other HDAC inhibitors could have therapeutic value in the treatment of lung cancers with Grg1 overexpression. - Highlights: • TSA suppresses lung tumorigenesis in Grg1 overexpressing transgenic mice. • TSA does not affect overall Grg1 protein levels in the mice and in A549 cells. • TSA reduces ErbB1 and ErbB2 expression in the mice and in A549 cells. • Lapatinib masks TSA-induced inhibition of A549 cell proliferation and migration. • TSA inhibits VEGF signaling, but not basic FGF

  1. Trichostatin A suppresses lung adenocarcinoma development in Grg1 overexpressing transgenic mice

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Ju, E-mail: ju.liu@sdu.edu.cn [Medical Research Center, Shandong Provincial Qianfoshan Hospital, Shandong University, 16766 Jingshi Road, Jinan (China); Molecular and Cellular Biology Division, Sunnybrook Health Science Centre, University of Toronto, 2075 Bayview Avenue, Toronto, Ontario M4N 3M5 (Canada); Li, Yan [Children' s Health Care Center, Shandong Provincial Qianfoshan Hospital, Shandong University, 16766 Jingshi Road, Jinan, Shandong 250014 (China); Dong, Fengyun; Li, Liqun [Medical Research Center, Shandong Provincial Qianfoshan Hospital, Shandong University, 16766 Jingshi Road, Jinan (China); Masuda, Takahiro; Allen, Thaddeus D. [Molecular and Cellular Biology Division, Sunnybrook Health Science Centre, University of Toronto, 2075 Bayview Avenue, Toronto, Ontario M4N 3M5 (Canada); Lobe, Corrinne G. [Molecular and Cellular Biology Division, Sunnybrook Health Science Centre, University of Toronto, 2075 Bayview Avenue, Toronto, Ontario M4N 3M5 (Canada); Miami Mice Research Corp., MaRS Centre, Heritage Bldg., 101 College Street, Toronto, Ontario M5G 1L7 (Canada)

    2015-08-07

    Trichostatin A (TSA) is a histone deacetylase inhibitor and a potential therapeutic for various malignancies. The in vivo effect of TSA, however, has not been investigated in a transgenic lung cancer model. Previously, we generated transgenic mice with overexpression of Groucho-related-gene 1 (Grg1) and these mice all developed mucinous lung adenocarcinoma. Grg1 is a transcriptional co-repressor protein, the function of which is thought to depend on HDAC activity. However, functions outside the nucleus have also been proposed. We tested the supposition that Grg1-induced tumorigenesis is HDAC-dependent by assaying the therapeutic effect of TSA in the Grg1 transgenic mouse model. We found that TSA significantly inhibited lung tumorigenesis in Grg1 transgenic mice (p < 0.01). TSA did not affect overall Grg1 protein levels, but instead reduced ErbB1 and ErbB2 expression, which are upregulated by Grg1 in the absence of TSA. We confirmed this effect in A549 cells. Furthermore, lapatinib, an inhibitor of both ErbB1 and ErbB2, effectively masked the effect of TSA on the inhibition of A549 cell proliferation and migration, suggesting TSA does work, at least in part, by downregulating ErbB receptors. We additionally found that TSA reduced the expression of VEGF and VEGFR2, but not basic FGF and FGFR1. Our findings indicate that TSA effectively inhibits Grg1-induced lung tumorigenesis through the down-regulation of ErbB1 and ErbB2, as well as reduced VEGF signaling. This suggests TSA and other HDAC inhibitors could have therapeutic value in the treatment of lung cancers with Grg1 overexpression. - Highlights: • TSA suppresses lung tumorigenesis in Grg1 overexpressing transgenic mice. • TSA does not affect overall Grg1 protein levels in the mice and in A549 cells. • TSA reduces ErbB1 and ErbB2 expression in the mice and in A549 cells. • Lapatinib masks TSA-induced inhibition of A549 cell proliferation and migration. • TSA inhibits VEGF signaling, but not basic FGF

  2. The photosynthetic response of tobacco plants overexpressing ice plant aquaporin McMIPB to a soil water deficit and high vapor pressure deficit.

    Science.gov (United States)

    Kawase, Miki; Hanba, Yuko T; Katsuhara, Maki

    2013-07-01

    We investigated the photosynthetic capacity and plant growth of tobacco plants overexpressing ice plant (Mesembryanthemum crystallinum L.) aquaporin McMIPB under (1) a well-watered growth condition, (2) a well-watered and temporal higher vapor pressure deficit (VPD) condition, and (3) a soil water deficit growth condition to investigate the effect of McMIPB on photosynthetic responses under moderate soil and atmospheric humidity and water deficit conditions. Transgenic plants showed a significantly higher photosynthesis rate (by 48 %), higher mesophyll conductance (by 52 %), and enhanced growth under the well-watered growth condition than those of control plants. Decreases in the photosynthesis rate and stomatal conductance from ambient to higher VPD were slightly higher in transgenic plants than those in control plants. When plants were grown under the soil water deficit condition, decreases in the photosynthesis rate and stomatal conductance were less significant in transgenic plants than those in control plants. McMIPB is likely to work as a CO2 transporter, as well as control the regulation of stomata to water deficits.

  3. A wheat WRKY transcription factor TaWRKY10 confers tolerance to multiple abiotic stresses in transgenic tobacco.

    Directory of Open Access Journals (Sweden)

    Chen Wang

    Full Text Available WRKY transcription factors are reported to be involved in defense regulation, stress response and plant growth and development. However, the precise role of WRKY transcription factors in abiotic stress tolerance is not completely understood, especially in crops. In this study, we identified and cloned 10 WRKY genes from genome of wheat (Triticum aestivum L.. TaWRKY10, a gene induced by multiple stresses, was selected for further investigation. TaWRKY10 was upregulated by treatment with polyethylene glycol, NaCl, cold and H2O2. Result of Southern blot indicates that the wheat genome contains three copies of TaWRKY10. The TaWRKY10 protein is localized in the nucleus and functions as a transcriptional activator. Overexpression of TaWRKY10 in tobacco (Nicotiana tabacum L. resulted in enhanced drought and salt stress tolerance, mainly demonstrated by the transgenic plants exhibiting of increased germination rate, root length, survival rate, and relative water content under these stress conditions. Further investigation showed that transgenic plants also retained higher proline and soluble sugar contents, and lower reactive oxygen species and malonaldehyde contents. Moreover, overexpression of the TaWRKY10 regulated the expression of a series of stress related genes. Taken together, our results indicate that TaWRKY10 functions as a positive factor under drought and salt stresses by regulating the osmotic balance, ROS scavenging and transcription of stress related genes.

  4. Transgenic mice display hair loss and regrowth overexpressing mutant Hr gene.

    Science.gov (United States)

    Zhu, Kuicheng; Xu, Cunshuan; Zhang, Jintao; Chen, Yingying; Liu, Mengduan

    2017-10-30

    Mutations in the hairless (Hr) gene in both mice and humans have been implicated in the development of congenital atrichia, but the role of Hr in skin and hair follicle (HF) biology remains unknown. Here, we established transgenic mice (TG) overexpressing mutant Hr to investigate its specific role in the development of HF. Three transgenic lines were successfully constructed, and two of them (TG3 and TG8) displayed a pattern of hair loss and regrowth with alternation in the expression of HR protein. The mutant Hr gene inhibited the expression of the endogenous gene in transgenic individuals, which led to the development of alopecia. Interestingly, the hair regrew with the increase in the endogenous expression levels resulting from decreased mutant Hr expression. The findings of our study indicate that the changes in the expression of Hr result in hair loss or regrowth.

  5. Overexpression of BiP in tobacco alleviates endoplasmic reticulum stress

    NARCIS (Netherlands)

    Leborgne-Castel, N.; Jelitto- Dooren, E.P.W.M. van; Crofts, A.J.; Denecke, J.

    1999-01-01

    To study the role of the lumenal binding protein (BiP) in the transport and secretion of proteins, we have produced plants with altered BiP levels. Transgenic plants overexpressing BiP showed dramatically increased BiP mRNA levels but only a modest increase in BiP protein levels. The presence of

  6. The signal peptide-like segment of hpaXm is required for its association to the cell wall in transgenic tobacco plants.

    Science.gov (United States)

    Li, Le; Miao, Weiguo; Liu, Wenbo; Zhang, Shujian

    2017-01-01

    Harpins, encoded by hrp (hypersensitive response and pathogenicity) genes of Gram-negative plant pathogens, are elicitors of hypersensitive response (HR). HpaXm is a novel harpin-like protein described from cotton leaf blight bacteria, Xanthomonas citri subsp. malvacearum-a synonym of X. campestris pv. malvacearum (Smith 1901-1978). A putative signal peptide (1-MNSLNTQIGANSSFL-15) of hpaXm was predicted in the nitroxyl-terminal (N-terminal)by SignalP (SignalP 3.0 server). Here, we explored the function of the N-terminal leader peptide like segment of hpaXm using transgenic tobacco (Nicotiana tabacum cv. Xanthi nc.). Transgenic tobacco lines expressing the full-length hpaXm and the signal peptide-like segment-deleted mutant hpaXmΔLP were developed using transformation mediated by Agrobacterium tumefaciens. The target genes were confirmed integrated into the tobacco genomes and expressed normally. Using immune colloidal-gold detection technique, hpaXm protein was found to be transferred to the cytoplasm, the cell membrane, and organelles such as chloroplasts, mitochondria, and nucleus, as well as the cell wall. However, the deletion mutant hpaXmΔLP expressed in transgenic tobacco was found unable to cross the membrane to reach the cell wall. Additionally, soluble proteins extracted from plants transformed with hpaXm and hpaXmΔLP were bio-active. Defensive micro-HR induced by the transgene expression of hpaXm and hpaXmΔLP were observed on transgenic tobacco leaves. Disease resistance bioassays to tobacco mosaic virus (TMV) showed that tobacco plants transformed with hpaXm and with hpaXmΔLP exhibited enhanced resistance to TMV. In summary, the N-terminal signal peptide-like segment (1-45 bp) in hpaXm sequence is not necessary for transgene expression, bioactivity of hpaXm and resistance to TMV in transgenic tobacco, but is required for the protein to be translocated to the cell wall.

  7. Compensation of the AKT signaling by ERK signaling in transgenic mice hearts overexpressing TRIM72

    Energy Technology Data Exchange (ETDEWEB)

    Ham, Young-Mi, E-mail: youngmi_ham@hms.harvard.edu [College of Life Science and Biotechnology, Korea University, Seoul (Korea, Republic of); Department of Cell Biology, Harvard Medical School, Boston, MA 02115 (United States); Mahoney, Sarah Jane [Department of Cell Biology, Harvard Medical School, Boston, MA 02115 (United States)

    2013-06-10

    The AKT and ERK signaling pathways are known to be involved in cell hypertrophy, proliferation, survival and differentiation. Although there is evidence for crosstalk between these two signaling pathways in cellulo, there is less evidence for cross talk in vivo. Here, we show that crosstalk between AKT and ERK signaling in the hearts of TRIM72-overexpressing transgenic mice (TRIM72-Tg) with alpha-MHC promoter regulates and maintains their heart size. TRIM72, a heart- and skeletal muscle-specific protein, downregulates AKT-mTOR signaling via IRS-1 degradation and reduces the size of rat cardiomyocytes and the size of postnatal TRIM72-Tg hearts. TRIM72 expression was upregulated by hypertrophic inducers in cardiomyocytes, while IRS-1 was downregulated by IGF-1. TRIM72 specifically regulated IGF-1-dependent AKT-mTOR signaling, resulting in a reduction of the size of cardiomyocytes. Postnatal TRIM72-Tg hearts were smaller than control-treated hearts with inhibition of AKT-mTOR signaling. However, adult TRIM72-Tg hearts were larger than of control despite the suppression of AKT-mTOR signaling. Activation of ERK, PKC-α, and JNK were observed to be elevated in adult TRIM72-Tg, and these signals were mediated by ET-1 via the ET receptors A and B. Altogether, these results suggest that AKT signaling regulates cardiac hypertrophy in physiological conditions, and ERK signaling compensates for the absence of AKT signaling during TRIM72 overexpression, leading to pathological hypertrophy. -- Highlights: • TRIM72 inhibits AKT signaling through ubiquitination of IRS-1 in cardiac cells. • TRIM72 regulates the size of cardiac cells. • TRIM72 regulates size of postnatal TRIM72-overexpressing transgenic mice hearts. • Adult TRIM72-overexpressing transgenic mice hearts showed cardiac dysfunction. • Adult TRIM72 transgenic mice hearts showed higher expression of endothelin receptors.

  8. Compensation of the AKT signaling by ERK signaling in transgenic mice hearts overexpressing TRIM72

    International Nuclear Information System (INIS)

    Ham, Young-Mi; Mahoney, Sarah Jane

    2013-01-01

    The AKT and ERK signaling pathways are known to be involved in cell hypertrophy, proliferation, survival and differentiation. Although there is evidence for crosstalk between these two signaling pathways in cellulo, there is less evidence for cross talk in vivo. Here, we show that crosstalk between AKT and ERK signaling in the hearts of TRIM72-overexpressing transgenic mice (TRIM72-Tg) with alpha-MHC promoter regulates and maintains their heart size. TRIM72, a heart- and skeletal muscle-specific protein, downregulates AKT-mTOR signaling via IRS-1 degradation and reduces the size of rat cardiomyocytes and the size of postnatal TRIM72-Tg hearts. TRIM72 expression was upregulated by hypertrophic inducers in cardiomyocytes, while IRS-1 was downregulated by IGF-1. TRIM72 specifically regulated IGF-1-dependent AKT-mTOR signaling, resulting in a reduction of the size of cardiomyocytes. Postnatal TRIM72-Tg hearts were smaller than control-treated hearts with inhibition of AKT-mTOR signaling. However, adult TRIM72-Tg hearts were larger than of control despite the suppression of AKT-mTOR signaling. Activation of ERK, PKC-α, and JNK were observed to be elevated in adult TRIM72-Tg, and these signals were mediated by ET-1 via the ET receptors A and B. Altogether, these results suggest that AKT signaling regulates cardiac hypertrophy in physiological conditions, and ERK signaling compensates for the absence of AKT signaling during TRIM72 overexpression, leading to pathological hypertrophy. -- Highlights: • TRIM72 inhibits AKT signaling through ubiquitination of IRS-1 in cardiac cells. • TRIM72 regulates the size of cardiac cells. • TRIM72 regulates size of postnatal TRIM72-overexpressing transgenic mice hearts. • Adult TRIM72-overexpressing transgenic mice hearts showed cardiac dysfunction. • Adult TRIM72 transgenic mice hearts showed higher expression of endothelin receptors

  9. Overexpression of the OsIMP Gene Increases the Accumulation of Inositol and Confers Enhanced Cold Tolerance in Tobacco through Modulation of the Antioxidant Enzymes' Activities.

    Science.gov (United States)

    Zhang, Rong-Xiang; Qin, Li-Jun; Zhao, De-Gang

    2017-07-20

    Inositol is a cyclic polyol that is involved in various physiological processes, including signal transduction and stress adaptation in plants. l- myo -inositol monophosphatase (IMPase) is one of the metal-dependent phosphatase family members and catalyzes the last reaction step of biosynthesis of inositol. Although increased IMPase activity induced by abiotic stress has been reported in chickpea plants, the role and regulation of the IMP gene in rice ( Oryza sativa L.) remains poorly understood. In the present work, we obtained a full-length cDNA sequence coding IMPase in the cold tolerant rice landraces in Gaogonggui, which is named as OsIMP . Multiple alignment results have displayed that this sequence has characteristic signature motifs and conserved enzyme active sites of the phosphatase super family. Phylogenetic analysis showed that IMPase is most closely related to that of the wild rice Oryza brachyantha , while transcript analysis revealed that the expression of the OsIMP is significantly induced by cold stress and exogenous abscisic acid (ABA) treatment. Meanwhile, we cloned the 5' flanking promoter sequence of the OsIMP gene and identified several important cis -acting elements, such as LTR (low-temperature responsiveness), TCA-element (salicylic acid responsiveness), ABRE-element (abscisic acid responsiveness), GARE-motif (gibberellin responsive), MBS (MYB Binding Site) and other cis -acting elements related to defense and stress responsiveness. To further investigate the potential function of the OsIMP gene, we generated transgenic tobacco plants overexpressing the OsIMP gene and the cold tolerance test indicated that these transgenic tobacco plants exhibit improved cold tolerance. Furthermore, transgenic tobacco plants have a lower level of hydrogen peroxide (H₂O₂) and malondialdehyde (MDA), and a higher content of total chlorophyll as well as increased antioxidant enzyme activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD

  10. Cloning and functional analysis in transgenic tobacco of a tapetum ...

    African Journals Online (AJOL)

    The 5'-flanking region of 1174 bp upstream of the translation start point (TSP) of a reported Arabidopsis anther-specific gene, Anther7 gene (ATA7), which putatively encodes a protein related to lipid transfer protein, was cloned and functionally analyzed in transgenic tobacco after been fused with β- glucuronidase (GUS) ...

  11. Transgenic soybean plants overexpressing O-acetylserine sulfhydrylase accumulate enhanced levels of cysteine and Bowman-Birk protease inhibitor in seeds.

    Science.gov (United States)

    Kim, Won-Seok; Chronis, Demosthenis; Juergens, Matthew; Schroeder, Amy C; Hyun, Seung Won; Jez, Joseph M; Krishnan, Hari B

    2012-01-01

    Soybeans provide an excellent source of protein in animal feed. Soybean protein quality can be enhanced by increasing the concentration of sulfur-containing amino acids. Previous attempts to increase the concentration of sulfur-containing amino acids through the expression of heterologous proteins have met with limited success. Here, we report a successful strategy to increase the cysteine content of soybean seed through the overexpression of a key sulfur assimilatory enzyme. We have generated several transgenic soybean plants that overexpress a cytosolic isoform of O-acetylserine sulfhydrylase (OASS). These transgenic soybean plants exhibit a four- to tenfold increase in OASS activity when compared with non-transformed wild-type. The OASS activity in the transgenic soybeans was significantly higher at all the stages of seed development. Unlike the non-transformed soybean plants, there was no marked decrease in the OASS activity even at later stages of seed development. Overexpression of cytosolic OASS resulted in a 58-74% increase in protein-bound cysteine levels compared with non-transformed wild-type soybean seeds. A 22-32% increase in the free cysteine levels was also observed in transgenic soybeans overexpressing OASS. Furthermore, these transgenic soybean plants showed a marked increase in the accumulation of Bowman-Birk protease inhibitor, a cysteine-rich protein. The overall increase in soybean total cysteine content (both free and protein-bound) satisfies the recommended levels required for the optimal growth of monogastric animals.

  12. Overexpression of ZmIRT1 and ZmZIP3 Enhances Iron and Zinc Accumulation in Transgenic Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Suzhen Li

    Full Text Available Iron and zinc are important micronutrients for both the growth and nutrient availability of crop plants, and their absorption is tightly controlled by a metal uptake system. Zinc-regulated transporters, iron-regulated transporter-like proteins (ZIP, is considered an essential metal transporter for the acquisition of Fe and Zn in graminaceous plants. Several ZIPs have been identified in maize, although their physiological function remains unclear. In this report, ZmIRT1 was shown to be specifically expressed in silk and embryo, whereas ZmZIP3 was a leaf-specific gene. Both ZmIRT1 and ZmZIP3 were shown to be localized to the plasma membrane and endoplasmic reticulum. In addition, transgenic Arabidopsis plants overexpressing ZmIRT1 or ZmZIP3 were generated, and the metal contents in various tissues of transgenic and wild-type plants were examined based on ICP-OES and Zinpyr-1 staining. The Fe and Zn concentration increased in roots and seeds of ZmIRT1-overexpressing plants, while the Fe content in shoots decreased. Overexpressing ZmZIP3 enhanced Zn accumulation in the roots of transgenic plants, while that in shoots was repressed. In addition, the transgenic plants showed altered tolerance to various Fe and Zn conditions compared with wild-type plants. Furthermore, the genes associated with metal uptake were stimulated in ZmIRT1 transgenic plants, while those involved in intra- and inter- cellular translocation were suppressed. In conclusion, ZmIRT1 and ZmZIP3 are functional metal transporters with different ion selectivities. Ectopic overexpression of ZmIRT1 may stimulate endogenous Fe uptake mechanisms, which may facilitate metal uptake and homeostasis. Our results increase our understanding of the functions of ZIP family transporters in maize.

  13. Over-expression of a tobacco nitrate reductase gene in wheat (Triticum aestivum L.) increases seed protein content and weight without augmenting nitrogen supplying.

    Science.gov (United States)

    Zhao, Xiao-Qiang; Nie, Xuan-Li; Xiao, Xing-Guo

    2013-01-01

    Heavy nitrogen (N) application to gain higher yield of wheat (Triticum aestivum L.) resulted in increased production cost and environment pollution. How to diminish the N supply without losing yield and/or quality remains a challenge. To meet the challenge, we integrated and expressed a tobacco nitrate reductase gene (NR) in transgenic wheat. The 35S-NR gene was transferred into two winter cultivars, "Nongda146" and "Jimai6358", by Agrobacterium-mediation. Over-expression of the transgene remarkably enhanced T1 foliar NR activity and significantly augmented T2 seed protein content and 1000-grain weight in 63.8% and 68.1% of T1 offspring (total 67 individuals analyzed), respectively. Our results suggest that constitutive expression of foreign nitrate reductase gene(s) in wheat might improve nitrogen use efficiency and thus make it possible to increase seed protein content and weight without augmenting N supplying.

  14. Over-expression of a tobacco nitrate reductase gene in wheat (Triticum aestivum L. increases seed protein content and weight without augmenting nitrogen supplying.

    Directory of Open Access Journals (Sweden)

    Xiao-Qiang Zhao

    Full Text Available Heavy nitrogen (N application to gain higher yield of wheat (Triticum aestivum L. resulted in increased production cost and environment pollution. How to diminish the N supply without losing yield and/or quality remains a challenge. To meet the challenge, we integrated and expressed a tobacco nitrate reductase gene (NR in transgenic wheat. The 35S-NR gene was transferred into two winter cultivars, "Nongda146" and "Jimai6358", by Agrobacterium-mediation. Over-expression of the transgene remarkably enhanced T1 foliar NR activity and significantly augmented T2 seed protein content and 1000-grain weight in 63.8% and 68.1% of T1 offspring (total 67 individuals analyzed, respectively. Our results suggest that constitutive expression of foreign nitrate reductase gene(s in wheat might improve nitrogen use efficiency and thus make it possible to increase seed protein content and weight without augmenting N supplying.

  15. Overexpression of GmDREB1 improves salt tolerance in transgenic wheat and leaf protein response to high salinity

    Directory of Open Access Journals (Sweden)

    Qiyan Jiang

    2014-04-01

    Full Text Available The transcription factor dehydration-responsive element binding protein (DREB is able to improve tolerance to abiotic stress in plants by regulating the expression of downstream genes involved in environmental stress resistance. The objectives of this study were to evaluate the salt tolerance of GmDREB1 transgenic wheat (Triticum aestivum L. and to evaluate its physiological and protein responses to salt stress. Compared with the wild type, the transgenic lines overexpressing GmDREB1 showed longer coleoptiles and radicles and a greater radicle number at the germination stage, as well as greater root length, fresh weight, and tiller number per plant at the seedling stage. The yield-related traits of transgenic lines were also improved compared with the wild type, indicating enhanced salt tolerance in transgenic lines overexpressing GmDREB1. Proteomics analysis revealed that osmotic- and oxidative-stress-related proteins were up-regulated in transgenic wheat leaves under salt stress conditions. Transgenic wheat had higher levels of proline and betaine and lower levels of malondialdehyde and relative electrolyte leakage than the wild type. These results suggest that GmDREB1 regulates the expression of osmotic- and oxidative-stress-related proteins that reduce the occurrence of cell injury caused by high salinity, thus improving the salt tolerance of transgenic wheat.

  16. Enhanced tolerance and remediation of anthracene by transgenic tobacco plants expressing a fungal glutathione transferase gene

    International Nuclear Information System (INIS)

    Dixit, Prachy; Mukherjee, Prasun K.; Sherkhane, Pramod D.; Kale, Sharad P.; Eapen, Susan

    2011-01-01

    Highlights: → Transgenic plants expressing a TvGST gene were tested for tolerance, uptake and degradation of anthracene. → Transgenic plants were more tolerant to anthracene and take up more anthracene from soil and solutions compared to control plants. → Using in vitro T 1 seedlings, we showed that anthracene-a three fused benzene ring compound was phytodegraded to naphthalene derivatives, having two benzene rings. → This is the first time that a transgenic plant was shown to have the potential to phytodegrade anthracene. - Abstract: Plants can be used for remediation of polyaromatic hydrocarbons, which are known to be a major concern for human health. Metabolism of xenobiotic compounds in plants occurs in three phases and glutathione transferases (GST) mediate phase II of xenobiotic transformation. Plants, although have GSTs, they are not very efficient for degradation of exogenous recalcitrant xenobiotics including polyaromatic hydrocarbons. Hence, heterologous expression of efficient GSTs in plants may improve their remediation and degradation potential of xenobiotics. In the present study, we investigated the potential of transgenic tobacco plants expressing a Trichoderma virens GST for tolerance, remediation and degradation of anthracene-a recalcitrant polyaromatic hydrocarbon. Transgenic plants with fungal GST showed enhanced tolerance to anthracene compared to control plants. Remediation of 14 C uniformly labeled anthracene from solutions and soil by transgenic tobacco plants was higher compared to wild-type plants. Transgenic plants (T 0 and T 1 ) degraded anthracene to naphthalene derivatives, while no such degradation was observed in wild-type plants. The present work has shown that in planta expression of a fungal GST in tobacco imparted enhanced tolerance as well as higher remediation potential of anthracene compared to wild-type plants.

  17. Overexpressing the Sedum alfredii Cu/Zn Superoxide Dismutase Increased Resistance to Oxidative Stress in Transgenic Arabidopsis

    Directory of Open Access Journals (Sweden)

    Zhen Li

    2017-06-01

    Full Text Available Superoxide dismutase (SOD is a very important reactive oxygen species (ROS-scavenging enzyme. In this study, the functions of a Cu/Zn SOD gene (SaCu/Zn SOD, from Sedum alfredii, a cadmium (Cd/zinc/lead co-hyperaccumulator of the Crassulaceae, was characterized. The expression of SaCu/Zn SOD was induced by Cd stress. Compared with wild-type (WT plants, overexpression of SaCu/Zn SOD gene in transgenic Arabidopsis plants enhanced the antioxidative defense capacity, including SOD and peroxidase activities. Additionally, it reduced the damage associated with the overproduction of hydrogen peroxide (H2O2 and superoxide radicals (O2•-. The influence of Cd stress on ion flux across the root surface showed that overexpressing SaCu/Zn SOD in transgenic Arabidopsis plants has greater Cd uptake capacity existed in roots. A co-expression network based on microarray data showed possible oxidative regulation in Arabidopsis after Cd-induced oxidative stress, suggesting that SaCu/Zn SOD may participate in this network and enhance ROS-scavenging capability under Cd stress. Taken together, these results suggest that overexpressing SaCu/Zn SOD increased oxidative stress resistance in transgenic Arabidopsis and provide useful information for understanding the role of SaCu/Zn SOD in response to abiotic stress.

  18. Characterization of transgenic tobacco plants containing bacterial bphC gene and study of their phytoremediation ability.

    Science.gov (United States)

    Viktorovtá, Jitka; Novakova, Martina; Trbolova, Ladislava; Vrchotova, Blanka; Lovecka, Petra; Mackova, Martina; Macek, Tomas

    2014-01-01

    Genetically modified plants can serve as an efficient tool for remediation of diverse dangerous pollutants of the environment such as pesticides, heavy metals, explosives and persistent organic compounds. Transgenic lines of Nicotiana tabacum containing bacterial bphC gene from the degradation pathway of polychlorinated biphenyls (PCBs) were tested. The product of the bphC gene - enzyme 2,3-dihydroxybiphenyl-1,2-dioxygenase is responsible for cleaving of the biphenyl ring. The presence of bphC gene in transgenic plants was detected on DNA, RNA and protein level. The expression of the bphC/His gene was verified afterpurification of the enzyme from plants by affinity chromatography followed by a Western blot and immunochemical assay. The enzyme activity of isolated protein was detected. Efficient transformation of 2,3-DHB by transgenic plants was achieved and the lines also exhibited high production of biomass. The transgenic plants were more tolerant to the commercial PCBs mixture Delor 103 than non-transgenic tobacco. And finally, the higher decrease of total PCB content and especially congener 28 in real contaminated soil from a dumpsite was determined after cultivation of transgenic plant in comparison with nontransgenic tobacco. The substrate specificity of transgenic plants was the same as substrate specificity of BphC enzyme.

  19. Heterogenous expression of Pyrus pyrifolia PpCAD2 and PpEXP2 in tobacco impacts lignin accumulation in transgenic plants.

    Science.gov (United States)

    Wang, Yuling; Zhang, Xinfu; Yang, Shaolan; Wang, Caihong; Lu, Guilong; Wang, Ran; Yang, Yingjie; Li, Dingli

    2017-12-30

    Lignin, a natural macromolecular compound, plays an important role in the texture and taste of fruit. Hard end is a physiological disorder of pear fruit, in which the level of lignification in fruit tissues is dramatically elevated. Cinnamyl alcohol dehydrogenase and expansin genes (PpCAD2 and PpEXP2, respectively) exhibit higher levels of expression in 'Whangkeumbae' (Pyrus pyrifolia) pear fruit exhibiting this physiological disorder, relative to control fruit without symptoms. These genes were isolated from pear fruit and subsequently expressed in tobacco (Nicotiana tabacum) to investigate their function. Histochemical staining for lignin revealed that the degree of lignification in leaf veins and stem tissues increased in plants transformed with sense constructs and decreased in plants transformed with antisense constructs of PpCAD2. The expression of native NtCADs was also inhibited in the antisense PpCAD2 transgenic tobacco. Sense and antisense PpCAD2 transgenic tobacco exhibited an 86.7% increase and a 60% decrease in CAD activity, respectively, accompanied by a complementary response in lignin content in root tissues. The basal portion of the stem in PpEXP2 transgenic tobacco was bent and highly lignified. Additionally, the level of cellulose also increased in the stem of PpEXP2 transgenic tobacco. Collectively, these results suggested that PpCAD2 and PpEXP2 genes play a significant role in lignin accumulation in transgenic tobacco plants, and it is inferred that these two genes may also participate in the increased lignification observed in hard end pear fruit. Copyright © 2017. Published by Elsevier B.V.

  20. Tobacco randomly inserted tt8 differenly enhance light signals and flavonoid accumulation

    International Nuclear Information System (INIS)

    Sompornpailin, K.; Kanthang, S.

    2015-01-01

    The individual lines of tobacco over-expressing TT8, a bHLH gene, were constructed and cultured under tissue culture condition radiating with photosynthetically activation radiation (PAR) or PAR+UVA. They were compared to wild type (WT). Leaf of treated plants was extracted and analyzed for flavonoid accumulations using a spectrophotometer. The extract of TT8 plants significantly contained flavone, flavonol and anthocyanin level, higher than the WT extract did. The petal extracts of mature transgenic under PAR had a similar absorbance profile of each substance, but these extracts had higher flavonoid contents than the leaf extracts did. All flavonoid subgroups and p-coumaric acid biosynthesis were significantly enhanced after the additional UVA radiation to plant. This UVA condition slightly stimulated an accumulation of these substances in normal plant. Some transgenic greatly increased flavonoid accumulation in responding to PAR+UVA, but the others were slightly different compared to WT. The distinct insertion site is directly affected TT8 gene expression. Transgenic seeds had a dark brown color more than WT seed, which indicated high content of polymer flavonoids (proanthocyanins). This over-expressing TT8 in transgenic tobacco may directly or indirectly enhance the signal transductions of PAR and UVA and raise up flavonoid accumulation. (author)

  1. Expression and Chloroplast Targeting of Cholesterol Oxidase in Transgenic Tobacco Plants

    Science.gov (United States)

    Corbin, David R.; Grebenok, Robert J.; Ohnmeiss, Thomas E.; Greenplate, John T.; Purcell, John P.

    2001-01-01

    Cholesterol oxidase represents a novel type of insecticidal protein with potent activity against the cotton boll weevil (Anthonomus grandis grandis Boheman). We transformed tobacco (Nicotiana tabacum) plants with the cholesterol oxidase choM gene and expressed cytosolic and chloroplast-targeted versions of the ChoM protein. Transgenic leaf tissues expressing cholesterol oxidase exerted insecticidal activity against boll weevil larvae. Our results indicate that cholesterol oxidase can metabolize phytosterols in vivo when produced cytosolically or when targeted to chloroplasts. The transgenic plants exhibiting cytosolic expression accumulated low levels of saturated sterols known as stanols, and displayed severe developmental aberrations. In contrast, the transgenic plants expressing chloroplast-targeted cholesterol oxidase maintained a greater accumulation of stanols, and appeared phenotypically and developmentally normal. These results are discussed within the context of plant sterol distribution and metabolism. PMID:11457962

  2. Co-overexpressing a Plasma Membrane and a Vacuolar Membrane Sodium/Proton Antiporter Significantly Improves Salt Tolerance in Transgenic Arabidopsis Plants

    Science.gov (United States)

    Pehlivan, Necla; Sun, Li; Jarrett, Philip; Yang, Xiaojie; Mishra, Neelam; Chen, Lin; Kadioglu, Asim; Shen, Guoxin; Zhang, Hong

    2016-01-01

    The Arabidopsis gene AtNHX1 encodes a vacuolar membrane-bound sodium/proton (Na+/H+) antiporter that transports Na+ into the vacuole and exports H+ into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane-bound Na+/H+ antiporter that exports Na+ to the extracellular space and imports H+ into the plant cell. Plants rely on these enzymes either to keep Na+ out of the cell or to sequester Na+ into vacuoles to avoid the toxic level of Na+ in the cytoplasm. Overexpression of AtNHX1 or SOS1 could improve salt tolerance in transgenic plants, but the improved salt tolerance is limited. NaCl at concentration >200 mM would kill AtNHX1-overexpressing or SOS1-overexpressing plants. Here it is shown that co-overexpressing AtNHX1 and SOS1 could further improve salt tolerance in transgenic Arabidopsis plants, making transgenic Arabidopsis able to tolerate up to 250 mM NaCl treatment. Furthermore, co-overexpression of AtNHX1 and SOS1 could significantly reduce yield loss caused by the combined stresses of heat and salt, confirming the hypothesis that stacked overexpression of two genes could substantially improve tolerance against multiple stresses. This research serves as a proof of concept for improving salt tolerance in other plants including crops. PMID:26985021

  3. Ubiquitin fusion expression and tissue-dependent targeting of hG-CSF in transgenic tobacco

    Science.gov (United States)

    2011-01-01

    Background Human granulocyte colony-stimulating factor (hG-CSF) is an important human cytokine which has been widely used in oncology and infection protection. To satisfy clinical needs, expression of recombinant hG-CSF has been studied in several organisms, including rice cell suspension culture and transient expression in tobacco leaves, but there was no published report on its expression in stably transformed plants which can serve as a more economical expression platform with potential industrial application. Results In this study, hG-CSF expression was investigated in transgenic tobacco leaves and seeds in which the accumulation of hG-CSF could be enhanced through fusion with ubiquitin by up to 7 fold in leaves and 2 fold in seeds, leading to an accumulation level of 2.5 mg/g total soluble protein (TSP) in leaves and 1.3 mg/g TSP in seeds, relative to hG-CSF expressed without a fusion partner. Immunoblot analysis showed that ubiquitin was processed from the final protein product, and ubiquitination was up-regulated in all transgenic plants analyzed. Driven by CaMV 35S promoter and phaseolin signal peptide, hG-CSF was observed to be secreted into apoplast in leaves but deposited in protein storage vacuole (PSV) in seeds, indicating that targeting of the hG-CSF was tissue-dependent in transgenic tobacco. Bioactivity assay showed that hG-CSF expressed in both seeds and leaves was bioactive to support the proliferation of NFS-60 cells. Conclusions In this study, the expression of bioactive hG-CSF in transgenic plants was improved through ubiquitin fusion strategy, demonstrating that protein expression can be enhanced in both plant leaves and seeds through fusion with ubiquitin and providing a typical case of tissue-dependent expression of recombinant protein in transgenic plants. PMID:21985646

  4. Overexpression of a Panax ginseng tonoplast aquaporin alters salt tolerance, drought tolerance and cold acclimation ability in transgenic Arabidopsis plants.

    Science.gov (United States)

    Peng, Yanhui; Lin, Wuling; Cai, Weiming; Arora, Rajeev

    2007-08-01

    Water movement across cellular membranes is regulated largely by a family of water channel proteins called aquaporins (AQPs). Since several abiotic stresses such as, drought, salinity and freezing, manifest themselves via altering water status of plant cells and are linked by the fact that they all result in cellular dehydration, we overexpressed an AQP (tonoplast intrinsic protein) from Panax ginseng, PgTIP1, in transgenic Arabidopsis thaliana plants to test its role in plant's response to drought, salinity and cold acclimation (induced freezing tolerance). Under favorable conditions, PgTIP1 overexpression significantly increased plant growth as determined by the biomass production, and leaf and root morphology. PgTIP1 overexpression had beneficial effect on salt-stress tolerance as indicated by superior growth status and seed germination of transgenic plants under salt stress; shoots of salt-stressed transgenic plants also accumulated greater amounts of Na(+) compared to wild-type plants. Whereas PgTIP1 overexpression diminished the water-deficit tolerance of plants grown in shallow (10 cm deep) pots, the transgenic plants were significantly more tolerant to water stress when grown in 45 cm deep pots. The rationale for this contrasting response, apparently, comes from the differences in the root morphology and leaf water channel activity (speed of dehydration/rehydration) between the transgenic and wild-type plants. Plants overexpressed with PgTIP1 exhibited lower (relative to wild-type control) cold acclimation ability; however, this response was independent of cold-regulated gene expression. Our results demonstrate a significant function of PgTIP1 in growth and development of plant cells, and suggest that the water movement across tonoplast (via AQP) represents a rate-limiting factor for plant vigor under favorable growth conditions and also significantly affect responses of plant to drought, salt and cold stresses.

  5. Enhanced tolerance and remediation of anthracene by transgenic tobacco plants expressing a fungal glutathione transferase gene

    Energy Technology Data Exchange (ETDEWEB)

    Dixit, Prachy; Mukherjee, Prasun K.; Sherkhane, Pramod D.; Kale, Sharad P. [Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai 400085 (India); Eapen, Susan, E-mail: eapenhome@yahoo.com [Nuclear Agriculture and Biotechnology Division, Bhabha Atomic Research Centre, Mumbai 400085 (India)

    2011-08-15

    Highlights: {yields} Transgenic plants expressing a TvGST gene were tested for tolerance, uptake and degradation of anthracene. {yields} Transgenic plants were more tolerant to anthracene and take up more anthracene from soil and solutions compared to control plants. {yields} Using in vitro T{sub 1} seedlings, we showed that anthracene-a three fused benzene ring compound was phytodegraded to naphthalene derivatives, having two benzene rings. {yields} This is the first time that a transgenic plant was shown to have the potential to phytodegrade anthracene. - Abstract: Plants can be used for remediation of polyaromatic hydrocarbons, which are known to be a major concern for human health. Metabolism of xenobiotic compounds in plants occurs in three phases and glutathione transferases (GST) mediate phase II of xenobiotic transformation. Plants, although have GSTs, they are not very efficient for degradation of exogenous recalcitrant xenobiotics including polyaromatic hydrocarbons. Hence, heterologous expression of efficient GSTs in plants may improve their remediation and degradation potential of xenobiotics. In the present study, we investigated the potential of transgenic tobacco plants expressing a Trichoderma virens GST for tolerance, remediation and degradation of anthracene-a recalcitrant polyaromatic hydrocarbon. Transgenic plants with fungal GST showed enhanced tolerance to anthracene compared to control plants. Remediation of {sup 14}C uniformly labeled anthracene from solutions and soil by transgenic tobacco plants was higher compared to wild-type plants. Transgenic plants (T{sub 0} and T{sub 1}) degraded anthracene to naphthalene derivatives, while no such degradation was observed in wild-type plants. The present work has shown that in planta expression of a fungal GST in tobacco imparted enhanced tolerance as well as higher remediation potential of anthracene compared to wild-type plants.

  6. Transgenic tobacco plants having a higher level of methionine are more sensitive to oxidative stress.

    Science.gov (United States)

    Hacham, Yael; Matityahu, Ifat; Amir, Rachel

    2017-07-01

    Methionine is an essential amino acid the low level of which limits the nutritional quality of plants. We formerly produced transgenic tobacco (Nicotiana tabacum) plants overexpressing CYSTATHIONE γ-SYNTHASE (CGS) (FA plants), methionine's main regulatory enzyme. These plants accumulate significantly higher levels of methionine compared with wild-type (WT) plants. The aim of this study was to gain more knowledge about the effect of higher methionine content on the metabolic profile of vegetative tissue and on the morphological and physiological phenotypes. FA plants exhibit slightly reduced growth, and metabolic profiling analysis shows that they have higher contents of stress-related metabolites. Despite this, FA plants were more sensitive to short- and long-term oxidative stresses. In addition, compared with WT plants and transgenic plants expressing an empty vector, the primary metabolic profile of FA was altered less during oxidative stress. Based on morphological and metabolic phenotypes, we strongly proposed that FA plants having higher levels of methionine suffer from stress under non-stress conditions. This might be one of the reasons for their lesser ability to cope with oxidative stress when it appeared. The observation that their metabolic profiling is much less responsive to stress compared with control plants indicates that the delta changes in metabolite contents between non-stress and stress conditions is important for enabling the plants to cope with stress conditions. © 2017 Scandinavian Plant Physiology Society.

  7. Transgenic rats overexpressing the human MrgX3 gene show cataracts and an abnormal skin phenotype

    International Nuclear Information System (INIS)

    Kaisho, Yoshihiko; Watanabe, Takuya; Nakata, Mitsugu; Yano, Takashi; Yasuhara, Yoshitaka; Shimakawa, Kozo; Mori, Ikuo; Sakura, Yasufumi; Terao, Yasuko; Matsui, Hideki; Taketomi, Shigehisa

    2005-01-01

    The human MrgX3 gene, belonging to the mrgs/SNSRs (mass related genes/sensory neuron specific receptors) family, was overexpressed in transgenic rats using the actin promoter. Two animal lines showed cataracts with liquification/degeneration and swelling of the lens fiber cells. The transient epidermal desquamation was observed in line with higher gene expression. Histopathology of the transgenic rats showed acanthosis and focal parakeratosis. In the epidermis, there was an increase in cellular keratin 14, keratin 10, and loricrin, as well as PGP 9.5 in innervating nerve fibers. These phenotypes accompanied an increase in the number of proliferating cells. These results suggest that overexpression of the human MrgX3 gene causes a disturbance of the normal cell-differentiation process

  8. Parvalbumin overexpression alters immune-mediated increases in intracellular calcium, and delays disease onset in a transgenic model of familial amyotrophic lateral sclerosis

    Science.gov (United States)

    Beers, D. R.; Ho, B. K.; Siklos, L.; Alexianu, M. E.; Mosier, D. R.; Mohamed, A. H.; Otsuka, Y.; Kozovska, M. E.; McAlhany, R. E.; Smith, R. G.; hide

    2001-01-01

    Intracellular calcium is increased in vulnerable spinal motoneurons in immune-mediated as well as transgenic models of amyotrophic lateral sclerosis (ALS). To determine whether intracellular calcium levels are influenced by the calcium-binding protein parvalbumin, we developed transgenic mice overexpressing parvalbumin in spinal motoneurons. ALS immunoglobulins increased intracellular calcium and spontaneous transmitter release at motoneuron terminals in control animals, but not in parvalbumin overexpressing transgenic mice. Parvalbumin transgenic mice interbred with mutant SOD1 (mSOD1) transgenic mice, an animal model of familial ALS, had significantly reduced motoneuron loss, and had delayed disease onset (17%) and prolonged survival (11%) when compared with mice with only the mSOD1 transgene. These results affirm the importance of the calcium binding protein parvalbumin in altering calcium homeostasis in motoneurons. The increased motoneuron parvalbumin can significantly attenuate the immune-mediated increases in calcium and to a lesser extent compensate for the mSOD1-mediated 'toxic-gain-of-function' in transgenic mice.

  9. Transgenic ipt tobacco overproducing cytokinins overaccumulates phenolic compounds during in vitro growth

    Czech Academy of Sciences Publication Activity Database

    Schnablová, Renáta; Synková, Helena; Vičánková, Anna; Burketová, Lenka; Eder, Josef; Cvikrová, Milena

    2006-01-01

    Roč. 44, - (2006), s. 526-534 ISSN 0981-9428 R&D Projects: GA ČR GA206/03/0310 Institutional research plan: CEZ:AV0Z50380511 Keywords : cytokinins * ipt * transgenic tobacco * in vitro cultivation Subject RIV: EF - Botanics Impact factor: 1.847, year: 2006

  10. Overexpression of the OsIMP Gene Increases the Accumulation of Inositol and Confers Enhanced Cold Tolerance in Tobacco through Modulation of the Antioxidant Enzymes’ Activities

    Directory of Open Access Journals (Sweden)

    Rong-Xiang Zhang

    2017-07-01

    Full Text Available Inositol is a cyclic polyol that is involved in various physiological processes, including signal transduction and stress adaptation in plants. l-myo-inositol monophosphatase (IMPase is one of the metal-dependent phosphatase family members and catalyzes the last reaction step of biosynthesis of inositol. Although increased IMPase activity induced by abiotic stress has been reported in chickpea plants, the role and regulation of the IMP gene in rice (Oryza sativa L. remains poorly understood. In the present work, we obtained a full-length cDNA sequence coding IMPase in the cold tolerant rice landraces in Gaogonggui, which is named as OsIMP. Multiple alignment results have displayed that this sequence has characteristic signature motifs and conserved enzyme active sites of the phosphatase super family. Phylogenetic analysis showed that IMPase is most closely related to that of the wild rice Oryza brachyantha, while transcript analysis revealed that the expression of the OsIMP is significantly induced by cold stress and exogenous abscisic acid (ABA treatment. Meanwhile, we cloned the 5’ flanking promoter sequence of the OsIMP gene and identified several important cis-acting elements, such as LTR (low-temperature responsiveness, TCA-element (salicylic acid responsiveness, ABRE-element (abscisic acid responsiveness, GARE-motif (gibberellin responsive, MBS (MYB Binding Site and other cis-acting elements related to defense and stress responsiveness. To further investigate the potential function of the OsIMP gene, we generated transgenic tobacco plants overexpressing the OsIMP gene and the cold tolerance test indicated that these transgenic tobacco plants exhibit improved cold tolerance. Furthermore, transgenic tobacco plants have a lower level of hydrogen peroxide (H2O2 and malondialdehyde (MDA, and a higher content of total chlorophyll as well as increased antioxidant enzyme activities of superoxide dismutase (SOD, catalase (CAT and peroxidase (POD

  11. Ectopic Expression of the Coleus R2R3 MYB-Type Proanthocyanidin Regulator Gene SsMYB3 Alters the Flower Color in Transgenic Tobacco.

    Directory of Open Access Journals (Sweden)

    Qinlong Zhu

    Full Text Available Proanthocyanidins (PAs play an important role in plant disease defense and have beneficial effects on human health. We isolated and characterized a novel R2R3 MYB-type PA-regulator SsMYB3 from a well-known ornamental plant, coleus (Solenostemon scutellarioides, to study the molecular regulation of PAs and to engineer PAs biosynthesis. The expression level of SsMYB3 was correlated with condensed tannins contents in various coleus tissues and was induced by wounding and light. A complementation test in the Arabidopsis tt2 mutant showed that SsMYB3 could restore the PA-deficient seed coat phenotype and activated expression of the PA-specific gene ANR and two related genes, DFR and ANS. In yeast two-hybrid assays, SsMYB3 interacted with the Arabidopsis AtTT8 and AtTTG1 to reform the ternary transcriptional complex, and also interacted with two tobacco bHLH proteins (NtAn1a and NtJAF13-1 and a WD40 protein, NtAn11-1. Ectopic overexpression of SsMYB3 in transgenic tobacco led to almost-white flowers by greatly reducing anthocyanin levels and enhancing accumulation of condensed tannins. This overexpression of SsMYB3 upregulated the key PA genes (NtLAR and NtANR and late anthocyanin structural genes (NtDFR and NtANS, but downregulated the expression of the final anthocyanin gene NtUFGT. The formative SsMYB3-complex represses anthocyanin accumulation by directly suppressing the expression of the final anthocyanin structural gene NtUFGT, through competitive inhibition or destabilization of the endogenous NtAn2-complex formation. These results suggested that SsMYB3 may form a transcription activation complex to regulate PA biosynthesis in the Arabidopsis tt2 mutant and transgenic tobacco. Our findings suggest that SsMYB3 is involved in the regulation of PA biosynthesis in coleus and has the potential as a molecular tool for manipulating biosynthesis of PAs in fruits and other crops using metabolic engineering.

  12. Pea p68, a DEAD-box helicase, provides salinity stress tolerance in transgenic tobacco by reducing oxidative stress and improving photosynthesis machinery.

    Science.gov (United States)

    Tuteja, Narendra; Banu, Mst Sufara Akhter; Huda, Kazi Md Kamrul; Gill, Sarvajeet Singh; Jain, Parul; Pham, Xuan Hoi; Tuteja, Renu

    2014-01-01

    The DEAD-box helicases are required mostly in all aspects of RNA and DNA metabolism and they play a significant role in various abiotic stresses, including salinity. The p68 is an important member of the DEAD-box proteins family and, in animal system, it is involved in RNA metabolism including pre-RNA processing and splicing. In plant system, it has not been well characterized. Here we report the cloning and characterization of p68 from pea (Pisum sativum) and its novel function in salinity stress tolerance in plant. The pea p68 protein self-interacts and is localized in the cytosol as well as the surrounding of cell nucleus. The transcript of pea p68 is upregulated in response to high salinity stress in pea. Overexpression of p68 driven by constitutive cauliflower mosaic virus-35S promoter in tobacco transgenic plants confers enhanced tolerances to salinity stress by improving the growth, photosynthesis and antioxidant machinery. Under stress treatment, pea p68 overexpressing tobacco accumulated higher K+ and lower Na+ level than the wild-type plants. Reactive oxygen species (ROS) accumulation was remarkably regulated by the overexpression of pea p68 under salinity stress conditions, as shown from TBARS content, electrolyte leakage, hydrogen peroxide accumulation and 8-OHdG content and antioxidant enzyme activities. To the best of our knowledge this is the first direct report, which provides the novel function of pea p68 helicase in salinity stress tolerance. The results suggest that p68 can also be exploited for engineering abiotic stress tolerance in crop plants of economic importance.

  13. TaPP2C1, a Group F2 Protein Phosphatase 2C Gene, Confers Resistance to Salt Stress in Transgenic Tobacco.

    Directory of Open Access Journals (Sweden)

    Wei Hu

    Full Text Available Group A protein phosphatases 2Cs (PP2Cs are essential components of abscisic acid (ABA signaling in Arabidopsis; however, the function of group F2 subfamily PP2Cs is currently less known. In this study, TaPP2C1 which belongs to group F2 was isolated and characterized from wheat. Expression of the TaPP2C1-GFP fusion protein suggested its ubiquitous localization within a cell. TaPP2C1 expression was downregulated by abscisic acid (ABA and NaCl treatments, but upregulated by H2O2 treatment. Overexpression of TaPP2C1 in tobacco resulted in reduced ABA sensitivity and increased salt resistance of transgenic seedlings. Additionally, physiological analyses showed that improved resistance to salt stress conferred by TaPP2C1 is due to the reduced reactive oxygen species (ROS accumulation, the improved antioxidant system, and the increased transcription of genes in the ABA-independent pathway. Finally, transgenic tobacco showed increased resistance to oxidative stress by maintaining a more effective antioxidant system. Taken together, these results demonstrated that TaPP2C1 negatively regulates ABA signaling, but positively regulates salt resistance. TaPP2C1 confers salt resistance through activating the antioxidant system and ABA-independent gene transcription process.

  14. Overexpression of a flower-specific aerolysin-like protein from the dioecious plant Rumex acetosa alters flower development and induces male sterility in transgenic tobacco.

    Science.gov (United States)

    Manzano, Susana; Megías, Zoraida; Martínez, Cecilia; García, Alicia; Aguado, Encarnación; Chileh, Tarik; López-Alonso, Diego; García-Maroto, Federico; Kejnovský, Eduard; Široký, Jiří; Kubát, Zdeněk; Králová, Tereza; Vyskot, Boris; Jamilena, Manuel

    2017-01-01

    Sex determination in Rumex acetosa, a dioecious plant with a complex XY 1 Y 2 sex chromosome system (females are XX and males are XY 1 Y 2 ), is not controlled by an active Y chromosome but depends on the ratio between the number of X chromosomes and autosomes. To gain insight into the molecular mechanisms of sex determination, we generated a subtracted cDNA library enriched in genes specifically or predominantly expressed in female floral buds in early stages of development, when sex determination mechanisms come into play. In the present paper, we report the molecular and functional characterization of FEM32, a gene encoding a protein that shares a common architecture with proteins in different plants, animals, bacteria and fungi of the aerolysin superfamily; many of these function as β pore-forming toxins. The expression analysis, assessed by northern blot, RT-PCR and in situ hybridization, demonstrates that this gene is specifically expressed in flowers in both early and late stages of development, although its transcripts accumulate much more in female flowers than in male flowers. The ectopic expression of FEM32 under both the constitutive promoter 35S and the flower-specific promoter AP3 in transgenic tobacco showed no obvious alteration in vegetative development but was able to alter floral organ growth and pollen fertility. The 35S::FEM32 and AP3::FEM32 transgenic lines showed a reduction in stamen development and pollen viability, as well as a diminution in fruit set, fruit development and seed production. Compared with other floral organs, pistil development was, however, enhanced in plants overexpressing FEM32. According to these effects, it is likely that FEM32 functions in Rumex by arresting stamen and pollen development during female flower development. The aerolysin-like pore-forming proteins of eukaryotes are mainly involved in defence mechanisms against bacteria, fungi and insects and are also involved in apoptosis and programmed cell death (PCD

  15. Co-overexpressing a Plasma Membrane and a Vacuolar Membrane Sodium/Proton Antiporter Significantly Improves Salt Tolerance in Transgenic Arabidopsis Plants.

    Science.gov (United States)

    Pehlivan, Necla; Sun, Li; Jarrett, Philip; Yang, Xiaojie; Mishra, Neelam; Chen, Lin; Kadioglu, Asim; Shen, Guoxin; Zhang, Hong

    2016-05-01

    The Arabidopsis gene AtNHX1 encodes a vacuolar membrane-bound sodium/proton (Na(+)/H(+)) antiporter that transports Na(+) into the vacuole and exports H(+) into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane-bound Na(+)/H(+) antiporter that exports Na(+) to the extracellular space and imports H(+) into the plant cell. Plants rely on these enzymes either to keep Na(+) out of the cell or to sequester Na(+) into vacuoles to avoid the toxic level of Na(+) in the cytoplasm. Overexpression of AtNHX1 or SOS1 could improve salt tolerance in transgenic plants, but the improved salt tolerance is limited. NaCl at concentration >200 mM would kill AtNHX1-overexpressing or SOS1-overexpressing plants. Here it is shown that co-overexpressing AtNHX1 and SOS1 could further improve salt tolerance in transgenic Arabidopsis plants, making transgenic Arabidopsis able to tolerate up to 250 mM NaCl treatment. Furthermore, co-overexpression of AtNHX1 and SOS1 could significantly reduce yield loss caused by the combined stresses of heat and salt, confirming the hypothesis that stacked overexpression of two genes could substantially improve tolerance against multiple stresses. This research serves as a proof of concept for improving salt tolerance in other plants including crops. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

  16. Herbicidal and antioxidant responses of transgenic rice overexpressing Myxococcus xanthus protoporphyrinogen oxidase.

    Science.gov (United States)

    Jung, Sunyo; Back, Kyoungwhan

    2005-05-01

    We analyzed the herbicidal and antioxidant defense responses of transgenic rice plants that overexpressed the Myxococcus xanthus protoporphyrinogen oxidase gene. Leaf squares of the wild-type incubated with oxyfluorfen were characterized by necrotic leaf lesions and increases in conductivity and malonyldialdehyde levels, whereas transgenic lines M4 and M7 did not show any change with up to 100 microM oxyfluorfen. The wild-type had decreased F(v)/F(m) and produced a high level of H(2)O(2) at 18 h after foliar application of oxyfluorfen, whereas transgenic lines M4 and M7 were unaffected. In response to oxyfluorfen, violaxanthin, beta-carotene, and chlorophylls (Chls) decreased in wild-type plants, whereas antheraxanthin and zeaxanthin increased. Only a slight decline in Chls was observed in transgenic lines at 48 h after oxyfluorfen treatment. Noticeable increases of cytosolic Cu/Zn-superoxide dismutase, peroxidase isozymes 1 and 2, and catalase were observed after at 48 h of oxyfluorfen treatment in the wild-type. Non-enzymatic antioxidants appeared to respond faster to oxyfluorfen-induced photodynamic stress than did enzymatic antioxidants. Protective responses for the detoxification of active oxygen species were induced to counteract photodynamic stress in oxyfluorfen-treated, wild-type plants. However, oxyfluorfen-treated, transgenic plants suffered less oxidative stress, confirming increased herbicidal resistance resulted from dual expression of M. xanthus Protox in chloroplasts and mitochondria.

  17. An Alcohol Dehydrogenase Gene from Synechocystis sp. Confers Salt Tolerance in Transgenic Tobacco

    Directory of Open Access Journals (Sweden)

    So Young Yi

    2017-11-01

    Full Text Available Synechocystis salt-responsive gene 1 (sysr1 was engineered for expression in higher plants, and gene construction was stably incorporated into tobacco plants. We investigated the role of Sysr1 [a member of the alcohol dehydrogenase (ADH superfamily] by examining the salt tolerance of sysr1-overexpressing (sysr1-OX tobacco plants using quantitative real-time polymerase chain reactions, gas chromatography-mass spectrometry, and bioassays. The sysr1-OX plants exhibited considerably increased ADH activity and tolerance to salt stress conditions. Additionally, the expression levels of several stress-responsive genes were upregulated. Moreover, airborne signals from salt-stressed sysr1-OX plants triggered salinity tolerance in neighboring wild-type (WT plants. Therefore, Sysr1 enhanced the interconversion of aldehydes to alcohols, and this occurrence might affect the quality of green leaf volatiles (GLVs in sysr1-OX plants. Actually, the Z-3-hexenol level was approximately twofold higher in sysr1-OX plants than in WT plants within 1–2 h of wounding. Furthermore, analyses of WT plants treated with vaporized GLVs indicated that Z-3-hexenol was a stronger inducer of stress-related gene expression and salt tolerance than E-2-hexenal. The results of the study suggested that increased C6 alcohol (Z-3-hexenol induced the expression of resistance genes, thereby enhancing salt tolerance of transgenic plants. Our results revealed a role for ADH in salinity stress responses, and the results provided a genetic engineering strategy that could improve the salt tolerance of crops.

  18. Transgenic overexpression of p23 induces spontaneous hydronephrosis in mice

    Science.gov (United States)

    Lee, Jaehoon; Kim, Hye Jin; Moon, Jung Ah; Sung, Young Hoon; Baek, In-Jeoung; Roh, Jae-il; Ha, Na Young; Kim, Seung-Yeon; Bahk, Young Yil; Lee, Jong Eun; Yoo, Tae Hyun; Lee, Han-Woong

    2011-01-01

    p23 is a cochaperone of heat shock protein 90 and also interacts functionally with numerous steroid receptors and kinases. However, the in vivo roles of p23 remain unclear. To explore its in vivo function, we generated the transgenic (TG) mice ubiquitously overexpressing p23. The p23 TG mice spontaneously developed kidney abnormalities closely resembling human hydronephrosis. Consistently, kidney functions deteriorate significantly in the p23 TG mice compared to their wild-type (WT) littermates. Furthermore, the expression of target genes for aryl hydrocarbon receptor (AhR), such as cytochrome P450, family 1, subfamily A, polypeptide 1 (Cyp1A1) and cytochrome P450, family 1, subfamily B, polypeptide 1 (Cyp1B1), were induced in the kidneys of the p23 TG mice. These results indicate that the overexpression of p23 contributes to the development of hydronephrosis through the upregulation of the AhR pathway in vivo. PMID:21323770

  19. Expression of kenaf mitochondrial chimeric genes HM184 causes male sterility in transgenic tobacco plants.

    Science.gov (United States)

    Zhao, Yanhong; Liao, Xiaofang; Huang, Zhipeng; Chen, Peng; Zhou, Bujin; Liu, Dongmei; Kong, Xiangjun; Zhou, Ruiyang

    2015-08-01

    Chimeric genes resulting from the rearrangement of a mitochondrial genome were generally thought to be a causal factor in the occurrence of cytoplasmic male sterility (CMS). In the study, earlier we reported that identifying a 47 bp deletion at 3'- flanking of atp9 that was linked to male sterile cytoplasm in kenaf. The truncated fragment was fused with atp9, a mitochondrial transit signal (MTS) and/or GFP, comprised two chimeric genes MTS-HM184-GFP and MTS-HM184. The plant expression vector pBI121 containing chimeric genes were then introduced to tobacco plants by Agrobacterium-mediated T-DNA transformation. The result showed that certain transgenic plants were male sterility or semi-sterility, while some were not. The expression analysis further demonstrated that higher level of expression were showed in the sterility plants, while no expression or less expression in fertility plants, the levels of expression of semi-sterility were in between. And the sterile plant (containing MTS-HM184-GFP) had abnormal anther produced malformed/shriveled pollen grains stained negative that failed to germinate (0%), the corresponding fruits was shrunken, the semi-sterile plants having normal anther shape produced about 10-50% normal pollen grains, the corresponding fruits were not full, and the germination rate was 58%. Meanwhile these transgenic plants which altered on fertility were further analyzed in phenotype. As a result, the metamorphosis leaves were observed in the seedling stage, the plant height of transgenic plants was shorter than wild type. The growth duration of transgenic tobacco was delayed 30-45 days compared to the wild type. The copy numbers of target genes of transgenic tobacco were analyzed using the real-time quantitative method. The results showed that these transgenic plants targeting-expression in mitochondrial containing MTS-HM184-GFP had 1 copy and 2 copies, the other two plants containing MTS-HM184 both had 3 copies, but 0 copy in wild type. In

  20. The garlic NF-YC gene, AsNF-YC8, positively regulates non-ionic hyperosmotic stress tolerance in tobacco.

    Science.gov (United States)

    Sun, Xiudong; Lian, Haifeng; Liu, Xingchen; Zhou, Shumei; Liu, Shiqi

    2017-05-01

    To investigate the relationship between nuclear factor Y (NF-Y) and stress tolerance in garlic, we cloned a NF-Y family gene AsNF-YC8 from garlic, which was largely upregulated at dehydrate stage. Expression pattern analyses in garlic revealed that AsNF-YC8 is induced through abscisic acid (ABA) and abiotic stresses, such as NaCl and PEG. Compared with wild-type plants, the overexpressing-AsNF-YC8 transgenic tobacco plants showed higher seed germination rates, longer root length and better plant growth under salt and drought stresses. Under drought stress, the transgenic plants maintained higher relative water content (RWC), net photosynthesis, lower levels of malondialdehyde (MDA), and less ion leakage (IL) than wild-type control plants. These results indicate the high tolerance of the transgenic plants to drought stress compared to the WT. The transgenic tobacco lines accumulated less reactive oxygen species (ROS) and exhibited higher antioxidative enzyme activities compared with wild-type (WT) plants under drought stress, which suggested that the overexpression of AsNF-YC8 improves the antioxidant defense system by regulating the activities of these antioxidant enzymes, which in turn protect transgenic lines against drought stress. These results suggest that AsNF-YC8 plays an important role in tolerance to drought and salt stresses.

  1. Overexpression of GsZFP1 enhances salt and drought tolerance in transgenic alfalfa (Medicago sativa L.).

    Science.gov (United States)

    Tang, Lili; Cai, Hua; Ji, Wei; Luo, Xiao; Wang, Zhenyu; Wu, Jing; Wang, Xuedong; Cui, Lin; Wang, Yang; Zhu, Yanming; Bai, Xi

    2013-10-01

    GsZFP1 encodes a Cys2/His2-type zinc-finger protein. In our previous study, when GsZFP1 was heterologously expressed in Arabidopsis, the transgenic Arabidopsis plants exhibited enhanced drought and cold tolerance. However, it is still unknown whether GsZFP1 is also involved in salt stress. GsZFP1 is from the wild legume Glycine soja. Therefore, the aims of this study were to further elucidate the functions of the GsZFP1 gene under salt and drought stress in the forage legume alfalfa and to investigate its biochemical and physiological functions under these stress conditions. Our data showed that overexpression of GsZFP1 in alfalfa resulted in enhanced salt tolerance. Under high salinity stress, greater relative membrane permeability and malondialdehyde (MDA) content were observed and more free proline and soluble sugars accumulated in transgenic alfalfa than in the wild-type (WT) plants; in addition, the transgenic lines accumulated less Na(+) and more K(+) in both the shoots and roots. Overexpression of GsZFP1 also enhanced the drought tolerance of alfalfa. The fold-inductions of stress-responsive marker gene expression, including MtCOR47, MtRAB18, MtP5CS, and MtRD2, were greater in transgenic alfalfa than those of WT under drought stress conditions. In conclusion, the transgenic alfalfa plants generated in this study could be used for farming in salt-affected as well as arid and semi-arid areas. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  2. Enhanced water stress tolerance of transgenic maize plants over-expressing LEA Rab28 gene.

    Science.gov (United States)

    Amara, Imen; Capellades, Montserrat; Ludevid, M Dolors; Pagès, Montserrat; Goday, Adela

    2013-06-15

    Late Embryogenesis Abundant (LEA) proteins participate in plant stress responses and contribute to the acquisition of desiccation tolerance. In this report Rab28 LEA gene has been over-expressed in maize plants under a constitutive maize promoter. The expression of Rab28 transcripts led to the accumulation and stability of Rab28 protein in the transgenic plants. Native Rab28 protein is localized to nucleoli in wild type maize embryo cells; here we find by whole-mount immunocytochemistry that in root cells of Rab28 transgenic and wild-type plants the protein is also associated to nucleolar structures. Transgenic plants were tested for stress tolerance and resulted in sustained growth under polyethyleneglycol (PEG)-mediated dehydration compared to wild-type controls. Under osmotic stress transgenic seedlings showed increased leaf and root areas, higher relative water content (RWC), reduced chlorophyll loss and lower Malondialdehyde (MDA) production in relation to wild-type plants. Moreover, transgenic seeds exhibited higher germination rates than wild-type seeds under water deficit. Overall, our results highlight the presence of transgenic Rab28 protein in nucleolar structures and point to the potential of group 5 LEA Rab28 gene as candidate to enhance stress tolerance in maize plants. Copyright © 2013 Elsevier GmbH. All rights reserved.

  3. Pea p68, a DEAD-box helicase, provides salinity stress tolerance in transgenic tobacco by reducing oxidative stress and improving photosynthesis machinery.

    Directory of Open Access Journals (Sweden)

    Narendra Tuteja

    Full Text Available The DEAD-box helicases are required mostly in all aspects of RNA and DNA metabolism and they play a significant role in various abiotic stresses, including salinity. The p68 is an important member of the DEAD-box proteins family and, in animal system, it is involved in RNA metabolism including pre-RNA processing and splicing. In plant system, it has not been well characterized. Here we report the cloning and characterization of p68 from pea (Pisum sativum and its novel function in salinity stress tolerance in plant.The pea p68 protein self-interacts and is localized in the cytosol as well as the surrounding of cell nucleus. The transcript of pea p68 is upregulated in response to high salinity stress in pea. Overexpression of p68 driven by constitutive cauliflower mosaic virus-35S promoter in tobacco transgenic plants confers enhanced tolerances to salinity stress by improving the growth, photosynthesis and antioxidant machinery. Under stress treatment, pea p68 overexpressing tobacco accumulated higher K+ and lower Na+ level than the wild-type plants. Reactive oxygen species (ROS accumulation was remarkably regulated by the overexpression of pea p68 under salinity stress conditions, as shown from TBARS content, electrolyte leakage, hydrogen peroxide accumulation and 8-OHdG content and antioxidant enzyme activities.To the best of our knowledge this is the first direct report, which provides the novel function of pea p68 helicase in salinity stress tolerance. The results suggest that p68 can also be exploited for engineering abiotic stress tolerance in crop plants of economic importance.

  4. Using quantitative real-time PCR to detect chimeras in transgenic tobacco and apricot and to monitor their dissociation

    Directory of Open Access Journals (Sweden)

    Burgos Lorenzo

    2010-07-01

    Full Text Available Abstract Background The routine generation of transgenic plants involves analysis of transgene integration into the host genome by means of Southern blotting. However, this technique cannot distinguish between uniformly transformed tissues and the presence of a mixture of transgenic and non-transgenic cells in the same tissue. On the other hand, the use of reporter genes often fails to accurately detect chimerical tissues because their expression can be affected by several factors, including gene silencing and plant development. So, new approaches based on the quantification of the amount of the transgene are needed urgently. Results We show here that chimeras are a very frequent phenomenon observed after regenerating transgenic plants. Spatial and temporal analyses of transformed tobacco and apricot plants with a quantitative, real-time PCR amplification of the neomycin phosphotransferase (nptII transgene as well as of an internal control (β-actin, used to normalise the amount of target DNA at each reaction, allowed detection of chimeras at unexpected rates. The amount of the nptII transgene differed greatly along with the sub-cultivation period of these plants and was dependent on the localisation of the analysed leaves; being higher in roots and basal leaves, while in the apical leaves it remained at lower levels. These data demonstrate that, unlike the use of the gus marker gene, real-time PCR is a powerful tool for detection of chimeras. Although some authors have proposed a consistent, positive Southern analysis as an alternative methodology for monitoring the dissociation of chimeras, our data show that it does not provide enough proof of uniform transformation. In this work, however, real-time PCR was applied successfully to monitor the dissociation of chimeras in tobacco plants and apricot callus. Conclusions We have developed a rapid and reliable method to detect and estimate the level of chimeras in transgenic tobacco and apricot

  5. The strawberry FaMYB1 transcription factor suppresses anthocyanin and flavonol accumulation in transgenic tobacco.

    Science.gov (United States)

    Aharoni, A; De Vos, C H; Wein, M; Sun, Z; Greco, R; Kroon, A; Mol, J N; O'Connell, A P

    2001-11-01

    Fruit ripening is characterized by dramatic changes in gene expression, enzymatic activities and metabolism. Although the process of ripening has been studied extensively, we still lack valuable information on how the numerous metabolic pathways are regulated and co-ordinated. In this paper we describe the characterization of FaMYB1, a ripening regulated strawberry gene member of the MYB family of transcription factors. Flowers of transgenic tobacco lines overexpressing FaMYB1 showed a severe reduction in pigmentation. A reduction in the level of cyanidin 3-rutinoside (an anthocyanin) and of quercetin-glycosides (flavonols) was observed. Expression of late flavonoid biosynthesis genes and their enzyme activities were adversely affected by FaMYB1 overexpression. Two-hybrid assays in yeast showed that FaMYB1 could interact with other known anthocyanin regulators, but it does not act as a transcriptional activator. Interestingly, the C-terminus of FaMYB1 contains the motif pdLNL(D)/(E)Lxi(G)/S. This motif is contained in a region recently proposed to be involved in the repression of transcription by AtMYB4, an Arabidopsis MYB protein. Our results suggest that FaMYB1 may play a key role in regulating the biosynthesis of anthocyanins and flavonols in strawberry. It may act to repress transcription in order to balance the levels of anthocyanin pigments produced at the latter stages of strawberry fruit maturation, and/or to regulate metabolite levels in various branches of the flavonoid biosynthetic pathway.

  6. Metallothionein-I overexpression decreases brain pathology in transgenic mice with astrocyte-targeted expression of interleukin-6

    DEFF Research Database (Denmark)

    Molinero, Amalia; Penkowa, Milena; Hernández, Joaquín

    2003-01-01

    in this report support the idea that the upregulation of MT-I observed in GFAP-IL6 mice is an important mechanism for coping with brain damage. Thus, GFAP-IL6 mice that were crossed with TgMTI transgenic mice (GFAP-IL6xTgMTI) and overexpressed MT-I in the brain showed a decreased upregulation of cytokines...... such as IL-6 and a diminished recruitment and activation of macrophages and T cells throughout the CNS but mainly in the cerebellum. The GFAP-IL6 mice showed clear evidence of increased oxidative stress, which was significantly decreased by MT-I overexpression. Interestingly, MT-I overexpression increased...

  7. FHL1 reduces dystrophy in transgenic mice overexpressing FSHD muscular dystrophy region gene 1 (FRG1.

    Directory of Open Access Journals (Sweden)

    Sandra J Feeney

    Full Text Available Facioscapulohumeral muscular dystrophy (FSHD is an autosomal-dominant disease with no effective treatment. The genetic cause of FSHD is complex and the primary pathogenic insult underlying the muscle disease is unknown. Several disease candidate genes have been proposed including DUX4 and FRG1. Expression analysis studies of FSHD report the deregulation of genes which mediate myoblast differentiation and fusion. Transgenic mice overexpressing FRG1 recapitulate the FSHD muscular dystrophy phenotype. Our current study selectively examines how increased expression of FRG1 may contribute to myoblast differentiation defects. We generated stable C2C12 cell lines overexpressing FRG1, which exhibited a myoblast fusion defect upon differentiation. To determine if myoblast fusion defects contribute to the FRG1 mouse dystrophic phenotype, this strain was crossed with skeletal muscle specific FHL1-transgenic mice. We previously reported that FHL1 promotes myoblast fusion in vitro and FHL1-transgenic mice develop skeletal muscle hypertrophy. In the current study, FRG1 mice overexpressing FHL1 showed an improvement in the dystrophic phenotype, including a reduced spinal kyphosis, increased muscle mass and myofiber size, and decreased muscle fibrosis. FHL1 expression in FRG1 mice, did not alter satellite cell number or activation, but enhanced myoblast fusion. Primary myoblasts isolated from FRG1 mice showed a myoblast fusion defect that was rescued by FHL1 expression. Therefore, increased FRG1 expression may contribute to a muscular dystrophy phenotype resembling FSHD by impairing myoblast fusion, a defect that can be rescued by enhanced myoblast fusion via expression of FHL1.

  8. Overexpression of GmDREB1 improves salt tolerance in transgenic wheat and leaf protein response to high salinity

    OpenAIRE

    Qiyan Jiang; Zheng Hu; Hui Zhang; Youzhi Ma

    2014-01-01

    The transcription factor dehydration-responsive element binding protein (DREB) is able to improve tolerance to abiotic stress in plants by regulating the expression of downstream genes involved in environmental stress resistance. The objectives of this study were to evaluate the salt tolerance of GmDREB1 transgenic wheat (Triticum aestivum L.) and to evaluate its physiological and protein responses to salt stress. Compared with the wild type, the transgenic lines overexpressing GmDREB1 showed...

  9. Overexpression of wheat lipid transfer protein gene TaLTP5 increases resistances to Cochliobolus sativus and Fusarium graminearum in transgenic wheat.

    Science.gov (United States)

    Zhu, Xiuliang; Li, Zhao; Xu, Huijun; Zhou, Miaoping; Du, Lipu; Zhang, Zengyan

    2012-08-01

    The fungus Cochliobolus sativus is the main pathogen of common root rot, a serious soil-borne disease of wheat (Triticum aestivum L.). The fungus Fusarium graminearum is the primary pathogen of Fusarium head blight, a devastating disease of wheat worldwide. In this study, the wheat lipid transfer protein gene, TaLTP5, was cloned and evaluated for its ability to suppress disease development in transgenic wheat. TaLTP5 expression was induced after C. sativus infection. The TaLTP5 expression vector, pA25-TaLTP5, was constructed and bombarded into Chinese wheat variety Yangmai 18. Six TaLTP5 transgenic wheat lines were established and characterized. PCR and Southern blot analyses indicated that the introduced TaLTP5 gene was integrated into the genomes of six transgenic wheat lines by distinct patterns, and heritable. RT-PCR and real-time quantitative RT-PCR revealed that the TaLTP5 gene was over-expressed in the transgenic wheat lines compared to segregants lacking the transgene and wild-type wheat plants. Following challenge with C. sativus or F. graminearum, all six transgenic lines overexpressing TaLTP5 exhibited significantly enhanced resistance to both common root rot and Fusarium head blight compared to the untransformed wheat Yangmai 18.

  10. Generation of Resistance to the Diphenyl Ether Herbicide, Oxyfluorfen, via Expression of the Bacillus subtilis Protoporphyrinogen Oxidase Gene in Transgenic Tobacco Plants.

    Science.gov (United States)

    Choi, K W; Han, O; Lee, H J; Yun, Y C; Moon, Y H; Kim, M; Kuk, Y I; Han, S U; Guh, J O

    1998-01-01

    In an effort to develop transgenic plants resistant to diphenyl ether herbicides, we introduced the protoporphyrinogen oxidase (EC 1.3.3.4) gene of Bacillus subtilis into tobacco plants. The results from a Northern analysis and leaf disc assay indicate that the expression of the B. subtilis protoporphyrinogen oxidase gene under the cauliflower mosaic virus 35S promoter generated resistance to the diphenyl ether herbicide, oxyfluorfen, in transgenic tobacco plants.

  11. Cloning of two individual cDNAS encoding 9-cis-epoxycarotenoid dioxygenase from Gentiana lutea, their tissue-specific expression and physiological effect in transgenic tobacco.

    Science.gov (United States)

    Zhu, Changfu; Kauder, Friedrich; Römer, Susanne; Sandmann, Gerhard

    2007-02-01

    Two 9-cis-epoxycarotenoid dioxygenase (NCED) cDNAs have been cloned from a petal library of Gentiana lutea. Both cDNAs carry a putative transit sequence for chloroplast import and differ mainly in their length and the 5'-flanking regions. GlNCED1 was evolutionary closely related to Arabidopsis thaliana NCED6 whereas GlNCED2 showed highest homology to tomato NCED1 and A. thaliana NCED3. The amounts of GlNCED2 transcript were below Northern detection in G. lutea. In contrast, GlNCED1 was specifically expressed at higher levels in developing flowers when petals start appearing. By genetic engineering of tobacco with coding regions of either gene under a constitutive promoter, their function was further analyzed. Although mRNA of both genes was detectable in the corresponding transgenic plants, a physiological effect was only found for GlNCED1 but not for GlNCED2. In germination experiments of GlNCED1 transgenic lines, delayed radicle formation and cotyledon appearance were observed. However, the transformants exhibited no improved tolerance against desiccation stress. In contrast to other plants with over-expressed NCEDs, prolonged delay of seed germination is the only abscisic-acid-related phenotypic effect in the GlNCED1 transgenic lines.

  12. Transgenic overexpression of adenine nucleotide translocase 1 protects ischemic hearts against oxidative stress.

    Science.gov (United States)

    Klumpe, Inga; Savvatis, Konstantinos; Westermann, Dirk; Tschöpe, Carsten; Rauch, Ursula; Landmesser, Ulf; Schultheiss, Heinz-Peter; Dörner, Andrea

    2016-06-01

    Ischemia impairs the adenine nucleotide translocase (ANT), which transports ADP and ATP across the inner mitochondrial membrane. We investigated whether ANT1 overexpression has protective effects on ischemic hearts. Myocardial infarction was induced in wild-type (WT) and heart-specific ANT1-transgenic (ANT1-TG) rats, and hypoxia was set in isolated cardiomyocytes. ANT1 overexpression reduced the myocardial infarct area and increased the survival rate of infarcted rats. Reduced ANT1 expression and increased 4-hydroxynonenal modification of ANT paralleled to impaired ANT function in infarcted WT hearts. ANT1 overexpression improved ANT expression and function. This was accompanied by reduced mitochondrial cytochrome C release and caspase-3 activation. ANT1-TG hearts suffered less from oxidative stress, as shown by lower protein carbonylation and 4-hydroxynonenal modification of ANT. ANT1 overexpression also increased cell survival of hypoxic cardiomyocytes and attenuated reactive oxygen species (ROS) production. This was linked to higher stability of mitochondrial membrane potential and lower activity of ROS detoxifying catalase. ANT1-TG cardiomyocytes also showed higher resistance against H2O2 treatment, which was independent of catalase activity. In conclusion, ANT1 overexpression compensates impaired ANT activity under oxygen-restricted conditions. It reduces ROS production and oxidative stress, stabilizes mitochondrial integrity, and increases survival, making ANT1 a component in ROS management and heart protection during ischemia. ANT1 overexpression reduces infarct size and increases survival after infarction. ANT1 overexpression compensates restricted ANT expression and function in infarcted hearts. Increased ANT1 expression enhances mitochondrial integrity. ANT1-overexpressing hearts reduce oxidative stress by decreasing ROS generation. ANT1 is a component in ROS management and heart protection.

  13. Overexpression of IGF-I in skeletal muscle of transgenic mice does not prevent unloading-induced atrophy

    Science.gov (United States)

    Criswell, D. S.; Booth, F. W.; DeMayo, F.; Schwartz, R. J.; Gordon, S. E.; Fiorotto, M. L.

    1998-01-01

    This study examined the association between local insulin-like growth factor I (IGF-I) overexpression and atrophy in skeletal muscle. We hypothesized that endogenous skeletal muscle IGF-I mRNA expression would decrease with hindlimb unloading (HU) in mice, and that transgenic mice overexpressing human IGF-I (hIGF-I) specifically in skeletal muscle would exhibit less atrophy after HU. Male transgenic mice and nontransgenic mice from the parent strain (FVB) were divided into four groups (n = 10/group): 1) transgenic, weight-bearing (IGF-I/WB); 2) transgenic, hindlimb unloaded (IGF-I/HU); 3) nontransgenic, weight-bearing (FVB/WB); and 4) nontransgenic, hindlimb unloaded (FVB/HU). HU groups were hindlimb unloaded for 14 days. Body mass was reduced (P < 0.05) after HU in both IGF-I (-9%) and FVB mice (-13%). Contrary to our hypothesis, we found that the relative abundance of mRNA for the endogenous rodent IGF-I (rIGF-I) was unaltered by HU in the gastrocnemius (GAST) muscle of wild-type FVB mice. High-level expression of hIGF-I peptide and mRNA was confirmed in the GAST and tibialis anterior (TA) muscles of the transgenic mice. Nevertheless, masses of the GAST and TA muscles were reduced (P < 0.05) in both FVB/HU and IGF-I/HU groups compared with FVB/WB and IGF-I/WB groups, respectively, and the percent atrophy in mass of these muscles did not differ between FVB and IGF-I mice. Therefore, skeletal muscle atrophy may not be associated with a reduction of endogenous rIGF-I mRNA level in 14-day HU mice. We conclude that high local expression of hIGF-I mRNA and peptide in skeletal muscle alone cannot attenuate unloading-induced atrophy of fast-twitch muscle in mice.

  14. Transgenic overexpression of NanogP8 in the mouse prostate is insufficient to initiate tumorigenesis but weakly promotes tumor development in the Hi-Myc mouse model.

    Science.gov (United States)

    Liu, Bigang; Gong, Shuai; Li, Qiuhui; Chen, Xin; Moore, John; Suraneni, Mahipal V; Badeaux, Mark D; Jeter, Collene R; Shen, Jianjun; Mehmood, Rashid; Fan, Qingxia; Tang, Dean G

    2017-08-08

    This project was undertaken to address a critical cancer biology question: Is overexpression of the pluripotency molecule Nanog sufficient to initiate tumor development in a somatic tissue? Nanog1 is critical for the self-renewal and pluripotency of ES cells, and its retrotransposed homolog, NanogP8 is preferentially expressed in somatic cancer cells. Our work has shown that shRNA-mediated knockdown of NanogP8 in prostate, breast, and colon cancer cells inhibits tumor regeneration whereas inducible overexpression of NanogP8 promotes cancer stem cell phenotypes and properties. To address the key unanswered question whether tissue-specific overexpression of NanogP8 is sufficient to promote tumor development in vivo , we generated a NanogP8 transgenic mouse model, in which the ARR 2 PB promoter was used to drive NanogP8 cDNA. Surprisingly, the ARR 2 PB-NanogP8 transgenic mice were viable, developed normally, and did not form spontaneous tumors in >2 years. Also, both wild type and ARR 2 PB-NanogP8 transgenic mice responded similarly to castration and regeneration and castrated ARR 2 PB-NanogP8 transgenic mice also did not develop tumors. By crossing the ARR 2 PB-NanogP8 transgenic mice with ARR 2 PB-Myc (i.e., Hi-Myc) mice, we found that the double transgenic (i.e., ARR 2 PB-NanogP8; Hi-Myc) mice showed similar tumor incidence and histology to the Hi-Myc mice. Interestingly, however, we observed white dots in the ventral lobes of the double transgenic prostates, which were characterized as overgrown ductules/buds featured by crowded atypical Nanog-expressing luminal cells. Taken together, our present work demonstrates that transgenic overexpression of NanogP8 in the mouse prostate is insufficient to initiate tumorigenesis but weakly promotes tumor development in the Hi-Myc mouse model.

  15. Inheritance and effectiveness of two transgenes determining PVY resistance in progeny from crossing independently transformed tobacco lines.

    Science.gov (United States)

    Czubacka, Anna; Sacco, Ermanno; Olszak-Przybyś, Hanna; Doroszewska, Teresa

    2017-05-01

    Genetic transformation of plants allows us to obtain improved genotypes enriched with the desired traits. However, if transgenic lines were to be used in breeding programs the stability of inserted transgenes is essential. In the present study, we followed the inheritance of transgenes in hybrids originated from crossing two transgenic tobacco lines resistant to Potato virus Y (PVY): MN 944 LMV with the transgene containing Lettuce mosaic virus coat protein gene (LMV CP) and AC Gayed ROKY2 with PVY replicase gene (ROKY2). Progeny populations generated by successive self-pollination were analyzed with respect to the transgene segregation ratio and resistance to Potato virus Y in tests carried out under greenhouse conditions. The presence of the virus in inoculated plants was detected by DAS-ELISA method. The results demonstrated the Mendelian fashion of inheritance of transgenes which were segregated independently and stably. As a result, we obtained T 4 generation of hybrid with both transgenes stacked and which was highly resistant to PVY.

  16. Transgenic tomato plants overexpressing tyramine N-hydroxycinnamoyltransferase exhibit elevated hydroxycinnamic acid amide levels and enhanced resistance to Pseudomonas syringae.

    Science.gov (United States)

    Campos, Laura; Lisón, Purificación; López-Gresa, María Pilar; Rodrigo, Ismael; Zacarés, Laura; Conejero, Vicente; Bellés, José María

    2014-10-01

    Hydroxycinnamic acid amides (HCAA) are secondary metabolites involved in plant development and defense that have been widely reported throughout the plant kingdom. These phenolics show antioxidant, antiviral, antibacterial, and antifungal activities. Hydroxycinnamoyl-CoA:tyramine N-hydroxycinnamoyl transferase (THT) is the key enzyme in HCAA synthesis and is induced in response to pathogen infection, wounding, or elicitor treatments, preceding HCAA accumulation. We have engineered transgenic tomato plants overexpressing tomato THT. These plants displayed an enhanced THT gene expression in leaves as compared with wild type (WT) plants. Consequently, leaves of THT-overexpressing plants showed a higher constitutive accumulation of the amide coumaroyltyramine (CT). Similar results were found in flowers and fruits. Moreover, feruloyltyramine (FT) also accumulated in these tissues, being present at higher levels in transgenic plants. Accumulation of CT, FT and octopamine, and noradrenaline HCAA in response to Pseudomonas syringae pv. tomato infection was higher in transgenic plants than in the WT plants. Transgenic plants showed an enhanced resistance to the bacterial infection. In addition, this HCAA accumulation was accompanied by an increase in salicylic acid levels and pathogenesis-related gene induction. Taken together, these results suggest that HCAA may play an important role in the defense of tomato plants against P. syringae infection.

  17. Characterization and subcellular compartmentation of recombinant 4-hydroxyphenylpyruvate dioxygenase from Arabidopsis in transgenic tobacco.

    Science.gov (United States)

    Garcia, I; Rodgers, M; Pepin, R; Hsieh, T F; Matringe, M

    1999-04-01

    4-Hydroxyphenylpyruvate dioxygenase (4HPPD) catalyzes the formation of homogentisate (2,5-dihydroxyphenylacetate) from p-hydroxyphenylpyruvate and molecular oxygen. In plants this enzyme activity is involved in two distinct metabolic processes, the biosynthesis of prenylquinones and the catabolism of tyrosine. We report here the molecular and biochemical characterization of an Arabidopsis 4HPPD and the compartmentation of the recombinant protein in chlorophyllous tissues. We isolated a 1508-bp cDNA with one large open reading frame of 1338 bp. Southern analysis strongly suggested that this Arabidopsis 4HPPD is encoded by a single-copy gene. We investigated the biochemical characteristics of this 4HPPD by overproducing the recombinant protein in Escherichia coli JM105. The subcellular localization of the recombinant 4HPPD in chlorophyllous tissues was examined by overexpressing its complete coding sequence in transgenic tobacco (Nicotiana tabacum), using Agrobacterium tumefaciens transformation. We performed western analyses for the immunodetection of protein extracts from purified chloroplasts and total leaf extracts and for the immunocytochemistry on tissue sections. These analyses clearly revealed that 4HPPD was confined to the cytosol compartment, not targeted to the chloroplast. Western analyses confirmed the presence of a cytosolic form of 4HPPD in cultured green Arabidopsis cells.

  18. Mammary gland tumor formation in transgenic mice overexpressing stromelysin-1

    Energy Technology Data Exchange (ETDEWEB)

    Sympson, Carolyn J; Bissell, Mina J; Werb, Zena

    1995-06-01

    An intact basement membrane (BM) is essential for the proper function, differentiation and morphology of many epithelial cells. The disruption or loss of this BM occurs during normal development as well as in the disease state. To examine the importance of BM during mammary gland development in vivo, we generated transgenic mice that inappropriately express autoactivating isoforms of the matrix metalloproteinase stromelysin-1. The mammary glands from these mice are both functionally and morphologically altered throughout development. We have now documented a dramatic incidence of breast tumors in several independent lines of these mice. These data suggest that overexpression of stromelysin-1 and disruption of the BM may be a key step in the multi-step process of breast cancer.

  19. Processing, Targeting, and Antifungal Activity of Stinging Nettle Agglutinin in Transgenic Tobacco

    Science.gov (United States)

    Does, Mirjam P.; Houterman, Petra M.; Dekker, Henk L.; Cornelissen, Ben J.C.

    1999-01-01

    The gene encoding the precursor to stinging nettle (Urtica dioica L.) isolectin I was introduced into tobacco (Nicotiana tabacum). In transgenic plants this precursor was processed to mature-sized lectin. The mature isolectin is deposited intracellularly, most likely in the vacuoles. A gene construct lacking the C-terminal 25 amino acids was also introduced in tobacco to study the role of the C terminus in subcellular trafficking. In tobacco plants that expressed this construct, the mutant precursor was correctly processed and the mature isolectin was targeted to the intercellular space. These results indicate the presence of a C-terminal signal for intracellular retention of stinging nettle lectin and most likely for sorting of the lectin to the vacuoles. In addition, correct processing of this lectin did not depend on vacuolar deposition. Isolectin I purified from tobacco displayed identical biological activities as isolectin I isolated from stinging nettle. In vitro antifungal assays on germinated spores of the fungi Botrytis cinerea, Trichoderma viride, and Colletotrichum lindemuthianum revealed that growth inhibition by stinging nettle isolectin I occurs at a specific phase of fungal growth and is temporal, suggesting that the fungi had an adaptation mechanism. PMID:10364393

  20. A ThDREB gene from Tamarix hispida improved the salt and drought tolerance of transgenic tobacco and T. hispida.

    Science.gov (United States)

    Yang, Guiyan; Yu, Lili; Zhang, Kaimin; Zhao, Yulin; Guo, Yucong; Gao, Caiqiu

    2017-04-01

    Dehydration-responsive element-binding (DREB) transcription factors are important abiotic stress tolerance related genes, and some reports on the roles of DREB have primarily addressed herbal plants. To explore the abiotic stress tolerance role of DREB (ThDREB) from Tamarix hispida, a ThDREB gene with a complete ORF of 783 bp that encodes a 28.74 kDa protein with 260 amino acids, was isolated and functionally annotated. ThDREB expression was highly induced by NaCl, PEG, NaHCO 3 and CdCl 2 treatments, and the highest expression level (369.2-fold of control) was found for the roots that were under NaCl stress for 6 h. The tobacco plants that were transformed by ThDREB were conferred with higher germination rates, fresh weights and root lengths than the wild type (WT) tobacco plants under NaCl and mannitol treatments. The total chlorophyll content (tcc), superoxide dismutase (SOD) and peroxidase (POD) activities were also higher in the transgenic lines in comparison with the WT, and the malondialdehyde (MDA) and H 2 O 2 content, electrolyte leakage (EL) rate and ROS as tracked by staining were generated to a lesser degree in ThDREB transgenic plants than in the WT under NaCl and mannitol stress. Furthermore, the transient overexpression analysis of ThDREB in T. hispida also improved plant salt and drought tolerance in comparison with the empty vector-transformed lines. Our results indicated that ThDREB expression could effectively improve tolerance to salt and drought stress by enhancing the antioxidase activity that keeps the ROS at a low accumulation level and makes them easy to scavenge. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  1. Improved antioxidant activity in transgenic Perilla frutescens plants via overexpression of the γ-tocopherol methyltransferase (γ-tmt) gene.

    Science.gov (United States)

    Ghimire, Bimal Kumar; Seong, Eun Soo; Lee, Chan Ok; Lee, Jae Geun; Yu, Chang Yeon; Kim, Seung Hyun; Chung, Ill Min

    2015-09-01

    The main goal of this study was to generate transgenic Perilla frutescens with enhanced antioxidant properties by overexpressing the γ-tocopherol methyltransferase (γ-tmt) gene. In this study, the antioxidant activity of methanolic crude extracts of transgenic and non-transgenic control plants was investigated using the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging method. Free radical scavenging activity was evaluated using α-tocopherol and butylated hydroxyl toluene as standard antioxidants. In general, the ethyl acetate fraction of transgenic P. frutescens showed stronger DPPH radical scavenging activity than the ethyl acetate fraction from non-transgenic control plants (IC50 2.00 ± 0.10 and 5.53 ± 0.40 μg ∙ ml(-1), respectively). High-performance liquid chromatography analysis of phenolic acids in leaf extracts confirmed increased levels of 16 individual phenolic compounds in two transgenic lines (pf47-5 and pf47-8) compared with control plants. Changes in the phenolic compound profile and α-tocopherol content were correlated with the antioxidant properties of transgenic plants, indicating that the introduction of transgene γ-tmt influenced the metabolism of phenolic compounds and subsequently produced biochemical changes in the transformants. There were no significant differences in photosynthetic rate in the transgenic plants as compared to the non-transgenic control plants, suggesting that the alteration of phenolic compounds and tocopherol composition had little impact on photosynthesis.

  2. Transgenic banana plants overexpressing a native plasma membrane aquaporin MusaPIP1;2 display high tolerance levels to different abiotic stresses.

    Science.gov (United States)

    Sreedharan, Shareena; Shekhawat, Upendra K S; Ganapathi, Thumballi R

    2013-10-01

    Water transport across cellular membranes is regulated by a family of water channel proteins known as aquaporins (AQPs). As most abiotic stresses like suboptimal temperatures, drought or salinity result in cellular dehydration, it is imperative to study the cause-effect relationship between AQPs and the cellular consequences of abiotic stress stimuli. Although plant cells have a high isoform diversity of AQPs, the individual and integrated roles of individual AQPs in optimal and suboptimal physiological conditions remain unclear. Herein, we have identified a plasma membrane intrinsic protein gene (MusaPIP1;2) from banana and characterized it by overexpression in transgenic banana plants. Cellular localization assay performed using MusaPIP1;2::GFP fusion protein indicated that MusaPIP1;2 translocated to plasma membrane in transformed banana cells. Transgenic banana plants overexpressing MusaPIP1;2 constitutively displayed better abiotic stress survival characteristics. The transgenic lines had lower malondialdehyde levels, elevated proline and relative water content and higher photosynthetic efficiency as compared to equivalent controls under different abiotic stress conditions. Greenhouse-maintained hardened transgenic plants showed faster recovery towards normal growth and development after cessation of abiotic stress stimuli, thereby underlining the importance of these plants in actual environmental conditions wherein the stress stimuli is often transient but severe. Further, transgenic plants where the overexpression of MusaPIP1;2 was made conditional by tagging it with a stress-inducible native dehydrin promoter also showed similar stress tolerance characteristics in in vitro and in vivo assays. Plants developed in this study could potentially enable banana cultivation in areas where adverse environmental conditions hitherto preclude commercial banana cultivation. © 2013 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons

  3. Multiple different defense mechanisms are activated in the young transgenic tobacco plants which express the full length genome of the Tobacco mosaic virus, and are resistant against this virus.

    Science.gov (United States)

    Jada, Balaji; Soitamo, Arto J; Siddiqui, Shahid Aslam; Murukesan, Gayatri; Aro, Eva-Mari; Salakoski, Tapio; Lehto, Kirsi

    2014-01-01

    Previously described transgenic tobacco lines express the full length infectious Tobacco mosaic virus (TMV) genome under the 35S promoter (Siddiqui et al., 2007. Mol Plant Microbe Interact, 20: 1489-1494). Through their young stages these plants exhibit strong resistance against both the endogenously expressed and exogenously inoculated TMV, but at the age of about 7-8 weeks they break into TMV infection, with typical severe virus symptoms. Infections with some other viruses (Potato viruses Y, A, and X) induce the breaking of the TMV resistance and lead to synergistic proliferation of both viruses. To deduce the gene functions related to this early resistance, we have performed microarray analysis of the transgenic plants during the early resistant stage, and after the resistance break, and also of TMV-infected wild type tobacco plants. Comparison of these transcriptomes to those of corresponding wild type healthy plants indicated that 1362, 1150 and 550 transcripts were up-regulated in the transgenic plants before and after the resistance break, and in the TMV-infected wild type tobacco plants, respectively, and 1422, 1200 and 480 transcripts were down-regulated in these plants, respectively. These transcriptome alterations were distinctly different between the three types of plants, and it appears that several different mechanisms, such as the enhanced expression of the defense, hormone signaling and protein degradation pathways contributed to the TMV-resistance in the young transgenic plants. In addition to these alterations, we also observed a distinct and unique gene expression alteration in these plants, which was the strong suppression of the translational machinery. This may also contribute to the resistance by slowing down the synthesis of viral proteins. Viral replication potential may also be suppressed, to some extent, by the reduction of the translation initiation and elongation factors eIF-3 and eEF1A and B, which are required for the TMV replication

  4. Overexpression of PSP1 enhances growth of transgenic Arabidopsis plants under ambient air conditions.

    Science.gov (United States)

    Han, Xiaofang; Peng, Keli; Wu, Haixia; Song, Shanshan; Zhu, Yerong; Bai, Yanling; Wang, Yong

    2017-07-01

    The importance of the phosphorylated pathway (PPSB) of L-serine (Ser) biosynthesis in plant growth and development has been demonstrated, but its specific role in leaves and interaction with photorespiration, the main leaf Ser biosynthetic pathway at daytime, are still unclear. To investigate whether changes in biosynthesis of Ser by the PPSB in leaves could have an impact on photorespiration and plant growth, we overexpressed PSP1, the last enzyme of this pathway, under control of the Cauliflower Mosaic Virus 35S promoter in Arabidopsis thaliana. Overexpressor plants grown in normal air displayed larger rosette diameter and leaf area as well as higher fresh and dry weight than the wild type. By contrast, no statistically significant differences to the wild type were observed when the overexpressor seedlings were transferred to elevated CO 2 , indicating a relationship between PSP1 overexpression and photorespiration. Additionally, the transgenic plants displayed higher photorespiration, an increase in CO 2 net-uptake and stronger expression in the light of genes encoding enzymes involved in photorespiration. We further demonstrated that expression of many genes involved in nitrogen assimilation was also promoted in leaves of transgenic plants and that leaf nitrate reductase activity increased in the light, too, although not in the dark. Our results suggest a close correlation between the function of PPSB and photorespiration, and also nitrogen metabolism in leaves.

  5. Resistance to BmNPV via overexpression of an exogenous gene controlled by an inducible promoter and enhancer in transgenic silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Liang Jiang

    Full Text Available The hycu-ep32 gene of Hyphantria cunea NPV can inhibit Bombyx mori nucleopolyhedrovirus (BmNPV multiplication in co-infected cells, but it is not known whether the overexpression of the hycu-ep32 gene has an antiviral effect in the silkworm, Bombyx mori. Thus, we constructed four transgenic vectors, which were under the control of the 39 K promoter of BmNPV (39 KP, Bombyx mori A4 promoter (A4P, hr3 enhancer of BmNPV combined with 39 KP, and hr3 combined with A4P. Transgenic lines were created via embryo microinjection using practical diapause silkworm. qPCR revealed that the expression level of hycu-ep32 could be induced effectively after BmNPV infection in transgenic lines where hycu-ep32 was controlled by hr3 combined with 39 KP (i.e., HEKG. After oral inoculation of BmNPV with 3 × 10(5 occlusion bodies per third instar, the mortality with HEKG-B was approximately 30% lower compared with the non-transgenic line. The economic characteristics of the transgenic lines remained unchanged. These results suggest that overexpression of an exogenous antiviral gene controlled by an inducible promoter and enhancer is a feasible method for breeding silkworms with a high antiviral capacity.

  6. Ectopic expression of phloem motor protein pea forisome PsSEO-F1 enhances salinity stress tolerance in tobacco.

    Science.gov (United States)

    Srivastava, Vineet Kumar; Raikwar, Shailendra; Tuteja, Renu; Tuteja, Narendra

    2016-05-01

    PsSEOF-1 binds to calcium and its expression is upregulated by salinity treatment. PsSEOF - 1 -overexpressing transgenic tobacco showed enhanced salinity stress tolerance by maintaining cellular ion homeostasis and modulating ROS-scavenging pathway. Calcium (Ca(2+)) plays important role in growth, development and stress tolerance in plants. Cellular Ca(2+) homeostasis is achieved by the collective action of channels, pumps, antiporters and by Ca(2+) chelators present in the cell like calcium-binding proteins. Forisomes are ATP-independent mechanically active motor proteins known to function in wound sealing of injured sieve elements of phloem tissue. The Ca(2+)-binding activity of forisome and its role in abiotic stress signaling were largely unknown. Here we report the Ca(2+)-binding activity of pea forisome (PsSEO-F1) and its novel function in promoting salinity tolerance in transgenic tobacco. Native PsSEO-F1 promoter positively responded in salinity stress as confirmed using GUS reporter. Overexpression of PsSEO-F1 tobacco plants confers salinity tolerance by alleviating ionic toxicity and increased ROS scavenging activity which probably results in reduced membrane damage and improved yield under salinity stress. Evaluation of several physiological indices shows an increase in relative water content, electrolyte leakage, proline accumulation and chlorophyll content in transgenic lines as compared with null-segregant control. Expression of several genes involved in cellular homeostasis is perturbed by PsSEO-F1 overexpression. These findings suggest that PsSEO-F1 provides salinity tolerance through cellular Ca(2+) homeostasis which in turn modulates ROS machinery providing indirect link between Ca(2+) and ROS signaling under salinity-induced perturbation. PsSEO-F1 most likely functions in salinity stress tolerance by improving antioxidant machinery and mitigating ion toxicity in transgenic lines. This finding should make an important contribution in our better

  7. Targeted Transgenic Overexpression of Mitochondrial Thymidine Kinase (TK2) Alters Mitochondrial DNA (mtDNA) and Mitochondrial Polypeptide Abundance : Transgenic TK2, mtDNA, and Antiretrovirals

    OpenAIRE

    Hosseini, Seyed H.; Kohler, James J.; Haase, Chad P.; Tioleco, Nina; Stuart, Tami; Keebaugh, Erin; Ludaway, Tomika; Russ, Rodney; Green, Elgin; Long, Robert; Wang, Liya; Eriksson, Staffan; Lewis, William

    2007-01-01

    Mitochondrial toxicity limits nucleoside reverse transcriptase inhibitors (NRTIs) for acquired immune deficiency syndrome. NRTI triphosphates, the active moieties, inhibit human immunodeficiency virus reverse transcriptase and eukaryotic mitochondrial DNA polymerase pol-γ. NRTI phosphorylation seems to correlate with mitochondrial toxicity, but experimental evidence is lacking. Transgenic mice (TGs) with cardiac overexpression of thymidine kinase isoforms (mitochondrial TK2 and cytoplasmic TK...

  8. Phytoremediation of arsenic from the contaminated soil using transgenic tobacco plants expressing ACR2 gene of Arabidopsis thaliana.

    Science.gov (United States)

    Nahar, Noor; Rahman, Aminur; Nawani, Neelu N; Ghosh, Sibdas; Mandal, Abul

    2017-11-01

    We have cloned, characterized and transformed the AtACR2 gene (arsenic reductase 2) of Arabidopsis thaliana into the genome of tobacco (Nicotiana tabacum, var Sumsun). Our results revealed that the transgenic tobacco plants are more tolerant to arsenic than the wild type ones. These plants can grow on culture medium containing 200μM arsenate, whereas the wild type can barely survive under this condition. Furthermore, when exposed to 100μM arsenate for 35days the amount of arsenic accumulated in the shoots of transgenic plants was significantly lower (28μg/g d wt.) than that found in the shoots of non-transgenic controls (40μg/g d wt.). However, the arsenic content in the roots of transgenic plants was significantly higher (2400μg/g d. wt.) than that (2100μg/g d. wt.) observed in roots of wild type plants. We have demonstrated that Arabidopsis thaliana AtACR2 gene is a potential candidate for genetic engineering of plants to develop new crop cultivars that can be grown on arsenic contaminated fields to reduce arsenic content of the soil and can become a source of food containing no arsenic or exhibiting substantially reduced amount of this metalloid. Copyright © 2017 Elsevier GmbH. All rights reserved.

  9. Expression of a pathogen-induced cysteine protease (AdCP) in tapetum results in male sterility in transgenic tobacco.

    Science.gov (United States)

    Shukla, Pawan; Singh, Naveen Kumar; Kumar, Dilip; Vijayan, Sambasivam; Ahmed, Israr; Kirti, Pulugurtha Bharadwaja

    2014-06-01

    Usable male sterility systems have immense potential in developing hybrid varieties in crop plants, which can also be used as a biological safety containment to prevent horizontal transgene flow. Barnase-Barstar system developed earlier was the first approach to engineer male sterility in plants. In an analogous situation, we have evolved a system of inducing pollen abortion and male sterility in transgenic tobacco by expressing a plant gene coding for a protein with known developmental function in contrast to the Barnase-Barstar system, which deploys genes of prokaryotic origin, i.e., from Bacillus amyloliquefaciens. We have used a plant pathogen-induced gene, cysteine protease for inducing male sterility. This gene was identified in the wild peanut, Arachis diogoi differentially expressed when it was challenged with the late leaf spot pathogen, Phaeoisariopsis personata. Arachis diogoi cysteine protease (AdCP) was expressed under the strong tapetum-specific promoter (TA29) and tobacco transformants were generated. Morphological and histological analysis of AdCP transgenic plants showed ablated tapetum and complete pollen abortion in three transgenic lines. Furthermore, transcript analysis displayed the expression of cysteine protease in these male sterile lines and the expression of the protein was identified in western blot analysis using its polyclonal antibody raised in the rabbit system.

  10. Reduced grain chalkiness and its possible physiological mechanism in transgenic rice overexpressing l-GalLDH

    Directory of Open Access Journals (Sweden)

    Le Yu

    2015-04-01

    Full Text Available Chalkiness is one of the key factors determining rice quality and price. Ascorbic acid (Asc is a major plant antioxidant that performs many functions in plants. l-Galactono-1,4-lactone dehydrogenase (l-GalLDH, EC1.3.2.3 is an enzyme that catalyzes the final step of Asc biosynthesis in plants. Here we show that the l-GalLDH-overexpressing transgenic rice, GO-2, which has constitutively higher leaf Asc content than wild-type (WT plants, exhibits significantly reduced grain chalkiness. Higher foliar ascorbate/dehydroascorbate (Asc/DHA ratios at 40, 60, 80, and 100 days of plant age were observed in GO-2. Further investigation showed that the enhanced level of Asc resulted in a significantly higher ribulose-1,5-bisphosphate (RuBP carboxylase/oxygenase (Rubisco protein level in GO-2 at 80 days. In addition, levels of abscisic acid (ABA and jasmonic acid (JA were lower in GO-2 at 60, 80, and 100 days. The results we present here indicate that the enhanced level of Asc is likely responsible for changing redox homeostasis in key developmental stages associated with grain filling and alters grain chalkiness in the l-GalLDH-overexpressing transgenic by maintaining photosynthetic function and affecting phytohormones associated with grain filling.

  11. Over-expression of VvWRKY1 in grapevines induces expression of jasmonic acid pathway-related genes and confers higher tolerance to the downy mildew.

    Directory of Open Access Journals (Sweden)

    Chloé Marchive

    Full Text Available Most WRKY transcription factors activate expression of defence genes in a salicylic acid- and/or jasmonic acid-dependent signalling pathway. We previously identified a WRKY gene, VvWRKY1, which is able to enhance tolerance to fungal pathogens when it is overexpressed in tobacco. The present work analyzes the effects of VvWRKY1 overexpression in grapevine. Microarray analysis showed that genes encoding defence-related proteins were up-regulated in the leaves of transgenic 35S::VvWRKY1 grapevines. Quantitative RT-PCR analysis confirmed that three genes putatively involved in jasmonic acid signalling pathway were overexpressed in the transgenic grapes. The ability of VvWRKY1 to trans-activate the promoters of these genes was demonstrated by transient expression in grape protoplasts. The resistance to the causal agent of downy mildew, Plasmopara viticola, was enhanced in the transgenic plants. These results show that VvWRKY1 can increase resistance of grapevine against the downy mildew through transcriptional reprogramming leading to activation of the jasmonic acid signalling pathway.

  12. Overexpression of the Synthetic Chimeric Native-T-phylloplanin-GFP Genes Optimized for Monocot and Dicot Plants Renders Enhanced Resistance to Blue Mold Disease in Tobacco (N. tabacum L.

    Directory of Open Access Journals (Sweden)

    Dipak K. Sahoo

    2014-01-01

    Full Text Available To enhance the natural plant resistance and to evaluate the antimicrobial properties of phylloplanin against blue mold, we have expressed a synthetic chimeric native-phylloplanin-GFP protein fusion in transgenic Nicotiana tabacum cv. KY14, a cultivar that is highly susceptible to infection by Peronospora tabacina. The coding sequence of the tobacco phylloplanin gene along with its native signal peptide was fused with GFP at the carboxy terminus. The synthetic chimeric gene (native-phylloplanin-GFP was placed between the modified Mirabilis mosaic virus full-length transcript promoter with duplicated enhancer domains and the terminator sequence from the rbcSE9 gene. The chimeric gene, expressed in transgenic tobacco, was stably inherited in successive plant generations as shown by molecular characterization, GFP quantification, and confocal fluorescent microscopy. Transgenic plants were morphologically similar to wild-type plants and showed no deleterious effects due to transgene expression. Blue mold-sensitivity assays of tobacco lines were performed by applying P. tabacina sporangia to the upper leaf surface. Transgenic lines expressing the fused synthetic native-phyllopanin-GFP gene in the leaf apoplast showed resistance to infection. Our results demonstrate that in vivo expression of a synthetic fused native-phylloplanin-GFP gene in plants can potentially achieve natural protection against microbial plant pathogens, including P. tabacina in tobacco.

  13. Novel AroA from Pseudomonas putida Confers Tobacco Plant with High Tolerance to Glyphosate

    Science.gov (United States)

    Yan, Hai-Qin; Chang, Su-Hua; Tian, Zhe-Xian; Zhang, Le; Sun, Yi-Cheng; Li, Yan; Wang, Jing; Wang, Yi-Ping

    2011-01-01

    Glyphosate is a non-selective broad-spectrum herbicide that inhibits 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS, also designated as AroA), a key enzyme in the aromatic amino acid biosynthesis pathway in microorganisms and plants. Previously, we reported that a novel AroA (PpAroA1) from Pseudomonas putida had high tolerance to glyphosate, with little homology to class I or class II glyphosate-tolerant AroA. In this study, the coding sequence of PpAroA1 was optimized for tobacco. For maturation of the enzyme in chloroplast, a chloroplast transit peptide coding sequence was fused in frame with the optimized aroA gene (PparoA1optimized) at the 5′ end. The PparoA1optimized gene was introduced into the tobacco (Nicotiana tabacum L. cv. W38) genome via Agrobacterium-mediated transformation. The transformed explants were first screened in shoot induction medium containing kanamycin. Then glyphosate tolerance was assayed in putative transgenic plants and its T1 progeny. Our results show that the PpAroA1 from Pseudomonas putida can efficiently confer tobacco plants with high glyphosate tolerance. Transgenic tobacco overexpressing the PparoA1optimized gene exhibit high tolerance to glyphosate, which suggest that the novel PpAroA1 is a new and good candidate applied in transgenic crops with glyphosate tolerance in future. PMID:21611121

  14. Stable Skin-specific Overexpression of Human CTLA4-Ig in Transgenic Mice through Seven Generations

    Institute of Scientific and Technical Information of China (English)

    Yong WANG; Yong NI; Hong WEI; Feng-Chao WANG; Liang-Peng GE; Xiang GAO

    2006-01-01

    Skin graft rejection is a typical cellular immune response, mainly mediated by T cells. Cytotoxic T lymphocyte associated antigen 4-immunoglobin (CTLA4-Ig) extends graft survival by blocking the T cell co-stimulation pathway and inhibiting T cell activation. To investigate the efficacy of CTLA4-Ig in prolonging skin graft survival, human CTLA4-Ig (hCTLA4-Ig) was engineered to overexpress in mouse skin by transgenesis using the K14 promoter. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot assay indicated that the expression of CTLA4-Ig remained skin-specific and relatively constant compared to the internal control protein, AKT, through seven generations. The presence and concentration of the hCTLA4-Ig protein in transgenic mouse sera was determined by enzyme-linked immunosorbent assay (ELISA), and the results indicated that the serum CTLA4-Ig concentration also remained constant through generations. Survival of transgenic mouse skins grafted onto rat wounds was remarkably prolonged compared to that of wild-type skins from the same mouse strain, and remained comparable among all seven generations. This suggested that the bioactive hCTLA4-Ig protein was stably expressed in transgenical mice through at least seven generations, which was consistent with the stable skin-specific CTLA4-Ig expression.The results demonstrated that the transgenic expression of hCTLA4-Ig in skin driven by the K14 promoter remained constant through generations, and a transgenic line can be established to provide transgenic skin with extended survival reproducibly.

  15. Transgenic soybean overexpressing GmSamT1 exhibits resistance to multiple-HG types of soybean cyst nematode Heterodera glycines

    Science.gov (United States)

    Soybean (Glycine max (L.) Merr.) salicylic acid methyl transferase (GmSAMT1) catalyzes the conversion of salicylic acid to methyl salicylate. Prior results showed that when GmSAMT1 was overexpressed in transgenic soybean hairy roots, resistance is conferred against soybean cyst nematode (SCN), Heter...

  16. Potential Impacts of Pharmaceutical Uses of Transgenic Tobacco: The Case of Human Serum Albumin and Gaucher's Disease Treatment

    OpenAIRE

    Kostandini, Gentian

    2004-01-01

    This thesis examines the size and distribution of benefits from the use of transgenic tobacco as a production vehicle for pharmaceutical proteins. Ex-ante welfare benefits are estimated for the introduction of two biotech innovations. In both cases economic surplus model with imperfect competition is employed to assess the size and distribution of benefits from these alternative uses of tobacco. An introductory chapter presents an overview of the topic followed by chapters 2 and 3 which conta...

  17. Cell culture-induced gradual and frequent epigenetic reprogramming of invertedly repeated tobacco transgene epialleles

    Czech Academy of Sciences Publication Activity Database

    Křížová, Kateřina; Fojtová, Miloslava; Depicker, A.; Kovařík, Aleš

    2009-01-01

    Roč. 149, č. 3 (2009), s. 1493-1504 ISSN 0032-0889 R&D Projects: GA AV ČR(CZ) IAA600040611; GA ČR(CZ) GD204/05/H505; GA ČR(CZ) GA521/07/0116 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : tobacco * cell culture * transgene silencing Subject RIV: BO - Biophysics Impact factor: 6.235, year: 2009

  18. A novel cold-regulated gene from Camellia sinensis, CsCOR1, enhances salt- and dehydration-tolerance in tobacco

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xian-Wen, E-mail: xianwenli01@sina.com [College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070 (China); College of Life Science, Xinyang Normal University, Xinyang 464000 (China); Key Laboratory of Horticultural Plant Biology of the Ministry of Education, Huazhong Agricultural University, Wuhan 430070 (China); Feng, Zhi-Guo; Yang, Hui-Min; Zhu, Xiao-Pei [College of Life Science, Xinyang Normal University, Xinyang 464000 (China); Liu, Jun, E-mail: liujun@mail.hzau.edu.cn [College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070 (China); Key Laboratory of Horticultural Plant Biology of the Ministry of Education, Huazhong Agricultural University, Wuhan 430070 (China); Yuan, Hong-Yu, E-mail: yhongyu92@163.com [College of Life Science, Xinyang Normal University, Xinyang 464000 (China)

    2010-04-02

    In present research, the full-length cDNA and the genomic sequence of a novel cold-regulated gene, CsCOR1, were isolated from Camellia sinensis L. The deduced protein CsCOR1 contains a hydrophobic N-terminus as a signal peptide and a hydrophilic C-terminal domain that is rich in glycine, arginine and proline. Two internal repetitive tridecapeptide fragments (HSVTAGRGGYNRG) exist in the middle of the C-terminal domain and the two nucleotide sequences encoding them are identical. CsCOR1 was localized in the cell walls of transgenic-tobaccos via CsCOR1::GFP fusion approach. The expression of CsCOR1 in tea leaves was enhanced dramatically by both cold- and dehydration-stress. And overexpression of CsCOR1 in transgenic-tobaccos improved obviously the tolerance to salinity and dehydration.

  19. The overexpression of insect endogenous small RNAs in transgenic rice inhibits growth and delays pupation of striped stem borer (Chilo suppressalis).

    Science.gov (United States)

    Jiang, Shan; Wu, Hao; Liu, Haoju; Zheng, Jie; Lin, Yongjun; Chen, Hao

    2017-07-01

    The striped stem borer (SSB), Chilo suppressalis Walker, is a major rice insect pest worldwide. RNA interference (RNAi) has become a promising strategy for developing insect-resistant crops. In a previous study, five double-stranded RNAs (dsRNAs) targeting important SSB housekeeping genes were overexpressed in rice, but none of the acquired dsRNA-transgenic rice plants showed significant effects on SSB. Thirteen selected SSB endogenous small RNAs, predicted as SSB novel microRNAs (miRNAs), were overexpressed in rice using artificial miRNA (amiRNA) expression technology. Feeding tests showed that two out of 13 selected SSB novel miRNAs caused significant growth inhibition for feeding SSB larvae based on transgenic rice expression. Pupation was delayed 4 days when SSB larvae consecutively fed on transgenic rice expressing the SSB novel miRNA candidate csu-novel-miR15 (csu-15 rice). Gene expression analysis confirmed that the expression levels of at least six SSB unigenes significantly changed (i.e., were up- or down-regulated) after feeding on csu-15 rice. Our research demonstrated a novel RNAi strategy using SSB endogenous small RNAs to develop RNAi crops for pest management; this strategy is different from the common RNAi resulting from transgenic dsRNAs or amiRNAs targeting certain insect endogenous genes. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  20. Enhanced disease resistance and drought tolerance in transgenic rice plants overexpressing protein elicitors from Magnaporthe oryzae.

    Science.gov (United States)

    Wang, Zhenzhen; Han, Qiang; Zi, Qian; Lv, Shun; Qiu, Dewen; Zeng, Hongmei

    2017-01-01

    Exogenous application of the protein elicitors MoHrip1 and MoHrip2, which were isolated from the pathogenic fungus Magnaporthe oryzae (M. oryzae), was previously shown to induce a hypersensitive response in tobacco and to enhance resistance to rice blast. In this work, we successfully transformed rice with the mohrip1 and mohrip2 genes separately. The MoHrip1 and MoHrip2 transgenic rice plants displayed higher resistance to rice blast and stronger tolerance to drought stress than wild-type (WT) rice and the vector-control pCXUN rice. The expression of salicylic acid (SA)- and abscisic acid (ABA)-related genes was also increased, suggesting that these two elicitors may trigger SA signaling to protect the rice from damage during pathogen infection and regulate the ABA content to increase drought tolerance in transgenic rice. Trypan blue staining indicated that expressing MoHrip1 and MoHrip2 in rice plants inhibited hyphal growth of the rice blast fungus. Relative water content (RWC), water usage efficiency (WUE) and water loss rate (WLR) were measured to confirm the high capacity for water retention in transgenic rice. The MoHrip1 and MoHrip2 transgenic rice also exhibited enhanced agronomic traits such as increased plant height and tiller number.

  1. Cloning of transgenic tobacco BY-2 cells; an efficient method to analyse and reduce high natural heterogeneity of transgene expression.

    Science.gov (United States)

    Nocarova, Eva; Fischer, Lukas

    2009-04-22

    Phenotypic characterization of transgenic cell lines, frequently used in plant biology studies, is complicated because transgene expression in individual cells is often heterogeneous and unstable. To identify the sources and to reduce this heterogeneity, we transformed tobacco (Nicotiana tabacum L.) BY-2 cells with a gene encoding green fluorescent protein (GFP) using Agrobacterium tumefaciens, and then introduced a simple cloning procedure to generate cell lines derived from the individual transformed cells. Expression of the transgene was monitored by analysing GFP fluorescence in the cloned lines and also in lines obtained directly after transformation. The majority ( approximately 90%) of suspension culture lines derived from calli that were obtained directly from transformation consisted of cells with various levels of GFP fluorescence. In contrast, nearly 50% of lines generated by cloning cells from the primary heterogeneous suspensions consisted of cells with homogenous GFP fluorescence. The rest of the lines exhibited "permanent heterogeneity" that could not be resolved by cloning. The extent of fluorescence heterogeneity often varied, even among genetically identical clones derived from the primary transformed lines. In contrast, the offspring of subsequent cloning of the cloned lines was uniform, showing GFP fluorescence intensity and heterogeneity that corresponded to the original clone. The results demonstrate that, besides genetic heterogeneity detected in some lines, the primary lines often contained a mixture of epigenetically different cells that could be separated by cloning. This indicates that a single integration event frequently results in various heritable expression patterns, which are probably accidental and become stabilized in the offspring of the primary transformed cells early after the integration event. Because heterogeneity in transgene expression has proven to be a serious problem, it is highly advisable to use transgenes tagged with

  2. Transgenic tobacco plants constitutively expressing peanut BTF3 exhibit increased growth and tolerance to abiotic stresses.

    Science.gov (United States)

    Pruthvi, V; Rama, N; Parvathi, M S; Nataraja, K N

    2017-05-01

    Abiotic stresses limit crop growth and productivity worldwide. Cellular tolerance, an important abiotic stress adaptive trait, involves coordinated activities of multiple proteins linked to signalling cascades, transcriptional regulation and other diverse processes. Basal transcriptional machinery is considered to be critical for maintaining transcription under stressful conditions. From this context, discovery of novel basal transcription regulators from stress adapted crops like peanut would be useful for improving tolerance of sensitive plant types. In this study, we prospected a basal transcription factor, BTF3 from peanut (Arachis hypogaea L) and studied its relevance in stress acclimation by over expression in tobacco. AhBTF3 was induced under PEG-, NaCl-, and methyl viologen-induced stresses in peanut. The constitutive expression of AhBTF3 in tobacco increased plant growth under non stress condition. The transgenic plants exhibited superior phenotype compared to wild type under mannitol- and NaCl-induced stresses at seedling level. The enhanced cellular tolerance of transgenic plants was evidenced by higher cell membrane stability, reactive oxygen species (ROS) scavenging activity, seedling survival and vigour than wild type. The transgenic lines showed better in vitro regeneration capacity on growth media supplemented with NaCl than wild type. Superior phenotype of transgenic plants under osmotic and salinity stresses seems to be due to constitutive activation of genes of multiple pathways linked to growth and stress adaptation. The study demonstrated that AhBTF3 is a positive regulator of growth and stress acclimation and hence can be considered as a potential candidate gene for crop improvement towards stress adaptation. © 2016 German Botanical Society and The Royal Botanical Society of the Netherlands.

  3. Overexpression of the PtSOS2 gene improves tolerance to salt stress in transgenic poplar plants.

    Science.gov (United States)

    Yang, Yang; Tang, Ren-Jie; Jiang, Chun-Mei; Li, Bei; Kang, Tao; Liu, Hua; Zhao, Nan; Ma, Xu-Jun; Yang, Lei; Chen, Shao-Liang; Zhang, Hong-Xia

    2015-09-01

    In higher plants, the salt overly sensitive (SOS) signalling pathway plays a crucial role in maintaining ion homoeostasis and conferring salt tolerance under salinity condition. Previously, we functionally characterized the conserved SOS pathway in the woody plant Populus trichocarpa. In this study, we demonstrate that overexpression of the constitutively active form of PtSOS2 (PtSOS2TD), one of the key components of this pathway, significantly increased salt tolerance in aspen hybrid clone Shanxin Yang (Populus davidiana × Populus bolleana). Compared to the wild-type control, transgenic plants constitutively expressing PtSOS2TD exhibited more vigorous growth and produced greater biomass in the presence of high concentrations of NaCl. The improved salt tolerance was associated with a decreased Na(+) accumulation in the leaves of transgenic plants. Further analyses revealed that plasma membrane Na(+) /H(+) exchange activity and Na(+) efflux in transgenic plants were significantly higher than those in the wild-type plants. Moreover, transgenic plants showed improved capacity in scavenging reactive oxygen species (ROS) generated by salt stress. Taken together, our results suggest that PtSOS2 could serve as an ideal target gene to genetically engineer salt-tolerant trees. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  4. [Induced expression of Serratia marcescens ribonuclease III gene in transgenic Nicotiana tabacum L. cv. SR1 tobacco plants].

    Science.gov (United States)

    Zhirnov, I V; Trifonova, E A; Romanova, A V; Filipenko, E A; Sapotsky, M V; Malinovsky, V I; Kochetov, A V; Shumny, V K

    2016-11-01

    Transgenic Nicotiana tabacum L. cv. SR1 plants, characterized by an increase in the level of dsRNA-specific hydrolytic activity after induction by wounding, were obtained. The Solanum lycopersicum anionic peroxidase gene promoter (new for plant genetic engineering) was for the first time used for the induced expression of the target Serratia marcescens RNase III gene. Upon infection with the tobacco mosaic virus (TMV), the transgenic plants of the obtained lines did not differ significantly from the control group in the level of TMV capsid protein accumulation. In general, no delay in the development of the infection symptoms was observed in transgenic plants as compared with the control group. The obtained transgenic plants represent a new model for the study of the biological role of endoribonucleases from the RNase III family, including in molecular mechanisms of resistance to pathogens.

  5. Overexpression of persimmon DkXTH1 enhanced tolerance to abiotic stress and delayed fruit softening in transgenic plants.

    Science.gov (United States)

    Han, Ye; Han, Shoukun; Ban, Qiuyan; He, Yiheng; Jin, Mijing; Rao, Jingping

    2017-04-01

    DkXTH1 promoted cell elongation and more strength to maintain structural integrity by involving in cell wall assembly, thus enhanced tolerance to abiotic stress with broader phenotype in transgenic plants. Xyloglucan endotransglucosylase/hydrolase (XTH) is thought to play a key role in cell wall modifications by cleaving and re-joining xyloglucan, and participates in the diverse physiological processes. DkXTH1 was found to peak in immature expanding persimmon fruit, and its higher expression level exhibited along with firmer fruit during storage. In the present study, transgenic Arabidopsis and tomato plants were generated with DkXTH1 constitutively expressed. Overexpression of DkXTH1 enhanced tolerance to salt, ABA and drought stresses in transgenic Arabidopsis plants with respect to root and leaf growth, and survival. Transgenic tomatoes collected at the mature green stage, presented delayed fruit softening coupled with postponed color change, a later and lower ethylene peak, and higher firmness in comparison with the wild-type tomatoes during storage. Furthermore, broader leaves and tomato fruit with larger diameter were gained in transgenic Arabidopsis and tomato, respectively. Most importantly, transgenic plants exhibited more large and irregular cells with higher density of cell wall and intercellular spaces, resulting from the overactivity of XET enzymes involving in cell wall assembly. We suggest that DkXTH1 expression resulted in cells with more strength and thickness to maintain structural integrity, and thus enhanced tolerance to abiotic stress and delayed fruit softening in transgenic plants.

  6. AgFNS overexpression increase apigenin and decrease anthocyanins in petioles of transgenic celery.

    Science.gov (United States)

    Tan, Guo-Fei; Ma, Jing; Zhang, Xin-Yue; Xu, Zhi-Sheng; Xiong, Ai-Sheng

    2017-10-01

    Apigenin and anthocyanin biosyntheses share common precursors in plants. Flavone synthase (FNS) converts naringenin into apigenin in higher plants. Celery is an important edible and medical vegetable crop that contains apigenin in its tissues. However, the effect of high AgFNS gene expression on the apigenin and anthocyanins contents of purple celery remains to be elucidated. In this study, the AgFNS gene was cloned from purple celery ('Nanxuan liuhe purple celery') and overexpressed in this purple celery to determine its influence on anthocyanins and apigenin contents. Results showed that the AgFNS gene was 1068bp, which encodes 355 amino acid residues. Evolution analysis showed that the AgFNS protein belongs to the FSN I type. In AgFNS transgenic celery, the anthocyanins content in petioles was lower than that wild-type celery plants. Apigenin content increased in the petioles of AgFNS transgenic celery. The transcript levels of the AgPAL, AgC4H, AgCHS, and AgCHI genes were up-regulated, whereas those of the AgF3H, AgF3'H, AgDFR, AgANS, and Ag3GT genes were down-regulated in the petioles of AgFNS transgenic plants compared with wild-type celery plants. This work provides basic knowledge about the function of the AgFNS gene in the anthocyanin and apigenin biosyntheses of celery. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Transgenic overexpression of 14-3-3 zeta protects hippocampus against endoplasmic reticulum stress and status epilepticus in vivo.

    Directory of Open Access Journals (Sweden)

    Gary P Brennan

    Full Text Available 14-3-3 proteins are ubiquitous molecular chaperones that are abundantly expressed in the brain where they regulate cell functions including metabolism, the cell cycle and apoptosis. Brain levels of several 14-3-3 isoforms are altered in diseases of the nervous system, including epilepsy. The 14-3-3 zeta (ζ isoform has been linked to endoplasmic reticulum (ER function in neurons, with reduced levels provoking ER stress and increasing vulnerability to excitotoxic injury. Here we report that transgenic overexpression of 14-3-3ζ in mice results in selective changes to the unfolded protein response pathway in the hippocampus, including down-regulation of glucose-regulated proteins 78 and 94, activating transcription factors 4 and 6, and Xbp1 splicing. No differences were found between wild-type mice and transgenic mice for levels of other 14-3-3 isoforms or various other 14-3-3 binding proteins. 14-3-3ζ overexpressing mice were potently protected against cell death caused by intracerebroventricular injection of the ER stressor tunicamycin. 14-3-3ζ overexpressing mice were also potently protected against neuronal death caused by prolonged seizures. These studies demonstrate that increased 14-3-3ζ levels protect against ER stress and seizure-damage despite down-regulation of the unfolded protein response. Delivery of 14-3-3ζ may protect against pathologic changes resulting from prolonged or repeated seizures or where injuries provoke ER stress.

  8. An N-terminal peptide extension results in efficient expression, but not secretion, of a synthetic horseradish peroxidase gene in transgenic tobacco.

    Science.gov (United States)

    Kis, Mihaly; Burbridge, Emma; Brock, Ian W; Heggie, Laura; Dix, Philip J; Kavanagh, Tony A

    2004-03-01

    Native horseradish (Armoracia rusticana) peroxidase, HRP (EC 1.11.1.7), isoenzyme C is synthesized with N-terminal and C-terminal peptide extensions, believed to be associated with protein targeting. This study aimed to explore the specific functions of these extensions, and to generate transgenic plants with expression patterns suitable for exploring the role of peroxidase in plant development and defence. Transgenic Nicotiana tabacum (tobacco) plants expressing different versions of a synthetic horseradish peroxidase, HRP, isoenzyme C gene were constructed. The gene was engineered to include additional sequences coding for either the natural N-terminal or the C-terminal extension or both. These constructs were placed under the control of a constitutive promoter (CaMV-35S) or the tobacco RUBISCO-SSU light inducible promoter (SSU) and introduced into tobacco using Agrobacterium-mediated transformation. To study the effects of the N- and C-terminal extensions, the localization of recombinant peroxidase was determined using biochemical and molecular techniques. Transgenic tobacco plants can exhibit a ten-fold increase in peroxidase activity compared with wild-type tobacco levels, and the majority of this activity is located in the symplast. The N-terminal extension is essential for the production of high levels of recombinant protein, while the C-terminal extension has little effect. Differences in levels of enzyme activity and recombinant protein are reflected in transcript levels. There is no evidence to support either preferential secretion or vacuolar targeting of recombinant peroxidase in this heterologous expression system. This leads us to question the postulated targeting roles of these peptide extensions. The N-terminal extension is essential for high level expression and appears to influence transcript stability or translational efficiency. Plants have been generated with greatly elevated cytosolic peroxidase activity, and smaller increases in apoplastic

  9. Light induced apoptosis is accelerated in transgenic retina overexpressing human EAT/mcl-1, an anti-apoptotic bcl-2 related gene.

    Science.gov (United States)

    Shinoda, K; Nakamura, Y; Matsushita, K; Shimoda, K; Okita, H; Fukuma, M; Yamada, T; Ohde, H; Oguchi, Y; Hata, J; Umezawa, A

    2001-10-01

    EAT/mcl-1 (EAT), an immediate early gene, functions in a similar way to bcl-2 in neutralising Bax mediated cytotoxicity, suggesting that EAT is a blocker of cell death. The aim of this study was to determine the effect of overexpression of the human EAT gene on light induced retinal cell apoptosis. EAT transgenic mice incorporating the EF-1alpha promoter were utilised, and expression of human EAT was detected by RT-PCR. Light damage was induced by raising mice under constant illumination. Two groups of animals, EAT transgenic mice (n=14) and littermates (n=13), were examined by ERG testing and histopathology at regular time points up to 20 weeks of constant light stimulation. Electrophysiological and histopathological findings were evaluated by established systems of arbitrary scoring as scores 0-2 and scores 0-3, respectively. The mean score (SD) of ERG response was significantly lower in EAT transgenic mice (0.79 (0.89)) than in littermates (1.69 (0.48)) (pstatistical significance (p=0.1156), the estimated incidence of electrophysiological retinal damage was higher in EAT mice (0.0495/mouse/week; 95% confidence interval (CI) 0.0347-0.0500) than in littermates (0. 0199/mouse/week; 95% CI 0.0035-0.0364). The mean scores (SD) for histopathological retinal degeneration were 2.31 (0.63) in littermates and 1.43 (1.22) in EAT transgenic mice (p=0.065). However, Kaplan-Meier curves for histopathological failure in two groups of mice showed that retinal photoreceptor cells were preserved significantly against constant light in the littermate compared with transgenic mice (p=0.0241). The estimated incidence of histopathological retinal damage was 0.0042/mouse/week in the littermates (95% CI 0-0.0120) and 0.0419/mouse/week in the EAT mice (95% CI 0.0286-0.0500). Retinal photoreceptor cell apoptosis under constant light stimulation is likely to be accelerated in transgenic retina overexpressing EAT.

  10. Overexpression of CaDSR6 increases tolerance to drought and salt stresses in transgenic Arabidopsis plants.

    Science.gov (United States)

    Kim, Eun Yu; Seo, Young Sam; Park, Ki Youl; Kim, Soo Jin; Kim, Woo Taek

    2014-11-15

    The partial CaDSR6 (Capsicum annuum Drought Stress Responsive 6) cDNA was previously identified as a drought-induced gene in hot pepper root tissues. However, the cellular role of CaDSR6 with regard to drought stress tolerance was unknown. In this report, full-length CaDSR6 cDNA was isolated. The deduced CaDSR6 protein was composed of 234 amino acids and contained an approximately 30 amino acid-long Asp-rich domain in its central region. This Asp-rich domain was highly conserved in all plant DSR6 homologs identified and shared a sequence identity with the N-terminal regions of yeast p23(fyp) and human hTCTP, which contain Rab protein binding sites. Transgenic Arabidopsis plants overexpressing CaDSR6 (35S:CaDSR6-sGFP) were tolerant to high salinity, as identified by more vigorous root growth and higher levels of total chlorophyll than wild type plants. CaDSR6-overexpressors were also more tolerant to drought stress compared to wild type plants. The 35S:CaDSR6-sGFP leaves retained their water content and chlorophyll more efficiently than wild type leaves in response to dehydration stress. The expression of drought-induced marker genes, such as RD20, RD22, RD26, RD29A, RD29B, RAB18, KIN2, ABF3, and ABI5, was markedly increased in CaDSR6-overexpressing plants relative to wild type plants under both normal and drought conditions. These results suggest that overexpression of CaDSR6 is associated with increased levels of stress-induced genes, which, in turn, conferred a drought tolerant phenotype in transgenic Arabidopsis plants. Overall, our data suggest that CaDSR6 plays a positive role in the response to drought and salt stresses. Copyright © 2014 Elsevier B.V. All rights reserved.

  11. Regulation of trichome development in tobacco by JcZFP8, a C2H2 zinc finger protein gene from Jatropha curcas L.

    Science.gov (United States)

    Shi, Xiaodong; Gu, Yuxi; Dai, Tingwei; Wu, Yang; Wu, Peng; Xu, Ying; Chen, Fang

    2018-06-05

    Trichomes are epidermal outgrowths of plant tissues that can secrete or store large quantities of secondary metabolites, which contribute to plant defense responses against stress. The use of bioengineering methods for regulating the development of trichomes and metabolism is a widely researched topic. In the present study, we demonstrate that JcZFP8, a C2H2 zinc finger protein gene from Jatropha curcas L., can regulate trichome development in transgenic tobacco. To understand the underlying mechanisms, we performed transcriptome profiling of overexpression JcZFP8 transgenic plants and wild-type tobacco. Based on the analysis of differentially expressed genes, we determined that genes of the plant hormone signal transduction pathway was significantly enriched, suggesting that these pathways were modulated in the transgenic plants. In addition, the transcript levels of the known trichome-related genes in Arabidopsis were not significantly changed, whereas CycB2 and MYB genes were differentially expressed in the transgenic plants. Despite tobacco and Arabidopsis have different types of trichomes, all the pathways were associated with C2H2 zinc finger protein genes. Our findings help us to understand the regulation of multicellular trichome formation and suggest a new metabolic engineering method for the improvement of plants. Copyright © 2018 Elsevier B.V. All rights reserved.

  12. Chloroplast overexpression of rice caffeic acid O-methyltransferase increases melatonin production in chloroplasts via the 5-methoxytryptamine pathway in transgenic rice plants.

    Science.gov (United States)

    Choi, Geun-Hee; Lee, Hyoung Yool; Back, Kyoungwhan

    2017-08-01

    Recent analyses of the enzymatic features of various melatonin biosynthetic genes from bacteria, animals, and plants have led to the hypothesis that melatonin could be synthesized via the 5-methoxytryptamine (5-MT) pathway. 5-MT is known to be synthesized in vitro from serotonin by the enzymatic action of O-methyltransferases, including N-acetylserotonin methyltransferase (ASMT) and caffeic acid O-methyltransferase (COMT), leading to melatonin synthesis by the subsequent enzymatic reaction with serotonin N-acetyltransferase (SNAT). Here, we show that 5-MT was produced and served as a precursor for melatonin synthesis in plants. When rice seedlings were challenged with senescence treatment, 5-MT levels and melatonin production were increased in transgenic rice seedlings overexpressing the rice COMT in chloroplasts, while no such increases were observed in wild-type or transgenic seedlings overexpressing the rice COMT in the cytosol, suggesting a 5-MT transport limitation from the cytosol to chloroplasts. In contrast, cadmium treatment led to results different from those in senescence. The enhanced melatonin production was not observed in the chloroplast COMT lines relative over the cytosol COMT lines although 5-MT levels were equally induced in all genotypes upon cadmium treatment. The transgenic seedlings with enhanced melatonin in their chloroplasts exhibited improved seedling growth vs the wild type under continuous light conditions. This is the first report describing enhanced melatonin production in chloroplasts via the 5-MT pathway with the ectopic overexpression of COMT in chloroplasts in plants. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  13. Increased yield of heterologous viral glycoprotein in the seeds of homozygous transgenic tobacco plants cultivated underground.

    Science.gov (United States)

    Tackaberry, Eilleen S; Prior, Fiona; Bell, Margaret; Tocchi, Monika; Porter, Suzanne; Mehic, Jelica; Ganz, Peter R; Sardana, Ravinder; Altosaar, Illimar; Dudani, Anil

    2003-06-01

    The use of transgenic plants in the production of recombinant proteins for human therapy, including subunit vaccines, is being investigated to evaluate the efficacy and safety of these emerging biopharmaceutical products. We have previously shown that synthesis of recombinant glycoprotein B (gB) of human cytomegalovirus can be targeted to seeds of transgenic tobacco when directed by the rice glutelin 3 promoter, with gB retaining critical features of immunological reactivity (E.S. Tackaberry et al. 1999. Vaccine, 17: 3020-3029). Here, we report development of second generation transgenic plant lines (T1) homozygous for the transgene. Twenty progeny plants from two lines (A23T(1)-2 and A24T(1)-3) were grown underground in an environmentally contained mine shaft. Based on yields of gB in their seeds, the A23T(1)-2 line was then selected for scale-up in the same facility. Analyses of mature seeds by ELISA showedthat gB specific activity in A23T(1)-2 seeds was over 30-fold greater than the best T0 plants from the same transformation series, representing 1.07% total seed protein. These data demonstrate stable inheritance, an absence of transgene inactivation, and enhanced levels of gB expression in a homozygous second generation plant line. They also provide evidence for the suitability of using this environmentally secure facility to grow transgenic plants producing therapeutic biopharmaceuticals.

  14. Cloning of Ammopiptanthus mongolicus C-repeat-binding factor gene and its cold-induced tolerance in transgenic tobacco

    Directory of Open Access Journals (Sweden)

    Lijiang Gu

    2013-12-01

    Full Text Available C-repeat-binding factors (CBFs are a type of important regulon in stress-related signal transduction pathways that control plant tolerance of abiotic stress. Ammopiptanthus mongolicus is the only evergreen broadleaf shrub in the northwest desert of China. The species shows strong resistance to environmental stress, especially to cold stress. An A. mongolicus CBF1 gene (AmCBF1 was cloned and transformed into tobacco. Expression of AmCBF1 could be detected in A. mongolicus shortly after exposure to low temperature of 4°C. Analysis on ratio of electrolytic leakage, soluble sugar content, free proline content, malondialdehyde (MDA content and peroxidase (POD activity before and after cold treatment (4°C for 24 h indicated AmCBF1 conferred higher cold tolerance to AmCBF1 transgenic tobacco compared with the wild type and empty vector transformed tobacco.

  15. Overexpression of rice thaumatin-like protein (Ostlp gene in transgenic cassava results in enhanced tolerance to Colletotrichum gloeosporioides f. sp. manihotis

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    Patroba Odeny Ojola

    2018-06-01

    Full Text Available Cassava (Manihot esculenta Crantz is the most important staple food for more than 300 million people in Africa, and anthracnose disease caused by Colletotrichum gloeosporioides f. sp. manihotis is the most destructive fungal disease affecting cassava production in sub-Saharan Africa. The main objective of this study was to improve anthracnose resistance in cassava through genetic engineering. Transgenic cassava plants harbouring rice thaumatin-like protein (Ostlp gene, driven by the constitutive CaMV35S promoter, were generated using Agrobacterium-mediated transformation of friable embryogenic calli (FEC of cultivar TMS 60444. Molecular analysis confirmed the presence, integration, copy number of the transgene all the independent transgenic events. Semi-quantitative RT-PCR confirmed high expression levels of Ostlp in six transgenic lines tested. The antifungal activity of the transgene against Colletotrichum gloeosporioides pathogen was evaluated using the leaves and stem cuttings bioassay. The results demonstrated significantly delayed disease development and reduced size of necrotic lesions in leaves and stem cuttings of all transgenic lines compared to the leaves and stem cuttingss of non-transgenic control plants. Therefore, constitutive overexpression of rice thaumatin-like protein in transgenic cassava confers enhanced tolerance to the fungal pathogen C. gloeosporioides f. sp. manihotis. These results can therefore serve as an initial step towards genetic engineering of farmer-preffered cassava cultivars for resistance to anthracnose disease. Keywords: Colletotrichum gloeosporioides f. sp. manihotis, Thaumatin-like protein, Transgenic cassava

  16. Protective effect of oral administration of transgenic tobacco seeds against verocytotoxic Escherichia coli strain in piglets.

    Science.gov (United States)

    Rossi, Luciana; Dell'Orto, Vittorio; Vagni, Simona; Sala, Vittorio; Reggi, Serena; Baldi, Antonella

    2014-03-01

    The use of transgenic plants as delivery system for antigenic proteins is attractive for its simplicity and increases likelihood for local immune response at sites of infection. The aim of this study was to evaluate the protective effect of oral administration of tobacco seeds, expressing the FedA, the major protein of the F18 adhesive fimbriae, and B subunit of verocytotoxin, against verocytotoxin-producing E. coli (VTEC) strain in piglets. Forty-three early weaned piglets, were randomly divided into 4 experimental groups: 3 test groups and a control. Treatment groups orally received a bolus, with different dose of tobacco seeds on 0, 1, 2, 14 days post primary administration. After challenge, with 1*10(10) CFU of O138 Escherichia coli strain, piglets showed clinical scores significantly higher in the control group compared to orally immunized groups (P administration of recombinant tobacco seeds expressing antigenic proteins against VTEC strains can induce a protective effect against challenger strain in piglets.

  17. Double overexpression of DREB and PIF transcription factors improves drought stress tolerance and cell elongation in transgenic plants.

    Science.gov (United States)

    Kudo, Madoka; Kidokoro, Satoshi; Yoshida, Takuya; Mizoi, Junya; Todaka, Daisuke; Fernie, Alisdair R; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

    2017-04-01

    Although a variety of transgenic plants that are tolerant to drought stress have been generated, many of these plants show growth retardation. To improve drought tolerance and plant growth, we applied a gene-stacking approach using two transcription factor genes: DEHYDRATION-RESPONSIVE ELEMENT-BINDING 1A (DREB1A) and rice PHYTOCHROME-INTERACTING FACTOR-LIKE 1 (OsPIL1). The overexpression of DREB1A has been reported to improve drought stress tolerance in various crops, although it also causes a severe dwarf phenotype. OsPIL1 is a rice homologue of Arabidopsis PHYTOCHROME-INTERACTING FACTOR 4 (PIF4), and it enhances cell elongation by activating cell wall-related gene expression. We found that the OsPIL1 protein was more stable than PIF4 under light conditions in Arabidopsis protoplasts. Transactivation analyses revealed that DREB1A and OsPIL1 did not negatively affect each other's transcriptional activities. The transgenic plants overexpressing both OsPIL1 and DREB1A showed the improved drought stress tolerance similar to that of DREB1A overexpressors. Furthermore, double overexpressors showed the enhanced hypocotyl elongation and floral induction compared with the DREB1A overexpressors. Metabolome analyses indicated that compatible solutes, such as sugars and amino acids, accumulated in the double overexpressors, which was similar to the observations of the DREB1A overexpressors. Transcriptome analyses showed an increased expression of abiotic stress-inducible DREB1A downstream genes and cell elongation-related OsPIL1 downstream genes in the double overexpressors, which suggests that these two transcription factors function independently in the transgenic plants despite the trade-offs required to balance plant growth and stress tolerance. Our study provides a basis for plant genetic engineering designed to overcome growth retardation in drought-tolerant transgenic plants. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology

  18. An N‐terminal Peptide Extension Results in Efficient Expression, but not Secretion, of a Synthetic Horseradish Peroxidase Gene in Transgenic Tobacco

    Science.gov (United States)

    KIS, MIHALY; BURBRIDGE, EMMA; BROCK, IAN W.; HEGGIE, LAURA; DIX, PHILIP J.; KAVANAGH, TONY A.

    2004-01-01

    • Background and Aims Native horseradish (Armoracia rusticana) peroxidase, HRP (EC 1.11.1.7), isoenzyme C is synthesized with N‐terminal and C‐terminal peptide extensions, believed to be associated with protein targeting. This study aimed to explore the specific functions of these extensions, and to generate transgenic plants with expression patterns suitable for exploring the role of peroxidase in plant development and defence. • Methods Transgenic Nicotiana tabacum (tobacco) plants expressing different versions of a synthetic horseradish peroxidase, HRP, isoenzyme C gene were constructed. The gene was engineered to include additional sequences coding for either the natural N‐terminal or the C‐terminal extension or both. These constructs were placed under the control of a constitutive promoter (CaMV‐35S) or the tobacco RUBISCO‐SSU light inducible promoter (SSU) and introduced into tobacco using Agrobacterium‐mediated transformation. To study the effects of the N‐ and C‐terminal extensions, the localization of recombinant peroxidase was determined using biochemical and molecular techniques. • Key Results Transgenic tobacco plants can exhibit a ten‐fold increase in peroxidase activity compared with wild‐type tobacco levels, and the majority of this activity is located in the symplast. The N‐terminal extension is essential for the production of high levels of recombinant protein, while the C‐terminal extension has little effect. Differences in levels of enzyme activity and recombinant protein are reflected in transcript levels. • Conclusions There is no evidence to support either preferential secretion or vacuolar targeting of recombinant peroxidase in this heterologous expression system. This leads us to question the postulated targeting roles of these peptide extensions. The N‐terminal extension is essential for high level expression and appears to influence transcript stability or translational efficiency. Plants have been

  19. Assessment of stress tolerance, productivity, and forage quality in T1 transgenic alfalfa co-overexpressing ZxNHX and ZxVP1-1 from Zygophyllum xanthoxylum

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    Peng Kang

    2016-10-01

    Full Text Available Salinization, desertification, and soil nutrient deprivation are threatening the production of alfalfa (Medicago sativa L. in northern China. We have previously generated T0 transgenic alfalfa co-overexpressing Zygophyllum xanthoxylum ZxNHX and ZxVP1-1 genes with enhanced salt and drought tolerance. To further develop this excellent breeding material into the new forage cultivar, stress tolerance, productivity, and forage quality of T1 transgenic alfalfa (GM were assessed in this study. The GM inherited the traits of salt and drought tolerance from T0 generation. Most importantly, co-overexpression of ZxNHX and ZxVP1-1 enhanced the tolerance to Pi deficiency in GM, which was associated with more Pi accumulation in plants. Meanwhile, T1 transgenic alfalfa developed a larger root system with increased root size, root dry weight and root/shoot ratio, which may be one important reason for the improvement of phosphorus nutrition and high biomass accumulation in GM under various conditions. GM also accumulated more crude protein, crude fibre, crude fat, and crude ash than wild-type (WT plants, especially under stress conditions and in the field. More interestingly, the crude fat contents sharply dropped in WT (by 66%-74%, whereas showed no change or decreased less in GM, when subjected to salinity, drought or low-Pi. Our results indicate that T1 transgenic alfalfa co-overexpressing ZxNHX and ZxVP1-1 shows stronger stress tolerance, higher productivity and better forage quality. This study provides a solid foundation for creating the alfalfa cultivars with high yield, good quality and wide adaptability on saline, dry and nutrient-deprived marginal lands of northern China.

  20. Enhanced quantitative resistance against fungal disease by combinatorial expression of different barley antifungal proteins in transgenic tobacco

    DEFF Research Database (Denmark)

    Jach, G; Görnhardt, B; Mundy, J

    1995-01-01

    cDNAs encoding three proteins from barley (Hordeum vulgare), a class-II chitinase (CHI), a class-II beta-1,3-glucanase (GLU) and a Type-I ribosome-inactivating protein (RIP) were expressed in tobacco plants under the control of the CaMV 35S-promoter. High-level expression of the transferred genes...... was detected in the transgenic plants by Northern and Western blot analysis. The leader peptides in CHI and GLU led to accumulation of these proteins in the intercellular space of tobacco leaves. RIP, which is naturally deposited in the cytosol of barley endosperm cells, was expressed either in its original...... cytosolic form or fused to a plant secretion peptide (spRIP). Fungal infection assays revealed that expression of the individual genes in each case resulted in an increased protection against the soilborne fungal pathogen Rhizoctonia solani, which infects a range of plant species including tobacco...

  1. Evaluation of deoxynivalenol production in dsRNA Carrying and Cured Fusarium graminearum isolates by AYT1 expressing transformed tobacco

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    Samira Shahbazi

    2015-12-01

    Full Text Available Introduction: Fusarium head blight (FHB, is the most destructive disease of wheat, producing the mycotoxin deoxynivalenol, a protein synthesis inhibitor, which is harmful to humans and livestock. dsRNAmycoviruses-infected-isolates of Fusariumgraminearum, showed changes in morphological and pathogenicity phenotypes including reduced virulence towards wheat and decreased production of trichothecene mycotoxin (deoxynivalenol: DON. Materials and methods: Previous studies indicated that over expression of yeast acetyl transferase gene (ScAYT1 encoding a 3-O trichothecene acetyl transferase that converts deoxynivalenol to a less toxic acetylated form, leads to suppression of the deoxynivalenol sensitivity in pdr5 yeast mutants. To identify whether ScAYT1 over-expression in transgenic tobacco plants can deal with mycotoxin (deoxynivalenol in fungal extract and studying the effect of dsRNA contamination on detoxification and resistance level, we have treated T1 AYT1 transgenic tobacco seedlings with complete extraction of normal F. graminearum isolate carrying dsRNA metabolites. First, we introduced AYT1into the model tobacco plants through Agrobacterium-mediated transformation in an attempt to detoxify deoxynivalenol. Results: In vitro tests with extraction of dsRNA carrying and cured isolates of F. graminearum and 10 ppm of deoxynivalenol indicated variable resistance levels in transgenic plants. Discussion and conclusion: The results of this study indicate that the transgene expression AYT1 and Fusarium infection to dsRNA can induce tolerance to deoxynivalenol, followed by increased resistance to Fusarium head blight disease of wheat.

  2. Constitutive and stress-inducible overexpression of a native aquaporin gene (MusaPIP2;6) in transgenic banana plants signals its pivotal role in salt tolerance.

    Science.gov (United States)

    Sreedharan, Shareena; Shekhawat, Upendra K Singh; Ganapathi, Thumballi R

    2015-05-01

    High soil salinity constitutes a major abiotic stress and an important limiting factor in cultivation of crop plants worldwide. Here, we report the identification and characterization of a aquaporin gene, MusaPIP2;6 which is involved in salt stress signaling in banana. MusaPIP2;6 was firstly identified based on comparative analysis of stressed and non-stressed banana tissue derived EST data sets and later overexpression in transgenic banana plants was performed to study its tangible functions in banana plants. The overexpression of MusaPIP2;6 in transgenic banana plants using constitutive or inducible promoter led to higher salt tolerance as compared to equivalent untransformed control plants. Cellular localization assay performed using transiently transformed onion peel cells indicated that MusaPIP2;6 protein tagged with green fluorescent protein was translocated to the plasma membrane. MusaPIP2;6-overexpressing banana plants displayed better photosynthetic efficiency and lower membrane damage under salt stress conditions. Our results suggest that MusaPIP2;6 is involved in salt stress signaling and tolerance in banana.

  3. Expression of chickpea CIPK25 enhances root growth and tolerance to dehydration and salt stress in transgenic tobacco

    Directory of Open Access Journals (Sweden)

    Mukesh Kumar Meena

    2015-09-01

    Full Text Available Calcium signaling plays an important role in adaptation and developmental processes in plants and animals. A class of calcium sensors, known as Calcineurin B-like (CBL proteins sense specific temporal changes in cytosolic Ca2+ concentration and regulate activities of a group of ser/thr protein kinases called CBL-interacting protein kinases (CIPKs. Although a number of CIPKs have been shown to play crucial roles in the regulation of stress signaling, no study on the function of CIPK25 or its orthologues has been reported so far. In the present study, an orthologue of Arabidopsis CIPK25 was cloned from chickpea (Cicer arietinum. CaCIPK25 gene expression in chickpea increased upon salt, dehydration, and different hormonal treatments. CaCIPK25 gene showed differential tissue-specific expression. 5'-upstream activation sequence (5'-UAS of the gene and its different truncated versions were fused to a reporter gene and studied in Arabidopsis to identify promoter regions directing its tissue-specific expression. Replacement of a conserved threonine residue with an aspartic acid at its catalytic site increased the kinase activity of CaCIPK25 by 2.5-fold. Transgenic tobacco plants overexpressing full-length and the high active versions of CaCIPK25 displayed a differential germination period and longer root length in comparison to the control plants. Expression of CaCIPK25 and its high active form differentially increased salt and water-deficit tolerance demonstrated by improved growth and reduced leaf chlorosis suggesting that the kinase activity of CaCIPK25 was required for these functions. Expressions of the abiotic stress marker genes were enhanced in the CaCIPK25-expressing tobacco plants. Our results suggested that CaCIPK25 functions in root development and abiotic stress tolerance.

  4. Expression of Finger Millet EcDehydrin7 in Transgenic Tobacco Confers Tolerance to Drought Stress.

    Science.gov (United States)

    Singh, Rajiv Kumar; Singh, Vivek Kumar; Raghavendrarao, Sanagala; Phanindra, Mullapudi Lakshmi Venkata; Venkat Raman, K; Solanke, Amolkumar U; Kumar, Polumetla Ananda; Sharma, Tilak Raj

    2015-09-01

    One of the critical alarming constraints for agriculture is water scarcity. In the current scenario, global warming due to climate change and unpredictable rainfall, drought is going to be a master player and possess a big threat to stagnating gene pool of staple food crops. So it is necessary to understand the mechanisms that enable the plants to cope with drought stress. In this study, effort was made to prospect the role of EcDehydrin7 protein from normalized cDNA library of drought tolerance finger millet in transgenic tobacco. Biochemical and molecular analyses of T0 transgenic plants were done for stress tolerance. Leaf disc assay, seed germination test, dehydration assay, and chlorophyll estimation showed EcDehydrin7 protein directly link to drought tolerance. Northern and qRT PCR analyses shows relatively high expression of EcDehydrin7 protein compare to wild type. T0 transgenic lines EcDehydrin7(11) and EcDehydrin7(15) shows superior expression among all lines under study. In summary, all results suggest that EcDehydrin7 protein has a remarkable role in drought tolerance and may be used for sustainable crop breeding program in other food crops.

  5. Functional analysis of multiple carotenogenic genes from Lycium barbarum and Gentiana lutea L. for their effects on beta-carotene production in transgenic tobacco.

    Science.gov (United States)

    Ji, Jing; Wang, Gang; Wang, Jiehua; Wang, Ping

    2009-02-01

    Carotenoids are red, yellow and orange pigments, which are widely distributed in nature and are especially abundant in yellow-orange fruits and vegetables and dark green leafy vegetables. Carotenoids are essential for photosynthesis and photoprotection in plant life and also have different beneficial effects in humans and animals (van den Berg et al. 2000). For example, beta-carotene plays an essential role as the main dietary source of vitamin A. To obtain further insight into beta-carotene biosynthesis in two important economic plant species, Lycium barbarum and Gentiana lutea L., and to investigate and prioritize potential genetic engineering targets in the pathway, the effects of five carotenogenic genes from these two species, encoding proteins including geranylgeranyl diphosphate synthase, phytoene synthase and delta-carotene desaturase gene, lycopene beta-cyclase, lycopene epsilon-cyclase were functionally analyzed in transgenic tobacco (Nicotiana tabacum) plants. All transgenic tobacco plants constitutively expressing these genes showed enhanced beta-carotene contents in their leaves and flowers to different extents. The addictive effects of co-ordinate expression of double transgenes have also been investigated.

  6. Targeted overexpression of amelotin disrupts the microstructure of dental enamel.

    Science.gov (United States)

    Lacruz, Rodrigo S; Nakayama, Yohei; Holcroft, James; Nguyen, Van; Somogyi-Ganss, Eszter; Snead, Malcolm L; White, Shane N; Paine, Michael L; Ganss, Bernhard

    2012-01-01

    We have previously identified amelotin (AMTN) as a novel protein expressed predominantly during the late stages of dental enamel formation, but its role during amelogenesis remains to be determined. In this study we generated transgenic mice that produce AMTN under the amelogenin (Amel) gene promoter to study the effect of AMTN overexpression on enamel formation in vivo. The specific overexpression of AMTN in secretory stage ameloblasts was confirmed by Western blot and immunohistochemistry. The gross histological appearance of ameloblasts or supporting cellular structures as well as the expression of the enamel proteins amelogenin (AMEL) and ameloblastin (AMBN) was not altered by AMTN overexpression, suggesting that protein production, processing and secretion occurred normally in transgenic mice. The expression of Odontogenic, Ameloblast-Associated (ODAM) was slightly increased in secretory stage ameloblasts of transgenic animals. The enamel in AMTN-overexpressing mice was much thinner and displayed a highly irregular surface structure compared to wild type littermates. Teeth of transgenic animals underwent rapid attrition due to the brittleness of the enamel layer. The microstructure of enamel, normally a highly ordered arrangement of hydroxyapatite crystals, was completely disorganized. Tomes' process, the hallmark of secretory stage ameloblasts, did not form in transgenic mice. Collectively our data demonstrate that the overexpression of amelotin has a profound effect on enamel structure by disrupting the formation of Tomes' process and the orderly growth of enamel prisms.

  7. Targeted overexpression of amelotin disrupts the microstructure of dental enamel.

    Directory of Open Access Journals (Sweden)

    Rodrigo S Lacruz

    Full Text Available We have previously identified amelotin (AMTN as a novel protein expressed predominantly during the late stages of dental enamel formation, but its role during amelogenesis remains to be determined. In this study we generated transgenic mice that produce AMTN under the amelogenin (Amel gene promoter to study the effect of AMTN overexpression on enamel formation in vivo. The specific overexpression of AMTN in secretory stage ameloblasts was confirmed by Western blot and immunohistochemistry. The gross histological appearance of ameloblasts or supporting cellular structures as well as the expression of the enamel proteins amelogenin (AMEL and ameloblastin (AMBN was not altered by AMTN overexpression, suggesting that protein production, processing and secretion occurred normally in transgenic mice. The expression of Odontogenic, Ameloblast-Associated (ODAM was slightly increased in secretory stage ameloblasts of transgenic animals. The enamel in AMTN-overexpressing mice was much thinner and displayed a highly irregular surface structure compared to wild type littermates. Teeth of transgenic animals underwent rapid attrition due to the brittleness of the enamel layer. The microstructure of enamel, normally a highly ordered arrangement of hydroxyapatite crystals, was completely disorganized. Tomes' process, the hallmark of secretory stage ameloblasts, did not form in transgenic mice. Collectively our data demonstrate that the overexpression of amelotin has a profound effect on enamel structure by disrupting the formation of Tomes' process and the orderly growth of enamel prisms.

  8. Expression of a finger millet transcription factor, EcNAC1, in tobacco confers abiotic stress-tolerance.

    Directory of Open Access Journals (Sweden)

    Venkategowda Ramegowda

    Full Text Available NAC (NAM, ATAF1-2, and CUC2 proteins constitute one of the largest families of plant-specific transcription factors and have been shown to be involved in diverse plant processes including plant growth, development, and stress-tolerance. In this study, a stress-responsive NAC gene, EcNAC1, was isolated from the subtracted stress cDNA library generated from a drought adapted crop, finger millet, and characterized for its role in stress-tolerance. The expression analysis showed that EcNAC1 was highly induced during water-deficit and salt stress. EcNAC1 shares high amino acid similarity with rice genes that have been phylogenetically classified into stress-related NAC genes. Our results demonstrated that tobacco transgenic plants expressing EcNAC1 exhibit tolerance to various abiotic stresses like simulated osmotic stress, by polyethylene glycol (PEG and mannitol, and salinity stress. The transgenic plants also showed enhanced tolerance to methyl-viologen (MV induced oxidative stress. Reduced levels of reactive oxygen species (ROS and ROS-induced damage were noticed in pot grown transgenic lines under water-deficit and natural high light conditions. Root growth under stress and recovery growth after stress alleviation was more in transgenic plants. Many stress-responsive genes were found to be up-regulated in transgenic lines expressing EcNAC1. Our results suggest that EcNAC1 overexpression confers tolerance against abiotic stress in susceptible species, tobacco.

  9. Transgenic overexpression of adenosine kinase in brain leads to multiple learning impairments and altered sensitivity to psychomimetic drugs.

    Science.gov (United States)

    Yee, Benjamin K; Singer, Philipp; Chen, Jiang-Fan; Feldon, Joram; Boison, Detlev

    2007-12-01

    The neuromodulator adenosine fulfills a unique role in the brain affecting glutamatergic neurotransmission and dopaminergic signaling via activation of adenosine A1 and A2A receptors, respectively. The adenosine system is thus ideally positioned to integrate glutamatergic and dopaminergic neurotransmission, which in turn could affect behavior and cognition. In the adult brain, adenosine levels are largely regulated by its key metabolic enzyme adenosine kinase (ADK), which may assume the role of an 'upstream regulator' of these two neurotransmitter pathways. To test this hypothesis, transgenic mice with an overexpression of ADK in brain (Adk-tg), and therefore reduced brain adenosine levels, were evaluated in a panel of behavioral and psychopharmacological assays to assess possible glutamatergic and dopaminergic dysfunction. In comparison to non-transgenic control mice, Adk-tg mice are characterized by severe learning deficits in the Morris water maze task and in Pavlovian conditioning. The Adk-tg mice also exhibited reduced locomotor reaction to systemic amphetamine, whereas their reaction to the non-competitive N-methyl-d-aspartate receptor antagonist MK-801 was enhanced. Our results confirmed that ADK overexpression could lead to functional concomitant alterations in dopaminergic and glutamatergic functions, which is in keeping with the hypothesized role of ADK in the balance and integration between glutamatergic and dopaminergic neurotransmission. The present findings are of relevance to current pathophysiological hypotheses of schizophrenia and its pharmacotherapy.

  10. Over-expression of Arabidopsis thaliana SFD1/GLY1, the gene encoding plastid localized glycerol-3-phosphate dehydrogenase, increases plastidic lipid content in transgenic rice plants.

    Science.gov (United States)

    Singh, Vijayata; Singh, Praveen Kumar; Siddiqui, Adnan; Singh, Subaran; Banday, Zeeshan Zahoor; Nandi, Ashis Kumar

    2016-03-01

    Lipids are the major constituents of all membranous structures in plants. Plants possess two pathways for lipid biosynthesis: the prokaryotic pathway (i.e., plastidic pathway) and the eukaryotic pathway (i.e., endoplasmic-reticulum (ER) pathway). Whereas some plants synthesize galactolipids from diacylglycerol assembled in the plastid, others, including rice, derive their galactolipids from diacylglycerols assembled by the eukaryotic pathway. Arabidopsis thaliana glycerol-3-phosphate dehydrogenase (G3pDH), coded by SUPPRESSOR OF FATTY ACID DESATURASE 1 (SFD1; alias GLY1) gene, catalyzes the formation of glycerol 3-phosphate (G3p), the backbone of many membrane lipids. Here SFD1 was introduced to rice as a transgene. Arabidopsis SFD1 localizes in rice plastids and its over-expression increases plastidic membrane lipid content in transgenic rice plants without any major impact on ER lipids. The results suggest that over-expression of plastidic G3pDH enhances biosynthesis of plastid-localized lipids in rice. Lipid composition in the transgenic plants is consistent with increased phosphatidylglycerol synthesis in the plastid and increased galactolipid synthesis from diacylglycerol produced via the ER pathway. The transgenic plants show a higher photosynthetic assimilation rate, suggesting a possible application of this finding in crop improvement.

  11. Comparative Study on Growth Performance of Transgenic (Over-Expressed OsNHX1 and Wild-Type Nipponbare under Different Salinity Regimes

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    Nurul Kahrani ISHAK

    2015-11-01

    Full Text Available Transgenic Nipponbare which over-expressed a Na+/H+ antiporter gene OsNHX1 was used to compare its growth performance, water status and photosynthetic efficiency with its wild type under varying salinity regimes. Chlorophyll content, quantum yield and photosynthetic rate were measured to assess the impact of salinity stress on photosynthetic efficiency for transgenic and wild-type Nipponbare. Effects of salinity on water status and gas exchange to both lines were studied by measuring water use efficiency, instantaneous transpiration rate and stomatal conductance. Dry shoot weight and leaf area were determined after three months of growth to assess the impacts of salinity on the growth of those two lines. Our study showed that both lines were affected by salinity stress, however, the transgenic line showed higher photosynthetic efficiency, better utilization of water, and better growth due to low transpiration rate and stomatal conductance. Reduction of photosynthetic efficiency exhibited by the wild-type Nipponbare was correlated to its poor growth under salinity stress.

  12. GhZFP1, a novel CCCH-type zinc finger protein from cotton, enhances salt stress tolerance and fungal disease resistance in transgenic tobacco by interacting with GZIRD21A and GZIPR5.

    Science.gov (United States)

    Guo, Ying-Hui; Yu, Yue-Ping; Wang, Dong; Wu, Chang-Ai; Yang, Guo-Dong; Huang, Jin-Guang; Zheng, Cheng-Chao

    2009-01-01

    * Zinc finger proteins are a superfamily involved in many aspects of plant growth and development. However, CCCH-type zinc finger proteins involved in plant stress tolerance are poorly understood. * A cDNA clone designated Gossypium hirsutum zinc finger protein 1 (GhZFP1), which encodes a novel CCCH-type zinc finger protein, was isolated from a salt-induced cotton (G. hirsutum) cDNA library using differential hybridization screening and further studied in transgenic tobacco Nicotiana tabacum cv. NC89. Using yeast two-hybrid screening (Y2H), proteins GZIRD21A (GhZFP1 interacting and responsive to dehydration protein 21A) and GZIPR5 (GhZFP1 interacting and pathogenesis-related protein 5), which interacted with GhZFP1, were isolated. * GhZFP1 contains two typical zinc finger motifs (Cx8Cx5Cx3H and Cx5Cx4Cx3H), a putative nuclear export sequence (NES) and a potential nuclear localization signal (NLS). Transient expression analysis using a GhZFP1::GFP fusion gene in onion epidermal cells indicated a nuclear localization for GhZFP1. RNA blot analysis showed that the GhZFP1 transcript was induced by salt (NaCl), drought and salicylic acid (SA). The regions in GhZFP1 that interact with GZIRD21A and GZIPR5 were identified using truncation mutations. * Overexpression of GhZFP1 in transgenic tobacco enhanced tolerance to salt stress and resistance to Rhizoctonia solani. Therefore, it appears that GhZFP1 might be involved as an important regulator in plant responses to abiotic and biotic stresses.

  13. Overexpression of ubiquitous 6-phosphofructo-2-kinase in the liver of transgenic mice results in weight gain

    International Nuclear Information System (INIS)

    Duran, Joan; Navarro-Sabate, Aurea; Pujol, Anna; Perales, Jose C.; Manzano, Anna; Obach, Merce; Gomez, Marta; Bartrons, Ramon

    2008-01-01

    Fructose 2,6-bisphosphate (Fru-2,6-P 2 ) is an important metabolite that controls glycolytic and gluconeogenic pathways in several cell types. Its synthesis and degradation are catalyzed by the bifunctional enzyme 6-phosphofructo-2-kinase/fructose 2,6-bisphosphatase (PFK-2). Four genes, designated Pfkfb1-4, codify the different PFK-2 isozymes. The Pfkfb3 gene product, ubiquitous PFK-2 (uPFK-2), has the highest kinase/bisphosphatase activity ratio and is associated with proliferation and tumor metabolism. A transgenic mouse model that overexpresses uPFK-2 under the control of the phosphoenolpyruvate carboxykinase promoter was designed to promote sustained and elevated Fru-2,6-P 2 levels in the liver. Our results demonstrate that in diet-induced obesity, high Fru-2,6-P 2 levels in transgenic livers caused changes in hepatic gene expression profiles for key gluconeogenic and lipogenic enzymes, as well as an accumulation of lipids in periportal cells, and weight gain

  14. Chronic wasting disease prions are not transmissible to transgenic mice overexpressing human prion protein.

    Science.gov (United States)

    Sandberg, Malin K; Al-Doujaily, Huda; Sigurdson, Christina J; Glatzel, Markus; O'Malley, Catherine; Powell, Caroline; Asante, Emmanuel A; Linehan, Jacqueline M; Brandner, Sebastian; Wadsworth, Jonathan D F; Collinge, John

    2010-10-01

    Chronic wasting disease (CWD) is a prion disease that affects free-ranging and captive cervids, including mule deer, white-tailed deer, Rocky Mountain elk and moose. CWD-infected cervids have been reported in 14 USA states, two Canadian provinces and in South Korea. The possibility of a zoonotic transmission of CWD prions via diet is of particular concern in North America where hunting of cervids is a popular sport. To investigate the potential public health risks posed by CWD prions, we have investigated whether intracerebral inoculation of brain and spinal cord from CWD-infected mule deer transmits prion infection to transgenic mice overexpressing human prion protein with methionine or valine at polymorphic residue 129. These transgenic mice have been utilized in extensive transmission studies of human and animal prion disease and are susceptible to BSE and vCJD prions, allowing comparison with CWD. Here, we show that these mice proved entirely resistant to infection with mule deer CWD prions arguing that the transmission barrier associated with this prion strain/host combination is greater than that observed with classical BSE prions. However, it is possible that CWD may be caused by multiple prion strains. Further studies will be required to evaluate the transmission properties of distinct cervid prion strains as they are characterized.

  15. The grapevine VvWRKY2 gene enhances salt and osmotic stress tolerance in transgenic Nicotiana tabacum.

    Science.gov (United States)

    Mzid, Rim; Zorrig, Walid; Ben Ayed, Rayda; Ben Hamed, Karim; Ayadi, Mariem; Damak, Yosra; Lauvergeat, Virginie; Hanana, Mohsen

    2018-06-01

    Our study aims to assess the implication of WRKY transcription factor in the molecular mechanisms of grapevine adaptation to salt and water stresses. In this respect, a full-length VvWRKY2 cDNA, isolated from a Vitis vinifera grape berry cDNA library, was constitutively over-expressed in Nicotiana tabacum seedlings. Our results showed that transgenic tobacco plants exhibited higher seed germination rates and better growth, under both salt and osmotic stress treatments, when compared to wild type plants. Furthermore, our analyses demonstrated that, under stress conditions, transgenic plants accumulated more osmolytes, such as soluble sugars and free proline, while no changes were observed regarding electrolyte leakage, H 2 O 2 , and malondialdehyde contents. The improvement of osmotic adjustment may be an important mechanism underlying the role of VvWRKY 2 in promoting tolerance and adaptation to abiotic stresses. Principal component analysis of our results highlighted a clear partition of plant response to stress. On the other hand, we observed a significant adaptation behaviour response for transgenic lines under stress. Taken together, all our findings suggest that over-expression of VvWRKY2 gene has a compelling role in abiotic stress tolerance and, therefore, would provide a useful strategy to promote abiotic stress tolerance in grape via molecular-assisted breeding and/or new biotechnology tools.

  16. Phytoremediation of small organic contaminants using transgenic plants

    Science.gov (United States)

    James, C Andrew; Strand, Stuart E

    2010-01-01

    The efficacy of transgenic plants in the phytoremediation of small organic contaminants has been investigated. Two principal strategies have been pursued (1) the manipulation of phase I metabolic activity to enhance in planta degradation rates, or to impart novel metabolic activity, and (2) the enhanced secretion of reactive enzymes from roots leading to accelerated ex planta degradation of organic contaminants. A pair of dehalogenase genes from Xanthobacter autotrophicus was expressed in tobacco resulting in the dehalogenation of 1,2-dichloroethane, which was otherwise recalcitrant. A laccase gene from cotton was overexpressed in Arabidopsis thaliana resulting in increased secretory laccase activity and the enhanced resistance to trichlorophenol in soils. Although the results to date are promising, much of the work has been limited to laboratory settings; field demonstrations are needed. PMID:19342219

  17. Resistance to the Beneficial Metabolic Effects and Hepatic Antioxidant Defense Actions of Fibroblast Growth Factor 21 Treatment in Growth Hormone-Overexpressing Transgenic Mice

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    Ravneet K. Boparai

    2015-01-01

    Full Text Available Fibroblast growth factor 21 (FGF21 modulates a diverse range of biological functions, including glucose and lipid metabolism, adaptive starvation response, and energy homeostasis, but with limited mechanistic insight. FGF21 treatment has been shown to inhibit hepatic growth hormone (GH intracellular signaling. To evaluate GH axis involvement in FGF21 actions, transgenic mice overexpressing bovine GH were used. Expectedly, in response to FGF21 treatment control littermates showed metabolic improvements whereas GH transgenic mice resisted most of the beneficial effects of FGF21, except an attenuation of the innate hyperinsulinemia. Since FGF21 is believed to exert its effects mostly at the transcriptional level, we analyzed and observed significant upregulation in expression of various genes involved in carbohydrate and lipid metabolism, energy homeostasis, and antioxidant defense in FGF21-treated controls, but not in GH transgenics. The resistance of GH transgenic mice to FGF21-induced changes underlines the necessity of normal GH signaling for the beneficial effects of FGF21.

  18. Overexpression of an alfalfa GDP-mannose 3, 5-epimerase gene enhances acid, drought and salt tolerance in transgenic Arabidopsis by increasing ascorbate accumulation.

    Science.gov (United States)

    Ma, Lichao; Wang, Yanrong; Liu, Wenxian; Liu, Zhipeng

    2014-11-01

    GDP-mannose 3', 5'-epimerase (GME) catalyses the conversion of GDP-D-mannose to GDP-L-galactose, an important step in the ascorbic acid (ascorbic acid) biosynthetic pathway in higher plants. In this study, a novel cDNA fragment (MsGME) encoding a GME protein was isolated and characterised from alfalfa (Medicago sativa). An expression analysis confirmed that MsGME expression was induced by salinity, PEG and acidity stresses. MsGME overexpression in Arabidopsis enhanced tolerance of the transgenic plants to salt, drought and acid. Real-time PCR analysis revealed that the transcript levels of GDP-D-mannose pyrophosphorylase (GMP), L-galactose-phosphate 1-P phosphatase (GP) and GDP-L-galactose phosphorylase (GGP) were increased in transgenic Arabidopsis (T3 generation). Moreover, the ascorbate content was increased in transgenic Arabidopsis. Our results suggest that MsGME can effectively enhance tolerance of transgenic Arabidopsis to acid, drought and salt by increasing ascorbate accumulation.

  19. Promoting scopolamine biosynthesis in transgenic Atropa belladonna plants with pmt and h6h overexpression under field conditions.

    Science.gov (United States)

    Xia, Ke; Liu, Xiaoqiang; Zhang, Qiaozhuo; Qiang, Wei; Guo, Jianjun; Lan, Xiaozhong; Chen, Min; Liao, Zhihua

    2016-09-01

    Atropa belladonna is one of the most important plant sources for producing pharmaceutical tropane alkaloids (TAs). T1 progeny of transgenic A. belladonna, in which putrescine N-methyltransferase (EC. 2.1.1.53) from Nicotiana tabacum (NtPMT) and hyoscyamine 6β-hydroxylase (EC. 1.14.11.14) from Hyoscyamus niger (HnH6H) were overexpressed, were established to investigate TA biosynthesis and distribution in ripe fruits, leaves, stems, primary roots and secondary roots under field conditions. Both NtPMT and HnH6H were detected at the transcriptional level in transgenic plants, whereas they were not detected in wild-type plants. The transgenes did not influence the root-specific expression patterns of endogenous TA biosynthetic genes in A. belladonna. All four endogenous TA biosynthetic genes (AbPMT, AbTRI, AbCYP80F1 and AbH6H) had the highest/exclusive expression levels in secondary roots, suggesting that TAs were mainly synthesized in secondary roots. T1 progeny of transgenic A. belladonna showed an impressive scopolamine-rich chemotype that greatly improved the pharmaceutical value of A. belladonna. The higher efficiency of hyoscyamine conversion was found in aerial than in underground parts. In aerial parts of transgenic plants, hyoscyamine was totally converted to downstream alkaloids, especially scopolamine. Hyoscyamine, anisodamine and scopolamine were detected in underground parts, but scopolamine and anisodamine were more abundant than hyoscyamine. The exclusively higher levels of anisodamine in roots suggested that it might be difficult for its translocation from root to aerial organs. T1 progeny of transgenic A. belladonna, which produces scopolamine at very high levels (2.94-5.13 mg g(-1)) in field conditions, can provide more valuable plant materials for scopolamine production. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  20. Overexpression of Populus trichocarpa CYP85A3 promotes growth and biomass production in transgenic trees.

    Science.gov (United States)

    Jin, Yan-Li; Tang, Ren-Jie; Wang, Hai-Hai; Jiang, Chun-Mei; Bao, Yan; Yang, Yang; Liang, Mei-Xia; Sun, Zhen-Cang; Kong, Fan-Jing; Li, Bei; Zhang, Hong-Xia

    2017-10-01

    Brassinosteroids (BRs) are essential hormones that play crucial roles in plant growth, reproduction and response to abiotic and biotic stress. In Arabidopsis, AtCYP85A2 works as a bifunctional cytochrome P450 monooxygenase to catalyse the conversion of castasterone to brassinolide, a final rate-limiting step in the BR-biosynthetic pathway. Here, we report the functional characterizations of PtCYP85A3, one of the three AtCYP85A2 homologous genes from Populus trichocarpa. PtCYP85A3 shares the highest similarity with AtCYP85A2 and can rescue the retarded-growth phenotype of the Arabidopsis cyp85a2-2 and tomato d x mutants. Constitutive expression of PtCYP85A3, driven by the cauliflower mosaic virus 35S promoter, increased the endogenous BR levels and significantly promoted the growth and biomass production in both transgenic tomato and poplar. Compared to the wild type, plant height, shoot fresh weight and fruit yield increased 50%, 56% and 43%, respectively, in transgenic tomato plants. Similarly, plant height and stem diameter increased 15% and 25%, respectively, in transgenic poplar plants. Further study revealed that overexpression of PtCYP85A3 enhanced xylem formation without affecting the composition of cellulose and lignin, as well as the cell wall thickness in transgenic poplar. Our finding suggests that PtCYP85A3 could be used as a potential candidate gene for engineering fast-growing trees with improved wood production. © 2017 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  1. The SbMT-2 gene from a halophyte confers abiotic stress tolerance and modulates ROS scavenging in transgenic tobacco.

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    Amit Kumar Chaturvedi

    Full Text Available Heavy metals are common pollutants of the coastal saline area and Salicornia brachiata an extreme halophyte is frequently exposed to various abiotic stresses including heavy metals. The SbMT-2 gene was cloned and transformed to tobacco for the functional validation. Transgenic tobacco lines (L2, L4, L6 and L13 showed significantly enhanced salt (NaCl, osmotic (PEG and metals (Zn++, Cu++ and Cd++ tolerance compared to WT plants. Transgenic lines did not show any morphological variation and had enhanced growth parameters viz. shoot length, root length, fresh weight and dry weight. High seed germination percentage, chlorophyll content, relative water content, electrolytic leakage and membrane stability index confirmed that transgenic lines performed better under salt (NaCl, osmotic (PEG and metals (Zn++, Cu++ and Cd++ stress conditions compared to WT plants. Proline, H2O2 and lipid peroxidation (MDA analyses suggested the role of SbMT-2 in cellular homeostasis and H2O2 detoxification. Furthermore in vivo localization of H2O2 and O2-; and elevated expression of key antioxidant enzyme encoding genes, SOD, POD and APX evident the possible role of SbMT-2 in ROS scavenging/detoxification mechanism. Transgenic lines showed accumulation of Cu++ and Cd++ in root while Zn++ in stem under stress condition. Under control (unstressed condition, Zn++ was accumulated more in root but accumulation of Zn++ in stem under stress condition suggested that SbMT-2 may involve in the selective translocation of Zn++ from root to stem. This observation was further supported by the up-regulation of zinc transporter encoding genes NtZIP1 and NtHMA-A under metal ion stress condition. The study suggested that SbMT-2 modulates ROS scavenging and is a potential candidate to be used for phytoremediation and imparting stress tolerance.

  2. The SbMT-2 gene from a halophyte confers abiotic stress tolerance and modulates ROS scavenging in transgenic tobacco.

    Science.gov (United States)

    Chaturvedi, Amit Kumar; Patel, Manish Kumar; Mishra, Avinash; Tiwari, Vivekanand; Jha, Bhavanath

    2014-01-01

    Heavy metals are common pollutants of the coastal saline area and Salicornia brachiata an extreme halophyte is frequently exposed to various abiotic stresses including heavy metals. The SbMT-2 gene was cloned and transformed to tobacco for the functional validation. Transgenic tobacco lines (L2, L4, L6 and L13) showed significantly enhanced salt (NaCl), osmotic (PEG) and metals (Zn++, Cu++ and Cd++) tolerance compared to WT plants. Transgenic lines did not show any morphological variation and had enhanced growth parameters viz. shoot length, root length, fresh weight and dry weight. High seed germination percentage, chlorophyll content, relative water content, electrolytic leakage and membrane stability index confirmed that transgenic lines performed better under salt (NaCl), osmotic (PEG) and metals (Zn++, Cu++ and Cd++) stress conditions compared to WT plants. Proline, H2O2 and lipid peroxidation (MDA) analyses suggested the role of SbMT-2 in cellular homeostasis and H2O2 detoxification. Furthermore in vivo localization of H2O2 and O2-; and elevated expression of key antioxidant enzyme encoding genes, SOD, POD and APX evident the possible role of SbMT-2 in ROS scavenging/detoxification mechanism. Transgenic lines showed accumulation of Cu++ and Cd++ in root while Zn++ in stem under stress condition. Under control (unstressed) condition, Zn++ was accumulated more in root but accumulation of Zn++ in stem under stress condition suggested that SbMT-2 may involve in the selective translocation of Zn++ from root to stem. This observation was further supported by the up-regulation of zinc transporter encoding genes NtZIP1 and NtHMA-A under metal ion stress condition. The study suggested that SbMT-2 modulates ROS scavenging and is a potential candidate to be used for phytoremediation and imparting stress tolerance.

  3. Overexpression of copper/zinc-superoxide dismutase in transgenic mice markedly impairs regeneration and increases development of neuropathic pain after sciatic nerve injury.

    Science.gov (United States)

    Kotulska, Katarzyna; LePecheur, Marie; Marcol, Wiesław; Lewin-Kowalik, Joanna; Larysz-Brysz, Magdalena; Paly, Evelyn; Matuszek, Iwona; London, Jacqueline

    2006-10-01

    Despite the general capacity of peripheral nervous system to regenerate, peripheral nerve injury is often followed by incomplete recovery of function, sometimes with the burden of neuropathic pain. The mechanisms of both regeneration and nociception have not been clarified, but it is known that inflammatory reactions are involved. Cu/Zn-superoxide dismutase (SOD1) is an important scavenger protein that acts against oxidative stress. It has been shown to play an important role in apoptosis and inflammation. The aim of this study was to examine the role of SOD1 overexpression in peripheral nerve regeneration and neuropathic pain-related behavior in mice. Sciatic nerves of SOD1-overexpressing and FVB/N wild type-mice were transected and immediately resutured. Evaluation of motor and sensory function and autotomy was carried out during 4 weeks of followup. We found markedly worse sciatic function index outcome as well as more significant atrophy of denervated muscles in SOD1-overexpressing animals compared with wild type. Autotomy was markedly worse in SOD1 transgenic mice than in wild-type animals. Histological evaluation revealed that the intensity of regeneration features, including numbers of GAP-43-positive growth cones, Schwann cells, and macrophages in the distal stump of the transected nerve, was also decreased in transgenic mice. Neuroma formation at the injury site was significantly more prominent in this group. Taken together, our findings suggest that SOD1 overexpression is deleterious for nerve regeneration processes and aggravates neuropathic pain-like state in mice. This can be at least partially ascribed to disturbed inflammatory reactions at the injury site. Copyright 2006 Wiley-Liss, Inc.

  4. Overexpression of a Novel Apple NAC Transcription Factor Gene, MdNAC1, Confers the Dwarf Phenotype in Transgenic Apple (Malus domestica)

    Science.gov (United States)

    Jia, Dongfeng; Gong, Xiaoqing; Li, Mingjun; Li, Chao; Sun, Tingting

    2018-01-01

    Plant height is an important trait for fruit trees. The dwarf characteristic is commonly associated with highly efficient fruit production, a major objective when breeding for apple (Malus domestica). We studied the function of MdNAC1, a novel NAC transcription factor (TF) gene in apple related to plant dwarfing. Localized primarily to the nucleus, MdNAC1 has transcriptional activity in yeast cells. Overexpression of the gene results in a dwarf phenotype in transgenic apple plants. Their reduction in size is manifested by shorter, thinner stems and roots, and a smaller leaf area. The transgenics also have shorter internodes and fewer cells in the stems. Levels of endogenous abscisic acid (ABA) and brassinosteroid (BR) are lower in the transgenic plants, and expression is decreased for genes involved in the biosynthesis of those phytohormones. All of these findings demonstrate that MdNAC1 has a role in plants dwarfism, probably by regulating ABA and BR production. PMID:29702625

  5. Overexpression of BID in thyroids of transgenic mice increases sensitivity to iodine-induced autoimmune thyroiditis

    Science.gov (United States)

    2014-01-01

    Background BID functions as a bridge molecule between death-receptor and mitochondrial related apoptotic pathways to amplify apoptotic signaling. Our previous studies have demonstrated a substantial increase in BID expression in primary normal thyroid epithelia cells treated with inflammatory cytokines, including the combination of IFNγ and IL-1β or IFNγ and TNFα. The aim of this study was to determine whether an increase in BID expression in thyroid can induce autoimmune thyroiditis. Methods A transgenic mouse line that expresses human BID in thyroid cells was established by fusing a mouse thyroglobulin (Tg) promoter upstream of human BID (Tg-BID). We tested whether the increased expression of pro-apoptotic BID in thyroid would induce autoimmune thyroiditis, both in the presence and absence of 0.3% iodine water. Results Our data show that Tg-BID mice in a CBA/J (H-2 k) background do not spontaneously develop autoimmune thyroiditis for over a year. However, upon ingestion of iodine in the drinking water, autoimmune thyroiditis does develop in Tg-BID transgenic mice, as shown by a significant increase in anti-Tg antibody and mononuclear cell infiltration in the thyroid glands in 30% of mice tested. Serum T4 levels, however, were similar between iodine-treated Tg-BID transgenic mice and the wild type mice. Conclusions Our data demonstrate that increased thyroid expression of BID facilitates the development of autoimmune thyroiditis induced by iodine uptake. However, the overexpression of BID itself is not sufficient to initiate thyroiditis in CBA/J (H-2 k) mice. PMID:24957380

  6. Chickpea WRKY70 Regulates the Expression of a Homeodomain-Leucine Zipper (HD-Zip) I Transcription Factor CaHDZ12, which Confers Abiotic Stress Tolerance in Transgenic Tobacco and Chickpea.

    Science.gov (United States)

    Sen, Senjuti; Chakraborty, Joydeep; Ghosh, Prithwi; Basu, Debabrata; Das, Sampa

    2017-11-01

    Drought and salinity are the two major environmental constraints that severely affect global agricultural productivity. Plant-specific HD-Zip transcription factors are involved in plant growth, development and stress responses. In the present study, we explored the functional characteristics and regulation of a novel HD-Zip (I) gene from chickpea, CaHDZ12, in response to water-deficit and salt-stress conditions. Transgenic tobacco lines over-expressing CaHDZ12 exhibited improved tolerance to osmotic stresses and increased sensitivity to abscisic acid (ABA). Physiological compatibility of transgenic lines was found to be more robust compared to the wild-type plants under drought and salinity stress. Additionally, expression of several stress-responsive genes was significantly induced in CaHDZ12 transgenic plants. On the other hand, silencing of CaHDZ12 in chickpea resulted in increased sensitivity to salt and drought stresses. Analysis of different promoter deletion mutants identified CaWRKY70 transcription factor as a transcriptional regulator of CaHDZ12 expression. In vivo and in vitro interaction studies detected an association between CaWRKY70 and CaHDZ12 promoter during stress responses. Epigenetic modifications underlying histone acetylation at the CaHDZ12 promoter region play a significant role in stress-induced activation of this gene. Collectively, our study describes a crucial and unique mechanistic link between two distinct transcription factors in regulating plant adaptive stress response. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  7. Biotechnological Reduction of Tobacco (Nicotiana Tabacum L. Toxicity

    Directory of Open Access Journals (Sweden)

    Samane Sattar

    2012-11-01

    Full Text Available Background: Nicotiana tobacco contains large amounts of alkaloid nicotine. Tobacco plant is used for smoking and causes many health problems since it is high in nicotine which is one of the widely-recognized toxic compounds with serious side effects for different body organs. Reducing nicotine content of this plant is a way to reduce its health hazards in cigarette smokers. Utilizing the new methods of genetic engineering can modify nicotine levels in the plant. In this study, through transferring the blocking gene, the pathway of nicotine biosynthesis was blocked to produce transgenic tobacco with low levels of nicotine. Methods: Transgenic plants carrying T DNA, and non-transgenic plants were grown on MS medium. Then their leaves were dried and powdered. The plants were extracted with alkali solution. Eventually, the nicotine content of the extract were analyzed using GC. Results: The analysis of extracts showed a reduction in the nicotine content of the transgenic plant (contain T-DNA in comparison with non-transgenic plants. Conclusion: Tobacco with lower nicotine reduction can reduce the toxic effects of smoking on smokers and can facilitate withdrawal from it.

  8. Overexpression of Nictaba-Like Lectin Genes from Glycine max Confers Tolerance towards Pseudomonas syringae Infection, Aphid Infestation and Salt Stress in Transgenic Arabidopsis Plants

    Directory of Open Access Journals (Sweden)

    Sofie Van Holle

    2016-10-01

    Full Text Available Plants have evolved a sophisticated immune system that allows them to recognize invading pathogens by specialized receptors. Carbohydrate-binding proteins or lectins are part of this immune system and especially the lectins that reside in the nucleocytoplasmic compartment are known to be implicated in biotic and abiotic stress responses. The class of Nictaba-like lectins (NLL groups all proteins with homology to the tobacco (Nicotiana tabacum lectin, known as a stress-inducible lectin. Here we focus on two Nictaba homologs from soybean (Glycine max, referred to as GmNLL1 and GmNLL2. Confocal laser scanning microscopy of fusion constructs with the green fluorescent protein either transiently expressed in Nicotiana benthamiana leaves or stably transformed in tobacco BY-2 suspension cells revealed a nucleocytoplasmic localization for the GmNLLs under study. RT-qPCR analysis of the transcript levels for the Nictaba-like lectins in soybean demonstrated that the genes are expressed in several tissues throughout the development of the plant. Furthermore, it was shown that salt treatment, Phytophthora sojae infection and Aphis glycines infestation trigger the expression of particular NLL genes. Stress experiments with Arabidopsis lines overexpressing the NLLs from soybean yielded an enhanced tolerance of the plant towards bacterial infection (Pseudomonas syringae, insect infestation (Myzus persicae and salinity. Our data showed a better performance of the transgenic lines compared to wild type plants, indicating that the NLLs from soybean are implicated in the stress response. These data can help to further elucidate the physiological importance of the Nictaba-like lectins from soybean, which can ultimately lead to the design of crop plants with a better tolerance to changing environmental conditions.

  9. Early thymic T cell development in young transgenic mice overexpressing human Cu/Zn superoxide dismutase, a model of Down syndrome.

    Science.gov (United States)

    Laurent, Julien; Paly, Evelyne; Marche, Patrice N; London, Jacqueline

    2006-06-01

    Previous studies have shown that transgenic mice overexpressing Cu/Zn superoxide dismutase, a model of Down syndrome, exhibit premature thymic involution. We have performed a flow cytometry analysis of the developing thymus in these homozygous transgenic mice (hSOD1/hSOD1: Tg-SOD). Longitudinal follow-up analysis from day 3 to day 280 showed an early thymic development in Tg-SOD mice compared with controls. This early thymic development was associated with an increased migration of mature T cells to peripheral lymphoid organs. BrdU labeling showed no difference between Tg-SOD and control mice, confirming that the greater number of peripheral T cells in Tg-SOD mice was not due to extensive proliferation of these cells but rather to a greater pool of emigrant T cells in Tg-SOD.

  10. Light-grown plants of transgenic tobacco expressing an introduced oat phytochrome A gene under the control of a constitutive viral promoter exhibit persistent growth inhibition by far-red light

    International Nuclear Information System (INIS)

    McCormac, A.; Whitelam, G.; Smith, H.

    1992-01-01

    A comparison of the photoregulation of development has been made for etiolated and light-grown plants of wild-type (WT) tobacco (Nicotiana tabacun L.) and an isogenic transgenic line which expresses an introduced oat phytochrome gene (phyA) under the control of a constitutive viral promoter. Etiolated seedlings of both the WT and transgenic line showed irradiance-dependent inhibition of hypocotyl growth under continuous far-red (FR) light; transgenic seedlings showed a greater level of inhibition under a given fluence rate and this is considered to be the result of the heterologous phytochrome protein (PhyA) functioning in a compatible manner with the native etiolated phytochrome. Deetiolation of WT seedlings resulted in a loss of responsiveness to prolonged FR. Light-grown transgenic seedlings, however, continued to respond in an irradiance-dependent manner to prolonged FR and it is proposed that this is a specific function of the constitutive PhyA. Mature green plants of the WT and transgenic lines showed a qualitatively similar growth promotion to a brief end-of-day FR-treatment but this response was abolished in the transgenic plants under prolonged irradiation by this same FR source. Growth inhibition (McCormac et al. 1991, Planta 185, 162-170) and enhanced levels of nitrate-reductase activity under irradiance of low red:far-red ratio, as achieved by the FR-supplementation of white light, emphasised that the introduced PhyA was eliciting an aberrant mode of photoresponse compared with the normal phytochrome population of light-grown plants. Total levels of the oat-encoded phytochrome in the etiolated transgenic tobacco were shown to be influenced by the wavelength of continuous irradiation in a manner which was qualitatively similar to that seen for the native, etiolated tobacco phytochrome, and distinct from that seen in etiolated oat tissues. These results are discussed in terms of the proposal that the constitutive oat-PhyA pool in the transgenic plants

  11. Salt tolerant SUV3 overexpressing transgenic rice plants conserve physicochemical properties and microbial communities of rhizosphere.

    Science.gov (United States)

    Sahoo, Ranjan K; Ansari, Mohammad W; Tuteja, Renu; Tuteja, Narendra

    2015-01-01

    Key concerns in the ecological evaluation of GM crops are undesirably spread, gene flow, other environmental impacts, and consequences on soil microorganism's biodiversity. Numerous reports have highlighted the effects of transgenic plants on the physiology of non-targeted rhizospheric microbes and the food chain via causing adverse effects. Therefore, there is an urgent need to develop transgenics with insignificant toxic on environmental health. In the present study, SUV3 overexpressing salt tolerant transgenic rice evaluated in New Delhi and Cuttack soil conditions for their effects on physicochemical and biological properties of rhizosphere. Its cultivation does not affect soil properties viz., pH, Eh, organic C, P, K, N, Ca, Mg, S, Na and Fe(2+). Additionally, SUV3 rice plants do not cause any change in the phenotype, species characteristics and antibiotic sensitivity of rhizospheric bacteria. The population and/or number of soil organisms such as bacteria, fungi and nematodes were unchanged in the soil. Also, the activity of bacterial enzymes viz., dehydrogenase, invertase, phenol oxidases, acid phosphatases, ureases and proteases was not significantly affected. Further, plant growth promotion (PGP) functions of bacteria such as siderophore, HCN, salicylic acid, IAA, GA, zeatin, ABA, NH3, phosphorus metabolism, ACC deaminase and iron tolerance were, considerably, not influenced. The present findings suggest ecologically pertinent of salt tolerant SUV3 rice to sustain the health and usual functions of the rhizospheric organisms. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Expression of the Native Cholera Toxin B Subunit Gene and Assembly as Functional Oligomers in Transgenic Tobacco Chloroplasts

    Science.gov (United States)

    Daniell, Henry; Lee, Seung-Bum; Panchal, Tanvi; Wiebe, Peter O.

    2012-01-01

    The B subunits of enterotoxigenic Escherichia coli (LTB) and cholera toxin of Vibrio cholerae (CTB) are candidate vaccine antigens. Integration of an unmodified CTB-coding sequence into chloroplast genomes (up to 10,000 copies per cell), resulted in the accumulation of up to 4.1% of total soluble tobacco leaf protein as functional oligomers (410-fold higher expression levels than that of the unmodified LTB gene expressed via the nuclear genome). However, expresssion levels reported are an underestimation of actual accumulation of CTB in transgenic chloroplasts, due to aggregation of the oligomeric forms in unboiled samples similar to the aggregation observed for purified bacterial antigen. PCR and Southern blot analyses confirmed stable integration of the CTB gene into the chloroplast genome. Western blot analysis showed that the chloroplast-synthesized CTB assembled into oligomers and were antigenically identical with purified native CTB. Also, binding assays confirmed that chloroplast- synthesized CTB binds to the intestinal membrane GM1-ganglioside receptor, indicating correct folding and disulfide bond formation of CTB pentamers within transgenic chloroplasts. In contrast to stunted nuclear transgenic plants, chloroplast transgenic plants were morphologically indistinguishable from untransformed plants, when CTB was constitutively expressed in chloroplasts. Introduced genes were inherited stably in subsequent generations, as confirmed by PCR and Southern blot analyses. Increased production of an efficient transmucosal carrier molecule and delivery system, like CTB, in transgenic chloroplasts makes plant-based oral vaccines and fusion proteins with CTB needing oral administration commercially feasible. Successful expression of foreign genes in transgenic chromoplasts and availability of marker-free chloroplast transformation techniques augurs well for development of vaccines in edible parts of transgenic plants. Furthermore, since the quaternary structure of

  13. Expression of TaWRKY44, a wheat WRKY gene, in transgenic tobacco confers multiple abiotic stress tolerances

    Directory of Open Access Journals (Sweden)

    Xiatian eWang

    2015-08-01

    Full Text Available The WRKY transcription factors have been reported to be involved in various plant physiological and biochemical processes. In this study, we successfully assembled ten unigenes from expressed sequence tags (ESTs of wheat and designated them as TaWRKY44–TaWRKY53, respectively. Among these genes, a subgroup I gene, TaWRKY44, was found to be upregulated by treatments with PEG6000, NaCl, 4°C, abscisic acid (ABA, H2O2 and gibberellin (GA. The TaWRKY44-GFP fusion protein was localized to the nucleus of onion epidermal cells, and TaWRKY44 was able to bind to the core DNA sequences of TTGACC and TTAACC in yeast. The N-terminal of TaWRKY44 showed transcriptional activation activity. Expression of TaWRKY44 in tobacco plants conferred drought and salt tolerance and transgenic tobacco exhibited a higher survival rate, relative water content (RWC, soluble sugar, proline and superoxide dismutase (SOD content, as well as higher activities of catalase (CAT and peroxidase (POD, but less ion leakage (IL, lower contents of malondialdehyde (MDA, and H2O2. In addition, expression of TaWRKY44 also increased the seed germination rate in the transgenic lines under osmotic stress conditions while exhibiting a lower H2O2 content and higher SOD, CAT and POD activities. Expression of TaWRKY44 upregulated the expression of some reactive oxygen species (ROS-related genes and stress-responsive genes in tobacco under osmotic stresses. These data demonstrate that TaWRKY44 may act as a positive regulator in drought/salt/osmotic stress responses by either efficient ROS elimination through direct or indirect activation of the cellular antioxidant systems or activation of stress-associated gene expression.

  14. Changes in melatonin levels in transgenic 'Micro-Tom' tomato overexpressing ovine AANAT and ovine HIOMT genes.

    Science.gov (United States)

    Wang, Lin; Zhao, Yu; Reiter, Russel J; He, Changjiu; Liu, Guoshi; Lei, Qiong; Zuo, Bixiao; Zheng, Xiao Dong; Li, Qingtian; Kong, Jin

    2014-03-01

    In animals, the melatonin biosynthesis pathway has been well defined after the isolation and identification of the four key genes that are involved in the conversion of tryptophan to melatonin. In plants, there are special alternative catalyzing steps, and plant genes share very low homology with the animal genes. It was of interest to examine the phenotype of transgenic Micro-Tom tomato plants overexpressing the homologous sheep oAANAT and oHIOMT genes responsible for the last two steps of melatonin synthesis. The oAANAT transgenic plants have higher melatonin levels and lower indoleacetic acid (IAA) contents than control due to the competition for tryptophan, the same precursor for both melatonin and IAA. Therefore, the oAANAT lines lose the 'apical dominance' inferring that melatonin likely lacks auxin activity. The significantly higher melatonin content in oHIOMT lines than oAANAT lines provides new proof for the important role of ASMT in plant melatonin synthesis. In addition, the enhanced drought tolerance of oHIOMT lines will also be an important contribution for plant engineering. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  15. Transcriptomic changes reveal gene networks responding to the overexpression of a blueberry DWARF AND DELAYED FLOWERING 1 gene in transgenic blueberry plants.

    Science.gov (United States)

    Song, Guo-Qing; Gao, Xuan

    2017-06-19

    Constitutive expression of the CBF/DREB1 for increasing freezing tolerance in woody plants is often associated with other phenotypic changes including dwarf plant and delayed flowering. These phenotypic changes have been observed when Arabidopsis DWARF AND DELAYED FLOWERING 1 (DDF1) was overexpressed in A. thaliana plants. To date, the DDF1 orthologues have not been studied in woody plants. The aim of this study is to investigate transcriptomic responses to the overexpression of blueberry (Vaccinium corymbosum) DDF1 (herein, VcDDF1-OX). The VcDDF1-OX resulted in enhanced freezing tolerance in tetraploid blueberry plants and did not result in significant changes in plant size, chilling requirement, and flowering time. Comparative transcriptome analysis of transgenic 'Legacy-VcDDF1-OX' plants containing an overexpressed VcDDF1 with non-transgenic highbush blueberry 'Legacy' plants revealed the VcDDF1-OX derived differentially expressed (DE) genes and transcripts in the pathways of cold-response, plant flowering, DELLA proteins, and plant phytohormones. The increase in freezing tolerance was associated to the expression of cold-regulated genes (CORs) and the ethylene pathway genes. The unchanged plant size, dormancy and flowering were due to the minimal effect of the VcDDF1-OX on the expression of DELLA proteins, flowering pathway genes, and the other phytohormone genes related to plant growth and development. The DE genes in auxin and cytokinin pathways suggest that the VcDDF1-OX has also altered plant tolerance to drought and high salinity. A DDF1 orthologue in blueberry functioned differently from the DDF1 reported in Arabidopsis. The overexpression of VcDDF1 or its orthologues is a new approach to increase freezing tolerance of deciduous woody plant species with no obvious effect on plant size and plant flowering time.

  16. Overexpression of NtWRKY50 Increases Resistance to Ralstonia solanacearum and Alters Salicylic Acid and Jasmonic Acid Production in Tobacco.

    Science.gov (United States)

    Liu, Qiuping; Liu, Ying; Tang, Yuanman; Chen, Juanni; Ding, Wei

    2017-01-01

    WRKY transcription factors (TFs) modulate plant responses to biotic and abiotic stresses. Here, we characterized a WRKY IIc TF, NtWRKY50, isolated from tobacco ( Nicotiana tabacum ) plants. The results showed that NtWRKY50 is a nuclear-localized protein and that its gene transcript is induced in tobacco when inoculated with the pathogenic bacterium Ralstonia solanacearum . Overexpression of NtWRKY50 enhanced bacterial resistance, which correlated with enhanced SA and JA/ET signaling genes. However, silencing of the NtWRKY50 gene had no obvious effects on plant disease resistance, implying functional redundancy of NtWRKY50 with other TFs. In addition, it was found that NtWRKY50 can be induced by various biotic or abiotic stresses, such as Potato virus Y, Rhizoctonia solani, Phytophthora parasitica , hydrogen peroxide, heat, cold, and wounding as well as the hormones salicylic acid (SA), jasmonic acid (JA), and ethylene (ET). Importantly, additional analysis suggests that NtWRKY50 overexpression markedly promotes SA levels but prevents pathogen-induced JA production. These data indicate that NtWRKY50 overexpression leads to altered SA and JA content, increased expression of defense-related genes and enhanced plant resistance to R. solanacearum. These probably due to increased activity of endogenous NtWRKY50 gene or could be gain-of-function phenotypes by altering the profile of genes affected by NtWRKY50 .

  17. An active role for endogenous beta-1,3-glucanase genes in transgene-mediated co-suppression in tobacco.

    Science.gov (United States)

    Sanders, Matthew; Maddelein, Wendy; Depicker, Anna; Van Montagu, Marc; Cornelissen, Marc; Jacobs, John

    2002-11-01

    Post-transcriptional gene silencing (PTGS) is characterized by the accumulation of short interfering RNAs that are proposed to mediate sequence-specific degradation of cognate and secondary target mRNAs. In plants, it is unclear to what extent endogenous genes contribute to this process. Here, we address the role of the endogenous target genes in transgene-mediated PTGS of beta-1,3-glucanases in tobacco. We found that mRNA sequences of the endogenous glucanase glb gene with varying degrees of homology to the Nicotiana plumbaginifolia gn1 transgene are targeted by the silencing machinery, although less efficiently than corresponding transgene regions. Importantly, we show that endogene-specific nucleotides in the glb sequence provide specificity to the silencing process. Consistent with this finding, small sense and antisense 21- to 23-nucleotide RNAs homologous to the endogenous glb gene were detected. Combined, these data demonstrate that a co-suppressed endogenous glucan ase gene is involved in signal amplification and selection of homologous targets, and show that endogenous genes can actively participate in PTGS in plants. The findings are introduced as a further sophistication of the post-transciptional silencing model.

  18. Isolation and characterization of a Vitis vinifera transcription factor, VvWRKY1, and its effect on responses to fungal pathogens in transgenic tobacco plants.

    Science.gov (United States)

    Marchive, Chloé; Mzid, Rim; Deluc, Laurent; Barrieu, François; Pirrello, Julien; Gauthier, Adrien; Corio-Costet, Marie-France; Regad, Farid; Cailleteau, Bernard; Hamdi, Saïd; Lauvergeat, Virginie

    2007-01-01

    Pathogen attack represents a major problem for viticulture and for agriculture in general. At present, the use of phytochemicals is more and more restrictive, and therefore it is becoming essential to control disease by having a thorough knowledge of resistance mechanisms. The present work focused on the trans-regulatory proteins potentially involved in the control of the plant defence response, the WRKY proteins. A full-length cDNA, designated VvWRKY1, was isolated from a grape berry library (Vitis vinifera L. cv. Cabernet Sauvignon). It encodes a polypeptide of 151 amino acids whose structure is characteristic of group IIc WRKY proteins. VvWRKY1 gene expression in grape is regulated in a developmental manner in berries and leaves and by various signal molecules involved in defence such as salicylic acid, ethylene, and hydrogen peroxide. Biochemical analysis indicates that VvWRKY1 specifically interacts with the W-box in various nucleotidic contexts. Functional analysis of VvWRKY1 was performed by overexpression in tobacco, and transgenic plants exhibited reduced susceptibility to various fungi but not to viruses. These results are consistent with a possible role for VvWRKY1 in grapevine defence against fungal pathogens.

  19. Transgenic APP expression during postnatal development causes persistent locomotor hyperactivity in the adult.

    Science.gov (United States)

    Rodgers, Shaefali P; Born, Heather A; Das, Pritam; Jankowsky, Joanna L

    2012-06-18

    Transgenic mice expressing disease-associated proteins have become standard tools for studying human neurological disorders. Transgenes are often expressed using promoters chosen to drive continuous high-level expression throughout life rather than temporal and spatial fidelity to the endogenous gene. This approach has allowed us to recapitulate diseases of aging within the two-year lifespan of the laboratory mouse, but has the potential for creating aberrant phenotypes by mechanisms unrelated to the human disorder. We show that overexpression of the Alzheimer's-related amyloid precursor protein (APP) during early postnatal development leads to severe locomotor hyperactivity that can be significantly attenuated by delaying transgene onset until adulthood. Our data suggest that exposure to transgenic APP during maturation influences the development of neuronal circuits controlling motor activity. Both when matched for total duration of APP overexpression and when matched for cortical amyloid burden, animals exposed to transgenic APP as juveniles are more active in locomotor assays than animals in which APP overexpression was delayed until adulthood. In contrast to motor activity, the age of APP onset had no effect on thigmotaxis in the open field as a rough measure of anxiety, suggesting that the interaction between APP overexpression and brain development is not unilateral. Our findings indicate that locomotor hyperactivity displayed by the tet-off APP transgenic mice and several other transgenic models of Alzheimer's disease may result from overexpression of mutant APP during postnatal brain development. Our results serve as a reminder of the potential for unexpected interactions between foreign transgenes and brain development to cause long-lasting effects on neuronal function in the adult. The tet-off APP model provides an easy means of avoiding developmental confounds by allowing transgene expression to be delayed until the mice reach adulthood.

  20. Inhibition of lipolysis in the novel transgenic quail model overexpressing G0/G1 switch gene 2 in the adipose tissue during feed restriction.

    Directory of Open Access Journals (Sweden)

    Sangsu Shin

    Full Text Available In addition to the issue of obesity in humans, the production of low-fat meat from domestic animals is important in the agricultural industry to satisfy consumer demand. Understanding the regulation of lipolysis in adipose tissue could advance our knowledge to potentially solve both issues. Although the G0/G1 switch gene 2 (G0S2 was recently identified as an inhibitor of adipose triglyceride lipase (ATGL in vitro, its role in vivo has not been fully clarified. This study was conducted to investigate the role of G0S2 gene in vivo by using two independent transgenic quail lines during different energy conditions. Unexpectedly, G0S2 overexpression had a negligible effect on plasma NEFA concentration, fat cell size and fat pad weight under ad libitum feeding condition when adipose lipolytic activity is minimal. A two-week feed restriction in non-transgenic quail expectedly caused increased plasma NEFA concentration and dramatically reduced fat cell size and fat pad weight. Contrary, G0S2 overexpression under a feed restriction resulted in a significantly less elevation of plasma NEFA concentration and smaller reductions in fat pad weights and fat cell size compared to non-transgenic quail, demonstrating inhibition of lipolysis and resistance to loss of fat by G0S2. Excessive G0S2 inhibits lipolysis in vivo during active lipolytic conditions, such as food restriction and fasting, suggesting G0S2 as a potential target for treatment of obesity. In addition, transgenic quail are novel models for studying lipid metabolism and mechanisms of obesity.

  1. A semi-automated motion-tracking analysis of locomotion speed in the C. elegans transgenics overexpressing beta-amyloid in neurons

    Directory of Open Access Journals (Sweden)

    Kevin eMachino

    2014-07-01

    Full Text Available Multi-Worm Tracker (MWT is a real-time computer vision system that can simultaneously quantify motional patterns of multiple worms. MWT provides several behavioral parameters, including analysis of accurate real-time locomotion speed in the nematode, Caenorhabditis elegans. Here, we determined locomotion speed of the Alzheimer’s disease (AD transgenic strain that over-expresses human beta-amyloid1-42 (Aβ in the neurons. The MWT analysis showed that the AD strain logged a slower average speed than the wild type worms. The results may be consistent with the observation that the AD patients with dementia tend to show deficits in physical activities, including frequent falls. The AD strain showed reduced ability of the eggs to hatch and slowed hatching of the eggs. Thus, over-expression of Aβ in neurons causes negative effects on locomotion and hatchability. This study sheds light on new examples of detrimental effects that Aβ deposits can exhibit using C. elegans as a model system. The information gathered from this study indicates that the motion tracking analysis is a cost-effective, efficient way to assess the deficits of Aβ over-expression in the C. elegans system.

  2. Overexpression of Cholesteryl Ester Transfer Protein Increases Macrophage-Derived Foam Cell Accumulation in Atherosclerotic Lesions of Transgenic Rabbits

    Directory of Open Access Journals (Sweden)

    Shoucui Gao

    2017-01-01

    Full Text Available High levels of plasma high-density lipoprotein-cholesterol (HDL-C are inversely associated with the risk of atherosclerosis and other cardiovascular diseases; thus, pharmacological inhibition of cholesteryl ester transfer protein (CETP is considered to be a therapeutic method of raising HDL-C levels. However, many CETP inhibitors have failed to achieve a clinical benefit despite raising HDL-C. In the study, we generated transgenic (Tg rabbits that overexpressed the human CETP gene to examine the influence of CETP on the development of atherosclerosis. Both Tg rabbits and their non-Tg littermates were fed a high cholesterol diet for 16 weeks. Plasma lipids and body weight were measured every 4 weeks. Gross lesion areas of the aortic atherosclerosis along with lesional cellular components were quantitatively analyzed. Overexpression of human CETP did not significantly alter the gross atherosclerotic lesion area, but the number of macrophages in lesions was significantly increased. Overexpression of human CETP did not change the plasma levels of total cholesterol or low-density lipoprotein cholesterol but lowered plasma HDL-C and increased triglycerides. These data revealed that human CETP may play an important role in the development of atherosclerosis mainly by decreasing HDL-C levels and increasing the accumulation of macrophage-derived foam cells.

  3. Transgenic overexpression of ADAM12 suppresses muscle regeneration and aggravates dystrophy in aged mdx mice

    DEFF Research Database (Denmark)

    Jørgensen, Louise Helskov; Jensen, Charlotte Harken; Wewer, Ulla M

    2007-01-01

    mice (ADAM12(+)) after a knife cut lesion and observed that the regeneration process was significantly impaired. ADAM12 seemed to inhibit the satellite cell response and delay myoblast differentiation. These results discourage long-term therapeutic use of ADAM12. They also point to impaired...... effect of ADAM12 was suggested to be mediated via a membrane-stabilizing up-regulation of utrophin, alpha7B integrin, and dystroglycans. Ectopic ADAM12 expression in normal mouse skeletal muscle also improved regeneration after freeze injury, presumably by the same mechanism. Hence, it was suggested...... overexpressing ADAM12 (ADAM12(+)/mdx mice), even though their utrophin levels were mildly elevated compared with age-matched controls. Thus, membrane stabilization was not sufficient to provide protection during prolonged disease. Consequently, we reinvestigated skeletal muscle regeneration in ADAM12 transgenic...

  4. Constitutive expression of the K-domain of a Vaccinium corymbosum SOC1-like (VcSOC1-K) MADS-box gene is sufficient to promote flowering in tobacco.

    Science.gov (United States)

    Song, Guo-qing; Walworth, Aaron; Zhao, Dongyan; Hildebrandt, Britton; Leasia, Michael

    2013-11-01

    The K-domain of a blueberry-derived SOC1 -like gene promotes flowering in tobacco without negatively impacting yield, demonstrating potential for manipulation of flowering time in horticultural crops. The SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) and SOC1-likes, belonging to the MIKC(c) (type II) MADS-box gene subfamily, are major floral activators and integrators of plant flowering. Both MADS-domains and K (Keratin)-domains are highly conserved in MIKC(c)-type MADS proteins. While there are many reports on overexpression of intact MIKC(c)-type MADS-box genes, few studies have been conducted to investigate the effects of the K-domains. In this report, a 474-bp K-domain of Vaccinium SOC1-like (VcSOC1-K) was cloned from the cDNA library of the northern highbush blueberry (Vaccinium corymbosum L.). Functional analysis of the VcSOC1-K was conducted by ectopically expressing of 35S:VcSOC1-K in tobacco. Reverse transcription PCR confirmed expression of the VcSOC1-K in T0 plants. Phenotypically, T1 transgenic plants (10 T1 plants/event) flowered sooner after seeding, and were shorter with fewer leaves at the time of flowering, than nontransgenic plants; but seed pod production of transgenic plants was not significantly affected. These results demonstrate that overexpression of the K-domain of a MIKC(c)-type MADS-box gene alone is sufficient to promote early flowering and more importantly without affecting seed production.

  5. Hyperactive mutant of a wheat plasma membrane Na+/H+ antiporter improves the growth and salt tolerance of transgenic tobacco.

    Science.gov (United States)

    Zhou, Yang; Lai, Zesen; Yin, Xiaochang; Yu, Shan; Xu, Yuanyuan; Wang, Xiaoxiao; Cong, Xinli; Luo, Yuehua; Xu, Haixia; Jiang, Xingyu

    2016-12-01

    Wheat SOS1 (TaSOS1) activity could be relieved upon deletion of the C-terminal 168 residues (the auto-inhibitory domain). This truncated form of wheat SOS1 (TaSOS1-974) was shown to increase compensation (compared to wild-type TaSOS1) for the salt sensitivity of a yeast mutant strain, AXT3K, via increased Na + transportation out of cells during salinity stress. Expression of the plasma membrane proteins TaSOS1-974 or TaSOS1 improved the growth of transgenic tobacco plants compared with wild-type plants under normal conditions. However, plants expressing TaSOS1-974 grew better than TaSOS1-transformed plants. Upon salinity stress, Na + efflux and K + influx rates in the roots of transgenic plants expressing TaSOS1-974 or TaSOS1 were greater than those of wild-type plants. Furthermore, compared to TaSOS1-transgenic plants, TaSOS1-974-expressing roots showed faster Na + efflux and K + influx, resulting in less Na + and more K + accumulation in TaSOS1-974-transgenic plants compared to TaSOS1-transgenic and wild-type plants. TaSOS1-974-expressing plants had the lowest MDA content and electrolyte leakage among all tested plants, indicating that TaSOS1-974 might protect the plasma membrane against oxidative damage generated by salt stress. Overall, TaSOS1-974 conferred higher salt tolerance in transgenic plants compared to TaSOS1. Consistent with this result, transgenic plants expressing TaSOS1-974 showed a better growth performance than TaSOS1-expressing and wild-type plants under saline conditions. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  6. Cotton Ascorbate Oxidase Promotes Cell Growth in Cultured Tobacco Bright Yellow-2 Cells through Generation of Apoplast Oxidation

    Science.gov (United States)

    Li, Rong; Xin, Shan; Tao, Chengcheng; Jin, Xiang; Li, Hongbin

    2017-01-01

    Ascorbate oxidase (AO) plays an important role in cell growth through the modulation of reduction/oxidation (redox) control of the apoplast. Here, a cotton (Gossypium hirsutum) apoplastic ascorbate oxidase gene (GhAO1) was obtained from fast elongating fiber tissues. GhAO1 belongs to the multicopper oxidase (MCO) family and includes a signal peptide and several transmembrane regions. Analyses of quantitative real-time polymerase chain reaction (QRT-PCR) and enzyme activity showed that GhAO1 was expressed abundantly in 15-day post-anthesis (dpa) wild-type (WT) fibers in comparison with fuzzless-lintless (fl) mutant ovules. Subcellular distribution analysis in onion cells demonstrated that GhAO1 is localized in the cell wall. In transgenic tobacco bright yellow-2 (BY-2) cells with ectopic overexpression of GhAO1, the enhancement of cell growth with 1.52-fold increase in length versus controls was indicated, as well as the enrichment of both total ascorbate in whole-cells and dehydroascorbate acid (DHA) in apoplasts. In addition, promoted activities of AO and monodehydroascorbate reductase (MDAR) in apoplasts and dehydroascorbate reductase (DHAR) in whole-cells were displayed in transgenic tobacco BY-2 cells. Accumulation of H2O2, and influenced expressions of Ca2+ channel genes with the activation of NtMPK9 and NtCPK5 and the suppression of NtTPC1B were also demonstrated in transgenic tobacco BY-2 cells. Finally, significant induced expression of the tobacco NtAO gene in WT BY-2 cells under indole-3-acetic acid (IAA) treatment appeared; however, the sensitivity of the NtAO gene expression to IAA disappeared in transgenic BY-2 cells, revealing that the regulated expression of the AO gene is under the control of IAA. Taken together, these results provide evidence that GhAO1 plays an important role in fiber cell elongation and may promote cell growth by generating the oxidation of apoplasts, via the auxin-mediated signaling pathway. PMID:28644407

  7. Over-Expression of Porcine Myostatin Missense Mutant Leads to A Gender Difference in Skeletal Muscle Growth between Transgenic Male and Female Mice.

    Science.gov (United States)

    Ma, Dezun; Gao, Pengfei; Qian, Lili; Wang, Qingqing; Cai, Chunbo; Jiang, Shengwang; Xiao, Gaojun; Cui, Wentao

    2015-08-24

    Myostatin, a transforming growth factor-β family member, is a negative regulator of skeletal muscle development and growth. Piedmontese cattle breeds have a missense mutation, which results in a cysteine to tyrosine substitution in the mature myostatin protein (C313Y). This loss-of-function mutation in myostatin results in a double-muscled phenotype in cattle. Myostatin propeptide is an inhibitor of myostatin activity and is considered a potential agent to stimulate muscle growth in livestock. In this study, we generated transgenic mice overexpressing porcine myostatin missense mutant (pmMS), C313Y, and wild-type porcine myostatin propeptide (ppMS), respectively, to examine their effects on muscle growth in mice. Enhanced muscle growth was observed in both pmMS and ppMS transgenic female mice and also in ppMS transgenic male mice. However, there was no enhanced muscle growth observed in pmMS transgenic male mice. To explore why there is such a big difference in muscle growth between pmMS and ppMS transgenic male mice, the expression level of androgen receptor (AR) mutant AR45 was measured by Western blot. Results indicated that AR45 expression significantly increased in pmMS transgenic male mice while it decreased dramatically in ppMS transgenic male mice. Our data demonstrate that both pmMS and ppMS act as myostatin inhibitors in the regulation of muscle growth, but the effect of pmMS in male mice is reversed by an increased AR45 expression. These results provide useful insight and basic theory to future studies on improving pork quality by genetically manipulating myostatin expression or by regulating myostatin activity.

  8. Resolving the role of plant glutamate dehydrogenase: II. Physiological characterization of plants overexpressing the two enzyme subunits individually or simultaneously.

    Science.gov (United States)

    Tercé-Laforgue, Thérèse; Bedu, Magali; Dargel-Grafin, Céline; Dubois, Frédéric; Gibon, Yves; Restivo, Francesco M; Hirel, Bertrand

    2013-10-01

    Glutamate dehydrogenase (GDH; EC 1.4.1.2) is able to carry out the deamination of glutamate in higher plants. In order to obtain a better understanding of the physiological function of GDH in leaves, transgenic tobacco (Nicotiana tabacum L.) plants were constructed that overexpress two genes from Nicotiana plumbaginifolia (GDHA and GDHB under the control of the Cauliflower mosiac virus 35S promoter), which encode the α- and β-subunits of GDH individually or simultaneously. In the transgenic plants, the GDH protein accumulated in the mitochondria of mesophyll cells and in the mitochondria of the phloem companion cells (CCs), where the native enzyme is normally expressed. Such a shift in the cellular location of the GDH enzyme induced major changes in carbon and nitrogen metabolite accumulation and a reduction in growth. These changes were mainly characterized by a decrease in the amount of sucrose, starch and glutamine in the leaves, which was accompanied by an increase in the amount of nitrate and Chl. In addition, there was an increase in the content of asparagine and a decrease in proline. Such changes may explain the lower plant biomass determined in the GDH-overexpressing lines. Overexpressing the two genes GDHA and GDHB individually or simultaneously induced a differential accumulation of glutamate and glutamine and a modification of the glutamate to glutamine ratio. The impact of the metabolic changes occurring in the different types of GDH-overexpressing plants is discussed in relation to the possible physiological function of each subunit when present in the form of homohexamers or heterohexamers.

  9. Enhancement of naphthalene tolerance in transgenic Arabidopsis plants overexpressing the ferredoxin-like protein (ADI1) from rice.

    Science.gov (United States)

    Fu, Xiao-Yan; Zhu, Bo; Han, Hong-Juan; Zhao, Wei; Tian, Yong-Sheng; Peng, Ri-He; Yao, Quan-Hong

    2016-01-01

    The ADI1 Arabidopsis plants enhanced tolerance and degradation efficiency to naphthalene and had great potential for phytoremediation of naphthalene in the plant material before composting or harvesting and removal. Naphthalene is a global environmental concern, because this substance is assumed to contribute considerably to human cancer risk. Cleaning up naphthalene contamination in the environment is crucial. Phytoremediation is an efficient technology to clean up contaminants. However, no gene that can efficiently degrade exogenous recalcitrant naphthalene in plants has yet been discovered. Ferredoxin (Fd) is a key player of biological electron transfer reaction in the PAH degradation process. The biochemical pathway for bacterial degradation of naphthalene has been well investigated. In this study, a rice gene, ADI1, which codes for a putative photosynthetic-type Fd, has been transformed into Arabidopsis thaliana. The transgenic Arabidopsis plants enhanced tolerance and degradation efficiency of naphthalene. Compared with wild-type plants, transgenic plants assimilated naphthalene from the culture media faster and removed more of this substance. When taken together, our findings suggest that breeding plants with overexpressed ADI1 gene is an effective strategy to degrade naphthalene in the environment.

  10. APP/SOD1 overexpressing mice present reduced neuropathic pain sensitivity.

    Science.gov (United States)

    Kotulska, Katarzyna; Larysz-Brysz, Magdalena; LePecheur, Marie; Marcol, Wiesław; Olakowska, Edyta; Lewin-Kowalik, Joanna; London, Jacqueline

    2011-07-15

    There are controversies regarding pain expression in mentally disabled people, including Down syndrome patients. The aim of this study was to examine neuropathic pain-related behavior and peripheral nerve regeneration in mouse model of Down syndrome. Sciatic nerves of double transgenic mice, overexpressing both amyloid precursor protein (APP) and Cu/Zn superoxide dismutase (SOD1) genes, and FVB/N wild type mice were transected and immediately resutured. Evaluation of autotomy and functional recovery was carried out during 4-week follow-up. We found markedly less severe autotomy in transgenic animals, although the onset of autotomy was significantly delayed in control mice. Interestingly, neuroma formation at the injury site was significantly more prominent in transgenic animals. Sciatic function index outcome was better in transgenic mice than in wild-type group. Histological evaluation revealed no statistically significant differences in the number of GAP-43-positive growth cones and macrophages in the distal stump of the transected nerve between groups. However, in transgenic animals, the regenerating axons were arranged more chaotically. The number of Schwann cells in the distal stump of the transected nerves was significantly lower in transgenic mice. The number of surviving motoneurons was markedly decreased in transgenic group. We measured also the atrophy of denervated muscles and found it decreased in APP/SOD1 overexpressing mice. Taken together, in this model of Down syndrome, we observed increased neuroma formation and decreased autotomy after peripheral nerve injury. Our findings suggest that APP/SOD1 overexpressing mice are less sensitive for neuropathic pain associated with neuroma. Copyright © 2011 Elsevier Inc. All rights reserved.

  11. Transgenic Mice Overexpressing Vitamin D Receptor (VDR) Show Anti-Inflammatory Effects in Lung Tissues.

    Science.gov (United States)

    Ishii, Masaki; Yamaguchi, Yasuhiro; Isumi, Kyoko; Ogawa, Sumito; Akishita, Masahiro

    2017-12-01

    Vitamin D insufficiency is increasingly recognized as a prevalent problem worldwide, especially in patients with a chronic lung disease. Chronic obstructive pulmonary disease (COPD) is a type of chronic inflammatory lung disease. Previous clinical studies have shown that COPD leads to low vitamin D levels, which further increase the severity of COPD. Vitamin D homeostasis represents one of the most important factors that potentially determine the severity of COPD. Nonetheless, the mechanisms underlying the anti-inflammatory effects of vitamin D receptor (VDR) in lung tissues are still unclear. To investigate the anti-inflammatory effects of VDR, we generated transgenic mice that show lung-specific VDR overexpression under the control of the surfactant protein C promoter (TG mice). The TG mice were used to study the expression patterns of proinflammatory cytokines using real-time polymerase chain reaction and immunohistochemistry. The TG mice had lower levels of T helper 1 (Th1)-related cytokines than wild-type (WT) mice did. No significant differences in the expression of Th2 cytokines were observed between TG and WT mice. This study is the first to achieve lung-specific overexpression of VDR in TG mice: an interesting animal model useful for studying the relation between airway cell inflammation and vitamin D signaling. VDR expression is an important factor that influences anti-inflammatory responses in lung tissues. Our results show the crucial role of VDR in anti-inflammatory effects in lungs; these data are potentially useful for the treatment or prevention of COPD.

  12. Light Intensity-Dependent Modulation of Chlorophyll b Biosynthesis and Photosynthesis by Overexpression of Chlorophyllide a Oxygenase in Tobacco1[C][OA

    Science.gov (United States)

    Biswal, Ajaya K.; Pattanayak, Gopal K.; Pandey, Shiv S.; Leelavathi, Sadhu; Reddy, Vanga S.; Govindjee; Tripathy, Baishnab C.

    2012-01-01

    Chlorophyll b is synthesized by the oxidation of a methyl group on the B ring of a tetrapyrrole molecule to a formyl group by chlorophyllide a oxygenase (CAO). The full-length CAO from Arabidopsis (Arabidopsis thaliana) was overexpressed in tobacco (Nicotiana tabacum) that grows well at light intensities much higher than those tolerated by Arabidopsis. This resulted in an increased synthesis of glutamate semialdehyde, 5-aminolevulinic acid, magnesium-porphyrins, and chlorophylls. Overexpression of CAO resulted in increased chlorophyll b synthesis and a decreased chlorophyll a/b ratio in low light-grown as well as high light-grown tobacco plants; this effect, however, was more pronounced in high light. The increased potential of the protochlorophyllide oxidoreductase activity and chlorophyll biosynthesis compensated for the usual loss of chlorophylls in high light. Increased chlorophyll b synthesis in CAO-overexpressed plants was accompanied not only by an increased abundance of light-harvesting chlorophyll proteins but also of other proteins of the electron transport chain, which led to an increase in the capture of light as well as enhanced (40%–80%) electron transport rates of photosystems I and II at both limiting and saturating light intensities. Although the quantum yield of carbon dioxide fixation remained unchanged, the light-saturated photosynthetic carbon assimilation, starch content, and dry matter accumulation increased in CAO-overexpressed plants grown in both low- and high-light regimes. These results demonstrate that controlled up-regulation of chlorophyll b biosynthesis comodulates the expression of several thylakoid membrane proteins that increase both the antenna size and the electron transport rates and enhance carbon dioxide assimilation, starch content, and dry matter accumulation. PMID:22419827

  13. Overexpression of a cotton (Gossypium hirsutum) WRKY gene, GhWRKY34, in Arabidopsis enhances salt-tolerance of the transgenic plants.

    Science.gov (United States)

    Zhou, Li; Wang, Na-Na; Gong, Si-Ying; Lu, Rui; Li, Yang; Li, Xue-Bao

    2015-11-01

    Soil salinity is one of the most serious threats in world agriculture, and often influences cotton growth and development, resulting in a significant loss in cotton crop yield. WRKY transcription factors are involved in plant response to high salinity stress, but little is known about the role of WRKY transcription factors in cotton so far. In this study, a member (GhWRKY34) of cotton WRKY family was functionally characterized. This protein containing a WRKY domain and a zinc-finger motif belongs to group III of cotton WRKY family. Subcellular localization assay indicated that GhWRKY34 is localized to the cell nucleus. Overexpression of GhWRKY34 in Arabidopsis enhanced the transgenic plant tolerance to salt stress. Several parameters (such as seed germination, green cotyledons, root length and chlorophyll content) in the GhWRKY34 transgenic lines were significantly higher than those in wild type under NaCl treatment. On the contrary, the GhWRKY34 transgenic plants exhibited a substantially lower ratio of Na(+)/K(+) in leaves and roots dealing with salt stress, compared with wild type. Growth status of the GhWRKY34 transgenic plants was much better than that of wild type under salt stress. Expressions of the stress-related genes were remarkably up-regulated in the transgenic plants under salt stress, compared with those in wild type. Based on the data presented in this study, we hypothesize that GhWRKY34 as a positive transcription regulator may function in plant response to high salinity stress through maintaining the Na(+)/K(+) homeostasis as well as activating the salt stress-related genes in cells. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  14. Overexpression of Pyrabactin Resistance-Like Abscisic Acid Receptors Enhances Drought, Osmotic, and Cold Tolerance in Transgenic Poplars

    Directory of Open Access Journals (Sweden)

    Jingling Yu

    2017-10-01

    Full Text Available Abscisic acid (ABA has been known participate in a wider range of adaptive responses to diverse environmental abiotic stresses such as drought, osmosis, and low temperatures. ABA signaling is initiated by its receptors PYR/PYL/RCARs, a type of soluble proteins with a conserved START domain which can bind ABA and trigger the downstream pathway. Previously, we discovered that poplar (Populus trichocarpa genome encodes 14 PYR/PYL/RCAR orthologs (PtPYRLs, and two of them, PtPYRL1 and PtPYRL5 have been functionally characterized to positively regulate drought tolerance. However, the physiological function of these ABA receptors in poplar remains uncharacterized. Here, we generated transgenic poplar plants overexpressing PtPYRL1 and PtPYRL5 and found that they exhibited more vigorous growth and produced greater biomass when exposed to drought stress. The improved drought tolerance was positively correlated with the key physiological responses dictated by the ABA signaling pathway, including increase in stomatal closure and decrease in leaf water loss. Further analyses revealed that overexpression lines showed improved capacity in scavenging reactive oxygen species and enhanced the activation of antioxidant enzymes under drought stress. Moreover, overexpression of PtPYRL1 or PtPYRL5 significantly increased the poplar resistance to osmotic and cold stresses. In summary, our results suggest that constitutive expression of PtPYRL1 and PtPYRL5 significantly enhances the resistance to drought, osmotic and cold stresses by positively regulating ABA signaling in poplar.

  15. Enhanced production of resveratrol derivatives in tobacco plants by improving the metabolic flux of intermediates in the phenylpropanoid pathway.

    Science.gov (United States)

    Jeong, Yu Jeong; An, Chul Han; Woo, Su Gyeong; Park, Ji Hye; Lee, Ki-Won; Lee, Sang-Hoon; Rim, Yeonggil; Jeong, Hyung Jae; Ryu, Young Bae; Kim, Cha Young

    2016-09-01

    The biosynthesis of flavonoids such as anthocyanin and stilbenes has attracted increasing attention because of their potential health benefits. Anthocyanins and stilbenes share common phenylpropanoid precursor pathways. We previously reported that the overexpression of sweetpotato IbMYB1a induced anthocyanin pigmentation in transgenic tobacco (Nicotiana tabacum) plants. In the present study, transgenic tobacco (Nicotiana tabacum SR1) plants (STS-OX and ROST-OX) expressing the RpSTS gene encoding stilbene synthase from rhubarb (Rheum palmatum L. cv. Jangyeop) and the RpSTS and VrROMT genes encoding resveratrol O-methyltransferase from frost grape (Vitis riparia) were generated under the control of 35S promoter. Phenotypic alterations in floral organs, such as a reduction in floral pigments and male sterility, were observed in STS-OX transgenic tobacco plants. However, we failed to obtain STS-OX and ROST-OX plants with high levels of resveratrol compounds. Therefore, to improve the production of resveratrol derivatives in plants, we cross-pollinated flowers of STS-OX or ROST-OX and IbMYB1a-OX transgenic lines (SM and RSM). Phenotypic changes in vegetative and reproductive development of SM and RSM plants were observed. Furthermore, by HPLC and LC-MS analyses, we found enhanced production of resveratrol derivatives such as piceid, piceid methyl ether, resveratrol methyl ether O-hexoside, and 5-methyl resveratrol-3,4'-O-β-D-diglucopyranoside in SM and RSM cross-pollinated lines. Here, total contents of trans- and cis-piceids ranged from approximately 104-240 µg/g fresh weight in SM (F2). Collectively, we suggest that coexpression of RpSTS and IbMYB1a via cross-pollination can induce enhanced production of resveratrol compounds in plants by increasing metabolic flux into stilbenoid biosynthesis.

  16. A chloroplast-localized and auxin-induced glutathione S-transferase from phreatophyte Prosopis juliflora confer drought tolerance on tobacco.

    Science.gov (United States)

    George, Suja; Venkataraman, Gayatri; Parida, Ajay

    2010-03-01

    Plant growth and productivity are adversely affected by various abiotic stress factors. In our previous study, we used Prosopis juliflora, a drought-tolerant tree species of Fabaceae, as a model plant system for mining genes functioning in abiotic stress tolerance. Large-scale random EST sequencing from a cDNA library obtained from drought-stressed leaves of 2-month-old P. juliflora plants resulted in identification of three different auxin-inducible glutathione S-transferases. In this paper, we report the cellular localization and the ability to confer drought tolerance in transgenic tobacco of one of these GSTs (PjGSTU1). PjGSTU1 was overexpressed in Escherichia coli and GST and GPX activities in total protein samples were assayed and compared with controls. The results indicated that PjGSTU1 protein forms a functional homo-dimer in recombinant bacteria with glutathione transferase as well as glutathione peroxidase activities. PjGSTU1 transgenic tobacco lines survived better under conditions of 15% PEG stress compared with control un-transformed plants. In vivo localization studies for PjGSTU1 using GFP fusion revealed protein localization in chloroplasts of transgenic plants. The peroxidase activity of PjGSTU1 and its localization in the chloroplast indicates a possible role for PjGSTU1 in ROS removal. Copyright 2009 Elsevier GmbH. All rights reserved.

  17. Altered Fruit and Seed Development of Transgenic Rapeseed (Brassica napus Over-Expressing MicroRNA394.

    Directory of Open Access Journals (Sweden)

    Jian Bo Song

    Full Text Available Fruit and seed development in plants is a complex biological process mainly involved in input and biosynthesis of many storage compounds such as proteins and oils. Although the basic biochemical pathways for production of the storage metabolites in plants are well characterized, their regulatory mechanisms are not fully understood. In this study, we functionally identified rapeseed (Brassica napus miR394 with its target gene Brassica napus leaf curling responsiveness (BnLCR to dissect a role of miR394 during the fruit and seed development. Transgenic rapeseed plants over-expressing miR394 under the control of the cauliflower mosaic virus 35S promoter were generated. miR394 over-expression plants exhibited a delayed flowering time and enlarged size of plants, leaf blade, pods and seed body, but developed seeds with higher contents of protein and glucosinolates (GLS and lower levels of oil accumulation as compared to wild-type. Over-expression of miR394 altered the fatty acid (FA composition by increasing several FA species such as C16:0 and C18:0 and unsaturated species of C20:1 and C22:1 but lowering C18:3. This change was accompanied by induction of genes coding for transcription factors of FA synthesis including leafy cotyledon1 (BnLEC1, BnLEC2, and FUSCA3 (FUS3. Because the phytohormone auxin plays a crucial role in fruit development and seed patterning, the DR5-GUS reporter was used for monitoring the auxin response in Arabidopsis siliques and demonstrated that the DR5 gene was strongly expressed. These results suggest that BnmiR394 is involved in rapeseed fruit and seed development.

  18. Heterologous expression of gentian MYB1R transcription factors suppresses anthocyanin pigmentation in tobacco flowers.

    Science.gov (United States)

    Nakatsuka, Takashi; Yamada, Eri; Saito, Misa; Fujita, Kohei; Nishihara, Masahiro

    2013-12-01

    Single-repeat MYB transcription factors, GtMYB1R1 and GtMYB1R9 , were isolated from gentian. Overexpression of these genes reduced anthocyanin accumulation in tobacco flowers, demonstrating their applicability to modification of flower color. RNA interference (RNAi) has recently been used to successfully modify flower color intensity in several plant species. In most floricultural plants, this technique requires prior isolation of target flavonoid biosynthetic genes from the same or closely related species. To overcome this limitation, we developed a simple and efficient method for reducing floral anthocyanin accumulation based on genetic engineering using novel transcription factor genes isolated from Japanese gentians. We identified two single-repeat MYB genes--GtMYB1R and GtMYB1R9--predominantly expressed in gentian petals. Transgenic tobacco plants expressing these genes were produced, and their flowers were analyzed for flavonoid components and expression of flavonoid biosynthetic genes. Transgenic tobacco plants expressing GtMYB1R1 or GtMYB1R9 exhibited significant reductions in floral anthocyanin accumulation, resulting in white-flowered phenotypes. Expression levels of chalcone isomerase (CHI), dihydroflavonol 4-reductase (DFR), and anthocyanidin synthase (ANS) genes were preferentially suppressed in these transgenic tobacco flowers. A yeast two-hybrid assay demonstrated that both GtMYB1R1 and GtMYB1R9 proteins interacted with the GtbHLH1 protein, previously identified as an anthocyanin biosynthesis regulator in gentian flowers. In addition, a transient expression assay indicated that activation of the gentian GtDFR promoter by the GtMYB3-GtbHLH1 complex was partly canceled by addition of GtMYB1R1 or GtMYB1R9. These results suggest that GtMYB1R1 and GtMYB1R9 act as antagonistic transcription factors of anthocyanin biosynthesis in gentian flowers. These genes should consequently be useful for manipulating anthocyanin accumulation via genetic engineering in

  19. Thioredoxin-1 overexpression in transgenic mice attenuates streptozotocin-induced diabetic osteopenia: a novel role of oxidative stress and therapeutic implications.

    Science.gov (United States)

    Hamada, Yasuhiro; Fujii, Hideki; Kitazawa, Riko; Yodoi, Junji; Kitazawa, Sohei; Fukagawa, Masafumi

    2009-05-01

    Diabetes mellitus is associated with increased risk of osteopenia and bone fracture. However, the mechanisms accounting for diabetic bone disorder are unclear. We have previously reported that streptozotocin-induced diabetic mice develop low turnover osteopenia associated with increased oxidative stress in the diabetic condition. To determine the role of oxidative stress in the development of diabetic osteopenia, we presently investigated the effect of overexpression of thioredoxin-1 (TRX), a major intracellular antioxidant, on the development of diabetic osteopenia, using TRX transgenic mice (TRX-Tg). TRX-Tg are C57BL/6 mice that carry the human TRX transgene under the control of beta-actin promoter. Eight-week-old male TRX-Tg mice and wild type (WT) littermates were intraperitoneally injected with either streptozotocin or vehicle. Mice were grouped as 1) non-diabetic WT, 2) non-diabetic TRX-Tg, 3) diabetic WT, and 4) diabetic TRX-Tg. After 12 weeks of streptozotocin treatment, oxidative stress on the whole body and bone was evaluated, and the physical properties of the femora, and histomorphometry parameters of the tibiae were assessed. TRX overexpression did not affect either body weight or hemoglobin A1c levels. There were no significant differences in renal function and in serum levels of calcium, phosphate, and intact parathyroid hormone among the four groups. On the other hand, urinary excretion of 8-hydroxydeoxyguanosine (8-OHdG), a marker of oxidative DNA damage, was significantly elevated in diabetic WT and attenuated in diabetic TRX-Tg. Immunohistochemical staining for 8-OHdG revealed marked intensity in the bone tissue of diabetic WT compared with non-diabetic WT, while staining was attenuated in diabetic TRX-Tg. TRX overexpression partially restored reduced bone mineral density and prevented the suppression of bone formation observed in diabetic WT. Increased oxidative stress in diabetic condition contributes to the development of diabetic osteopenia

  20. Targeted transgenic overexpression of mitochondrial thymidine kinase (TK2) alters mitochondrial DNA (mtDNA) and mitochondrial polypeptide abundance: transgenic TK2, mtDNA, and antiretrovirals.

    Science.gov (United States)

    Hosseini, Seyed H; Kohler, James J; Haase, Chad P; Tioleco, Nina; Stuart, Tami; Keebaugh, Erin; Ludaway, Tomika; Russ, Rodney; Green, Elgin; Long, Robert; Wang, Liya; Eriksson, Staffan; Lewis, William

    2007-03-01

    Mitochondrial toxicity limits nucleoside reverse transcriptase inhibitors (NRTIs) for acquired immune deficiency syndrome. NRTI triphosphates, the active moieties, inhibit human immunodeficiency virus reverse transcriptase and eukaryotic mitochondrial DNA polymerase pol-gamma. NRTI phosphorylation seems to correlate with mitochondrial toxicity, but experimental evidence is lacking. Transgenic mice (TGs) with cardiac overexpression of thymidine kinase isoforms (mitochondrial TK2 and cytoplasmic TK1) were used to study NRTI mitochondrial toxicity. Echocardiography and nuclear magnetic resonance imaging defined cardiac performance and structure. TK gene copy and enzyme activity, mitochondrial (mt) DNA and polypeptide abundance, succinate dehydrogenase and cytochrome oxidase histochemistry, and electron microscopy correlated with transgenesis, mitochondrial structure, and biogenesis. Antiretroviral combinations simulated therapy. Untreated hTK1 or TK2 TGs exhibited normal left ventricle mass. In TK2 TGs, cardiac TK2 gene copy doubled, activity increased 300-fold, and mtDNA abundance doubled. Abundance of the 17-kd subunit of complex I, succinate dehydrogenase histochemical activity, and cristae density increased. NRTIs increased left ventricle mass 20% in TK2 TGs. TK activity increased 3 logs in hTK1 TGs, but no cardiac phenotype resulted. NRTIs abrogated functional effects of transgenically increased TK2 activity but had no effect on TK2 mtDNA abundance. Thus, NRTI mitochondrial phosphorylation by TK2 is integral to clinical NRTI mitochondrial toxicity.

  1. Tetracycline-inducible system for regulation of skeletal muscle-specific gene expression in transgenic mice

    Science.gov (United States)

    Grill, Mischala A.; Bales, Mark A.; Fought, Amber N.; Rosburg, Kristopher C.; Munger, Stephanie J.; Antin, Parker B.

    2003-01-01

    Tightly regulated control of over-expression is often necessary to study one aspect or time point of gene function and, in transgenesis, may help to avoid lethal effects and complications caused by ubiquitous over-expression. We have utilized the benefits of an optimized tet-on system and a modified muscle creatine kinase (MCK) promoter to generate a skeletal muscle-specific, doxycycline (Dox) controlled over-expression system in transgenic mice. A DNA construct was generated in which the codon optimized reverse tetracycline transactivator (rtTA) was placed under control of a skeletal muscle-specific version of the mouse MCK promoter. Transgenic mice containing this construct expressed rtTA almost exclusively in skeletal muscles. These mice were crossed to a second transgenic line containing a bi-directional promoter centered on a tet responder element driving both a luciferase reporter gene and a tagged gene of interest; in this case the calpain inhibitor calpastatin. Compound hemizygous mice showed high level, Dox dependent muscle-specific luciferase activity often exceeding 10,000-fold over non-muscle tissues of the same mouse. Western and immunocytochemical analysis demonstrated similar Dox dependent muscle-specific induction of the tagged calpastatin protein. These findings demonstrate the effectiveness and flexibility of the tet-on system to provide a tightly regulated over-expression system in adult skeletal muscle. The MCKrtTA transgenic lines can be combined with other transgenic responder lines for skeletal muscle-specific over-expression of any target gene of interest.

  2. Salt tolerance and activity of antioxidative enzymes of transgenic finger millet overexpressing a vacuolar H(+)-pyrophosphatase gene (SbVPPase) from Sorghum bicolor.

    Science.gov (United States)

    Anjaneyulu, Ediga; Reddy, Palle Surender; Sunita, Merla Srilakshmi; Kishor, Polavarapu B Kavi; Meriga, Balaji

    2014-06-15

    A vacuolar proton pyrophosphatase cDNA clone was isolated from Sorghum bicolor (SbVPPase) using end-to-end gene-specific primer amplification. It showed 80-90% homology at the nucleotide and 85-95% homology at the amino acid level with other VPPases. The gene was introduced into expression vector pCAMBIA1301 under the control of the cauliflower mosaic virus 35S (CaMV35S) promoter and transformed into Agrobacterium tumifaciens strain LBA4404 to infect embryogenic calli of finger millet (Eleusine coracana). Successful transfer of SbVPPase was confirmed by a GUS histochemical assay and PCR analysis. Both, controls and transgenic plants were subjected to 100 and 200mM NaCl and certain biochemical and physiological parameters were studied. Relative water content (RWC), plant height, leaf expansion, finger length and width and grain weight were severely reduced (50-70%), and the flowering period was delayed by 20% in control plants compared to transgenic plants under salinity stress. With increasing salt stress, the proline and chlorophyll contents as well as the enzyme activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), guaiacol peroxidase (GPX) and glutathione reductase (GR) increased by 25-100% in transgenics, while malondialdehyde (MDA) showed a 2-4-fold decrease. The increased activities of antioxidant enzymes and the reduction in the MDA content suggest efficient scavenging of reactive oxygen species (ROS) in transgenics and, as a consequence, probably alleviation of salt stress. Also, the leaf tissues of the transgenics accumulated 1.5-2.5-fold higher Na(+) and 0.4-0.8-fold higher K(+) levels. Together, these results clearly demonstrate that overexpression of SbVPPase in transgenic finger millet enhances the plant's performance under salt stress. Copyright © 2014 Elsevier GmbH. All rights reserved.

  3. Overexpression of Thellungiella halophila H+-pyrophosphatase Gene Improves Low Phosphate Tolerance in Maize

    Science.gov (United States)

    Pei, Laming; Wang, Jiemin; Li, Kunpeng; Li, Yongjun; Li, Bei; Gao, Feng; Yang, Aifang

    2012-01-01

    Low phosphate availability is a major constraint on plant growth and agricultural productivity. Engineering a crop with enhanced low phosphate tolerance by transgenic technique could be one way of alleviating agricultural losses due to phosphate deficiency. In this study, we reported that transgenic maize plants that overexpressed the Thellungiella halophila vacuolar H+-pyrophosphatase gene (TsVP) were more tolerant to phosphate deficit stress than the wild type. Under phosphate sufficient conditions, transgenic plants showed more vigorous root growth than the wild type. When phosphate deficit stress was imposed, they also developed more robust root systems than the wild type, this advantage facilitated phosphate uptake, which meant that transgenic plants accumulated more phosphorus. So the growth and development in the transgenic maize plants were not damaged as much as in the wild type plants under phosphate limitation. Overexpression of TsVP increased the expression of genes involved in auxin transport, which indicated that the development of larger root systems in transgenic plants might be due in part to enhanced auxin transport which controls developmental events in plants. Moreover, transgenic plants showed less reproductive development retardation and a higher grain yield per plant than the wild type plants when grown in a low phosphate soil. The phenotypes of transgenic maize plants suggested that the overexpression of TsVP led to larger root systems that allowed transgenic maize plants to take up more phosphate, which led to less injury and better performance than the wild type under phosphate deficiency conditions. This study describes a feasible strategy for improving low phosphate tolerance in maize and reducing agricultural losses caused by phosphate deficit stress. PMID:22952696

  4. Transgenic plants with increased calcium stores

    Science.gov (United States)

    Wyatt, Sarah (Inventor); Tsou, Pei-Lan (Inventor); Robertson, Dominique (Inventor); Boss, Wendy (Inventor)

    2004-01-01

    The present invention provides transgenic plants over-expressing a transgene encoding a calcium-binding protein or peptide (CaBP). Preferably, the CaBP is a calcium storage protein and over-expression thereof does not have undue adverse effects on calcium homeostasis or biochemical pathways that are regulated by calcium. In preferred embodiments, the CaBP is calreticulin (CRT) or calsequestrin. In more preferred embodiments, the CaBP is the C-domain of CRT, a fragment of the C-domain, or multimers of the foregoing. In other preferred embodiments, the CaBP is localized to the endoplasmic reticulum by operatively associating the transgene encoding the CaBP with an endoplasmic reticulum localization peptide. Alternatively, the CaBP is targeted to any other sub-cellular compartment that permits the calcium to be stored in a form that is biologically available to the plant. Also provided are methods of producing plants with desirable phenotypic traits by transformation of the plant with a transgene encoding a CaBP. Such phenotypic traits include increased calcium storage, enhanced resistance to calcium-limiting conditions, enhanced growth and viability, increased disease and stress resistance, enhanced flower and fruit production, reduced senescence, and a decreased need for fertilizer production. Further provided are plants with enhanced nutritional value as human food or animal feed.

  5. Overexpression of an endo-1,4-β-glucanase V gene (EGV) from Trichoderma reesei leads to the accumulation of cellulase activity in transgenic rice.

    Science.gov (United States)

    Li, X Y; Liu, F; Hu, Y F; Xia, M; Cheng, B J; Zhu, S W; Ma, Q

    2015-12-21

    The ectopic expression of cellulase in biomass can reduce the cost of biofuel conversion. This trait modification technique is highly beneficial for biofuel production. In this study, we isolated an endo-1,4-beta-glucanase gene (EGV) from Trichoderma reesei and inserted this gene downstream of a fragment encoding the signal peptide Apo-SP in a modified pCAMBIA1301 vector to obtain an Apo-SP and AsRed fusion protein. Transient expression of this fusion protein in onion epidermal cells showed that the Apo-SP signal was localized to the plastids. EGV transgenic rice plants that did not carry screening marker genes were obtained through overexpression of the pDTB double T-DNA vector. Western blotting showed that EGV was expressed in the dry straw of T0 generation transgenic rice plants and in fresh leaves of the T1 generation. More importantly, our results also showed that the peptide product of EGV in the transgenic plants folded correctly and was capable of digesting the cellulase substrate CMC. Additionally, cellulase activity remained stable in the straw that had been dried at room temperature for three months. This study presents an important technical approach for the development of transgenic rice straw that has stable cellulase activity and can be used for biofuel conversion.

  6. MdATG18a overexpression improves tolerance to nitrogen deficiency and regulates anthocyanin accumulation through increased autophagy in transgenic apple.

    Science.gov (United States)

    Sun, Xun; Jia, Xin; Huo, Liuqing; Che, Runmin; Gong, Xiaoqing; Wang, Ping; Ma, Fengwang

    2018-02-01

    Nitrogen (N) availability is an essential factor for plant growth. Recycling and remobilization of N have strong impacts on crop yield and quality under N deficiency. Autophagy is a critical nutrient-recycling process that facilitates remobilization under starvation. We previously showed that an important AuTophaGy (ATG) protein from apple, MdATG18a, has a positive role in drought tolerance. In this study, we explored its biological role in response to low-N. Overexpression of MdATG18a in both Arabidopsis and apple improved tolerance to N-depletion and caused a greater accumulation of anthocyanin. The increased anthocyanin concentration in transgenic apple was possibly due to up-regulating flavonoid biosynthetic and regulatory genes (MdCHI, MdCHS, MdANS, MdPAL, MdUFGT, and MdMYB1) and higher soluble sugars concentration. MdATG18a overexpression enhanced starch degradation with up-regulating amylase gene (MdAM1) and up-regulated sugar metabolism related genes (MdSS1, MdHXKs, MdFK1, and MdNINVs). Furthermore, MdATG18a functioned in nitrate uptake and assimilation by up-regulating nitrate reductase MdNIA2 and 3 high-affinity nitrate transporters MdNRT2.1/2.4/2.5. MdATG18a overexpression also elevated other important MdATG genes expression and autophagosomes formation under N-depletion, which play key contributions to above changes. Together, these results demonstrate that overexpression of MdATG18a enhances tolerance to N-deficiencies and plays positive roles in anthocyanin biosynthesis through greater autophagic activity. © 2017 John Wiley & Sons Ltd.

  7. A novel bZIP gene from Tamarix hispida mediates physiological responses to salt stress in tobacco plants.

    Science.gov (United States)

    Wang, Yucheng; Gao, Caiqiu; Liang, Yenan; Wang, Chao; Yang, Chuanping; Liu, Guifeng

    2010-02-15

    Basic leucine zipper proteins (bZIPs) are transcription factors that bind abscisic acid (ABA)-responsive elements (ABREs) and enable plants to withstand adverse environmental conditions. In the present study, a novel bZIP gene, ThbZIP1 was cloned from Tamarix hispida. Expression studies in T. hispida showed differential regulation of ThbZIP1 in response to treatment with NaCl, polyethylene glycol (PEG) 6000, NaHCO(3), and CdCl(2), suggesting that ThbZIP1 is involved in abiotic stress responses. To identify the physiological responses mediated by ThbZIP1, transgenic tobacco plants overexpressing exogenous ThbZIP1 were generated. Various physiological parameters related to salt stress were measured and compared between transgenic and wild type (WT) plants. Our results indicate that overexpression of ThbZIP1 can enhance the activity of both peroxidase (POD) and superoxide dismutase (SOD), and increase the content of soluble sugars and soluble proteins under salt stress conditions. These results suggest that ThbZIP1 contributes to salt tolerance by mediating signaling through multiple physiological pathways. Furthermore, ThbZIP1 confers stress tolerance to plants by enhancing reactive oxygen species (ROS) scavenging, facilitating the accumulation of compatible osmolytes, and inducing and/or enhancing the biosynthesis of soluble proteins. Copyright 2009 Elsevier GmbH. All rights reserved.

  8. Regulatory approval and a first-in-human phase I clinical trial of a monoclonal antibody produced in transgenic tobacco plants.

    Science.gov (United States)

    Ma, Julian K-C; Drossard, Jürgen; Lewis, David; Altmann, Friedrich; Boyle, Julia; Christou, Paul; Cole, Tom; Dale, Philip; van Dolleweerd, Craig J; Isitt, Valerie; Katinger, Dietmar; Lobedan, Martin; Mertens, Hubert; Paul, Mathew J; Rademacher, Thomas; Sack, Markus; Hundleby, Penelope A C; Stiegler, Gabriela; Stoger, Eva; Twyman, Richard M; Vcelar, Brigitta; Fischer, Rainer

    2015-10-01

    Although plant biotechnology has been widely investigated for the production of clinical-grade monoclonal antibodies, no antibody products derived from transgenic plants have yet been approved by pharmaceutical regulators for clinical testing. In the Pharma-Planta project, the HIV-neutralizing human monoclonal antibody 2G12 was expressed in transgenic tobacco (Nicotiana tabacum). The scientific, technical and regulatory demands of good manufacturing practice (GMP) were addressed by comprehensive molecular characterization of the transgene locus, confirmation of genetic and phenotypic stability over several generations of transgenic plants, and by establishing standard operating procedures for the creation of a master seed bank, plant cultivation, harvest, initial processing, downstream processing and purification. The project developed specifications for the plant-derived antibody (P2G12) as an active pharmaceutical ingredient (API) based on (i) the guidelines for the manufacture of monoclonal antibodies in cell culture systems; (ii) the draft European Medicines Agency Points to Consider document on quality requirements for APIs produced in transgenic plants; and (iii) de novo guidelines developed with European national regulators. From the resulting process, a GMP manufacturing authorization was issued by the competent authority in Germany for transgenic plant-derived monoclonal antibodies for use in a phase I clinical evaluation. Following preclinical evaluation and ethical approval, a clinical trial application was accepted by the UK national pharmaceutical regulator. A first-in-human, double-blind, placebo-controlled, randomized, dose-escalation phase I safety study of a single vaginal administration of P2G12 was carried out in healthy female subjects. The successful completion of the clinical trial marks a significant milestone in the commercial development of plant-derived pharmaceutical proteins. © 2015 Society for Experimental Biology, Association of

  9. Transgenic plants with enhanced growth characteristics

    Energy Technology Data Exchange (ETDEWEB)

    Unkefer, Pat J.; Anderson, Penelope S.; Knight, Thomas J.

    2018-01-09

    The invention relates to transgenic plants exhibiting dramatically enhanced growth rates, greater seed and fruit/pod yields, earlier and more productive flowering, more efficient nitrogen utilization, increased tolerance to high salt conditions, and increased biomass yields. In one embodiment, transgenic plants engineered to over-express both glutamine phenylpyruvate transaminase (GPT) and glutamine synthetase (GS) are provided. The GPT+GS double-transgenic plants of the invention consistently exhibit enhanced growth characteristics, with T0 generation lines showing an increase in biomass over wild type counterparts of between 50% and 300%. Generations that result from sexual crosses and/or selfing typically perform even better, with some of the double-transgenic plants achieving an astounding four-fold biomass increase over wild type plants.

  10. Transgenic plants with enhanced growth characteristics

    Energy Technology Data Exchange (ETDEWEB)

    Unkefer, Pat J.; Anderson, Penelope S.; Knight, Thomas J.

    2016-09-06

    The invention relates to transgenic plants exhibiting dramatically enhanced growth rates, greater seed and fruit/pod yields, earlier and more productive flowering, more efficient nitrogen utilization, increased tolerance to high salt conditions, and increased biomass yields. In one embodiment, transgenic plants engineered to over-express both glutamine phenylpyruvate transaminase (GPT) and glutamine synthetase (GS) are provided. The GPT+GS double-transgenic plants of the invention consistently exhibit enhanced growth characteristics, with T0 generation lines showing an increase in biomass over wild type counterparts of between 50% and 300%. Generations that result from sexual crosses and/or selfing typically perform even better, with some of the double-transgenic plants achieving an astounding four-fold biomass increase over wild type plants.

  11. Overexpression of antioxidant enzymes in diaphragm muscle does not alter contraction-induced fatigue or recovery

    Science.gov (United States)

    McClung, Joseph M.; DeRuisseau, Keith C.; Whidden, Melissa A.; Van Remmen, Holly; Richardson, Arlan; Song, Wook; Vrabas, Ioannis S.; Powers, Scott K.

    2010-01-01

    Low levels of reactive oxygen species (ROS) production are necessary to optimize muscle force production in unfatigued muscle. In contrast, sustained high levels of ROS production have been linked to impaired muscle force production and contraction-induced skeletal muscle fatigue. Using genetically engineered mice, we tested the hypothesis that the independent transgenic overexpression of catalase (CAT), copper/zinc superoxide dismutase (CuZnSOD; SOD1) or manganese superoxide dismutase (MnSOD; SOD2) antioxidant enzymes would negatively affect force production in unfatigued diaphragm muscle but would delay the development of muscle fatigue and enhance force recovery after fatiguing contractions. Diaphragm muscle from wild-type littermates (WT) and from CAT, SOD1 and SOD2 overexpressing mice were subjected to an in vitro contractile protocol to investigate the force–frequency characteristics, the fatigue properties and the time course of recovery from fatigue. The CAT, SOD1 and SOD2 overexpressors produced less specific force (in N cm−2) at stimulation frequencies of 20–300 Hz and produced lower maximal tetanic force than WT littermates. The relative development of muscle fatigue and recovery from fatigue were not influenced by transgenic overexpression of any antioxidant enzyme. Morphologically, the mean cross-sectional area (in μm2) of diaphragm myofibres expressing myosin heavy chain type IIA was decreased in both CAT and SOD2 transgenic animals, and the percentage of non-contractile tissue increased in diaphragms from all transgenic mice. In conclusion, our results do not support the hypothesis that overexpression of independent antioxidant enzymes protects diaphragm muscle from contraction-induced fatigue or improves recovery from fatigue. Moreover, our data are consistent with the concept that a basal level of ROS is important to optimize muscle force production, since transgenic overexpression of major cellular antioxidants is associated with

  12. Transgenic overexpression of BAFF regulates the expression of ...

    Indian Academy of Sciences (India)

    LI ZHANG

    Hubei 430030, People's Republic of China. 3Second ... body homogenate of zebrafish and demonstrated a significant increase in BAFF-transgenic group. Therefore, our ... diet and brine shrimp according to the conditions in our sys- tem (Li et al. 2014). ... images were captured using a Leica DM3000B microscope. Embryos ...

  13. E2F-1-Induced p53-independent apoptosis in transgenic mice

    DEFF Research Database (Denmark)

    Holmberg, Christian Henrik; Helin, K.; Sehested, M.

    1998-01-01

    The E2F transcription factors are key targets for the retinoblastoma protein, pRB. By inactivation of E2Fs, pRB prevents progression to the S phase. To test proliferative functions of E2F, we generated transgenic mice expressing human E2F-1 and/or human DP-1. When the hydroxymethyl glutaryl...... involving increased apoptosis in the germinal epithelium. This effect was potentiated by simultaneous overexpression of DP-1. Testicular atrophy as a result of overexpression of E2F-1 and DP-1 is independent of functional p53, since p53-nullizygous transgenic mice overexpressing E2F-1 and DP-1 also suffered...

  14. Differential gene expression in ADAM10 and mutant ADAM10 transgenic mice

    Directory of Open Access Journals (Sweden)

    Postina Rolf

    2009-02-01

    Full Text Available Abstract Background In a transgenic mouse model of Alzheimer disease (AD, cleavage of the amyloid precursor protein (APP by the α-secretase ADAM10 prevented amyloid plaque formation, and alleviated cognitive deficits. Furthermore, ADAM10 overexpression increased the cortical synaptogenesis. These results suggest that upregulation of ADAM10 in the brain has beneficial effects on AD pathology. Results To assess the influence of ADAM10 on the gene expression profile in the brain, we performed a microarray analysis using RNA isolated from brains of five months old mice overexpressing either the α-secretase ADAM10, or a dominant-negative mutant (dn of this enzyme. As compared to non-transgenic wild-type mice, in ADAM10 transgenic mice 355 genes, and in dnADAM10 mice 143 genes were found to be differentially expressed. A higher number of genes was differentially regulated in double-transgenic mouse strains additionally expressing the human APP[V717I] mutant. Overexpression of proteolytically active ADAM10 affected several physiological pathways, such as cell communication, nervous system development, neuron projection as well as synaptic transmission. Although ADAM10 has been implicated in Notch and β-catenin signaling, no significant changes in the respective target genes were observed in adult ADAM10 transgenic mice. Real-time RT-PCR confirmed a downregulation of genes coding for the inflammation-associated proteins S100a8 and S100a9 induced by moderate ADAM10 overexpression. Overexpression of the dominant-negative form dnADAM10 led to a significant increase in the expression of the fatty acid-binding protein Fabp7, which also has been found in higher amounts in brains of Down syndrome patients. Conclusion In general, there was only a moderate alteration of gene expression in ADAM10 overexpressing mice. Genes coding for pro-inflammatory or pro-apoptotic proteins were not over-represented among differentially regulated genes. Even a decrease of

  15. Smad6/Smurf1 overexpression in cartilage delays chondrocyte hypertrophy and causes dwarfism with osteopenia

    Science.gov (United States)

    Horiki, Mitsuru; Imamura, Takeshi; Okamoto, Mina; Hayashi, Makoto; Murai, Junko; Myoui, Akira; Ochi, Takahiro; Miyazono, Kohei; Yoshikawa, Hideki; Tsumaki, Noriyuki

    2004-01-01

    Biochemical experiments have shown that Smad6 and Smad ubiquitin regulatory factor 1 (Smurf1) block the signal transduction of bone morphogenetic proteins (BMPs). However, their in vivo functions are largely unknown. Here, we generated transgenic mice overexpressing Smad6 in chondrocytes. Smad6 transgenic mice showed postnatal dwarfism with osteopenia and inhibition of Smad1/5/8 phosphorylation in chondrocytes. Endochondral ossification during development in these mice was associated with almost normal chondrocyte proliferation, significantly delayed chondrocyte hypertrophy, and thin trabecular bone. The reduced population of hypertrophic chondrocytes after birth seemed to be related to impaired bone growth and formation. Organ culture of cartilage rudiments showed that chondrocyte hypertrophy induced by BMP2 was inhibited in cartilage prepared from Smad6 transgenic mice. We then generated transgenic mice overexpressing Smurf1 in chondrocytes. Abnormalities were undetectable in Smurf1 transgenic mice. Mating Smad6 and Smurf1 transgenic mice produced double-transgenic pups with more delayed endochondral ossification than Smad6 transgenic mice. These results provided evidence that Smurf1 supports Smad6 function in vivo. PMID:15123739

  16. Composite potato plants with transgenic roots on non-transgenic shoots: a model system for studying gene silencing in roots.

    Science.gov (United States)

    Horn, Patricia; Santala, Johanna; Nielsen, Steen Lykke; Hühns, Maja; Broer, Inge; Valkonen, Jari P T

    2014-12-01

    Composite potato plants offer an extremely fast, effective and reliable system for studies on gene functions in roots using antisense or inverted-repeat but not sense constructs for gene inactivation. Composite plants, with transgenic roots on a non-transgenic shoot, can be obtained by shoot explant transformation with Agrobacterium rhizogenes. The aim of this study was to generate composite potato plants (Solanum tuberosum) to be used as a model system in future studies on root-pathogen interactions and gene silencing in the roots. The proportion of transgenic roots among the roots induced was high (80-100%) in the four potato cultivars tested (Albatros, Desirée, Sabina and Saturna). No wild-type adventitious roots were formed at mock inoculation site. All strains of A. rhizogenes tested induced phenotypically normal roots which, however, showed a reduced response to cytokinin as compared with non-transgenic roots. Nevertheless, both types of roots were infected to a similar high rate with the zoospores of Spongospora subterranea, a soilborne potato pathogen. The transgenic roots of composite potato plants expressed significantly higher amounts of β-glucuronidase (GUS) than the roots of a GUS-transgenic potato line event. Silencing of the uidA transgene (GUS) was tested by inducing roots on the GUS-transgenic cv. Albatros event with strains of A. rhizogenes over-expressing either the uidA sense or antisense transcripts, or inverted-repeat or hairpin uidA RNA. The three last mentioned constructs caused 2.5-4.0 fold reduction in the uidA mRNA expression. In contrast, over-expression of uidA resulted in over 3-fold increase in the uidA mRNA and GUS expression, indicating that sense-mediated silencing (co-suppression) was not functional in roots. The results suggest that composite plants offer a useful experimental system for potato research, which has gained little previous attention.

  17. Morphogenetic roles of perlecan in the tooth enamel organ: an analysis of overexpression using transgenic mice.

    Science.gov (United States)

    Ida-Yonemochi, Hiroko; Satokata, Ichiro; Ohshima, Hayato; Sato, Toshiya; Yokoyama, Minesuke; Yamada, Yoshihiko; Saku, Takashi

    2011-09-01

    Perlecan, a heparan sulfate proteoglycan, is enriched in the intercellular space of the enamel organ. To understand the role of perlecan in tooth morphogenesis, we used a keratin 5 promoter to generate transgenic (Tg) mice that over-express perlecan in epithelial cells, and examined their tooth germs at tissue and cellular levels. Immunohistochemistry showed that perlecan was more strongly expressed in the enamel organ cells of Tg mice than in wild-type mice. Histopathology showed wider intercellular spaces in the stellate reticulum of the Tg molars and loss of cellular polarity in the enamel organ, especially in its cervical region. Hertwig's epithelial root sheath (HERS) cells in Tg mice were irregularly aligned due to excessive deposits of perlecan along the inner, as well as on the outer sides of the HERS. Tg molars had dull-ended crowns and outward-curved tooth roots and their enamel was poorly crystallized, resulting in pronounced attrition of molar cusp areas. In Tg mice, expression of integrin β1 mRNA was remarkably higher at E18, while expression of bFGF, TGF-β1, DSPP and Shh was more elevated at P1. The overexpression of perlecan in the enamel organ resulted in irregular morphology of teeth, suggesting that the expression of perlecan regulates growth factor signaling in a stage-dependent manner during each step of the interaction between ameloblast-lineage cells and mesenchymal cells. Copyright © 2011 International Society of Matrix Biology. All rights reserved.

  18. Constitutive overexpression of the TaNF-YB4 gene in transgenic wheat significantly improves grain yield.

    Science.gov (United States)

    Yadav, Dinesh; Shavrukov, Yuri; Bazanova, Natalia; Chirkova, Larissa; Borisjuk, Nikolai; Kovalchuk, Nataliya; Ismagul, Ainur; Parent, Boris; Langridge, Peter; Hrmova, Maria; Lopato, Sergiy

    2015-11-01

    Heterotrimeric nuclear factors Y (NF-Ys) are involved in regulation of various vital functions in all eukaryotic organisms. Although a number of NF-Y subunits have been characterized in model plants, only a few have been functionally evaluated in crops. In this work, a number of genes encoding NF-YB and NF-YC subunits were isolated from drought-tolerant wheat (Triticum aestivum L. cv. RAC875), and the impact of the overexpression of TaNF-YB4 in the Australian wheat cultivar Gladius was investigated. TaNF-YB4 was isolated as a result of two consecutive yeast two-hybrid (Y2H) screens, where ZmNF-YB2a was used as a starting bait. A new NF-YC subunit, designated TaNF-YC15, was isolated in the first Y2H screen and used as bait in a second screen, which identified two wheat NF-YB subunits, TaNF-YB2 and TaNF-YB4. Three-dimensional modelling of a TaNF-YB2/TaNF-YC15 dimer revealed structural determinants that may underlie interaction selectivity. The TaNF-YB4 gene was placed under the control of the strong constitutive polyubiquitin promoter from maize and introduced into wheat by biolistic bombardment. The growth and yield components of several independent transgenic lines with up-regulated levels of TaNF-YB4 were evaluated under well-watered conditions (T1-T3 generations) and under mild drought (T2 generation). Analysis of T2 plants was performed in large deep containers in conditions close to field trials. Under optimal watering conditions, transgenic wheat plants produced significantly more spikes but other yield components did not change. This resulted in a 20-30% increased grain yield compared with untransformed control plants. Under water-limited conditions transgenic lines maintained parity in yield performance. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  19. Transgenic Overexpression of the Proprotein Convertase Furin Enhances Skin Tumor Growth

    Directory of Open Access Journals (Sweden)

    Jian Fu

    2012-04-01

    Full Text Available Furin, one of the members of the family of proprotein convertases (PCs, ubiquitously expressed as a type I membrane-bound proteinase, activates several proteins that contribute to tumor progression. In vitro studies using cancer cell lines and clinical specimens demonstrated that furin processes important substrates such as insulin-like growth factor 1 receptor (IGF-1R and transforming growth factor β, leading to increased tumor growth and progression. Despite the numerous studies associating furin with tumor development, its effects in preclinical models has not been comprehensively studied. In this study, we sought to determine the protumorigenic role of furin in vivo after a two-stage chemical carcinogenesis protocol in transgenic mice in which furin expression was targeted to the epidermal basal layer. We found that processing of the PC substrate IGF-1R and the proliferation rate of mouse epidermis was enhanced in transgenic mice when compared with their WT counterparts. Histopathologic diagnoses of the tumors demonstrated that furin transgenic mice (line F47 developed twice as many squamous carcinomas as the control, WT mice (P < .002. Similarly, tumors cells from transgenic mice were able to process PC substrates more efficiently than tumor cells from WT mice. Furthermore, furin expression resulted in a higher SCC volume in transgenic mice as well as an increase in the percentage of high-grade SCC, including poorly differentiated and spindle cell carcinomas. In conclusion, expression of furin in the basal layer of the epidermis increased tumor development and enhanced tumor growth, supporting the consideration of furin as a potential target for cancer treatment.

  20. Glutathione Transferase from Trichoderma virens Enhances Cadmium Tolerance without Enhancing Its Accumulation in Transgenic Nicotiana tabacum

    Science.gov (United States)

    Dixit, Prachy; Mukherjee, Prasun K.; Ramachandran, V.; Eapen, Susan

    2011-01-01

    Background Cadmium (Cd) is a major heavy metal pollutant which is highly toxic to plants and animals. Vast agricultural areas worldwide are contaminated with Cd. Plants take up Cd and through the food chain it reaches humans and causes toxicity. It is ideal to develop plants tolerant to Cd, without enhanced accumulation in the edible parts for human consumption. Glutathione transferases (GST) are a family of multifunctional enzymes known to have important roles in combating oxidative stresses induced by various heavy metals including Cd. Some GSTs are also known to function as glutathione peroxidases. Overexpression/heterologous expression of GSTs is expected to result in plants tolerant to heavy metals such as Cd. Results Here, we report cloning of a glutathione transferase gene from Trichoderma virens, a biocontrol fungus and introducing it into Nicotiana tabacum plants by Agrobacterium-mediated gene transfer. Transgenic nature of the plants was confirmed by Southern blot hybridization and expression by reverse transcription PCR. Transgene (TvGST) showed single gene Mendelian inheritance. When transgenic plants expressing TvGST gene were exposed to different concentrations of Cd, they were found to be more tolerant compared to wild type plants, with transgenic plants showing lower levels of lipid peroxidation. Levels of different antioxidant enzymes such as glutathione transferase, superoxide dismutase, ascorbate peroxidase, guiacol peroxidase and catalase showed enhanced levels in transgenic plants expressing TvGST compared to control plants, when exposed to Cd. Cadmium accumulation in the plant biomass in transgenic plants were similar or lower than wild-type plants. Conclusion The results of the present study suggest that transgenic tobacco plants expressing a Trichoderma virens GST are more tolerant to Cd, without enhancing its accumulation in the plant biomass. It should be possible to extend the present results to crop plants for developing Cd tolerance and

  1. Glutathione transferase from Trichoderma virens enhances cadmium tolerance without enhancing its accumulation in transgenic Nicotiana tabacum.

    Directory of Open Access Journals (Sweden)

    Prachy Dixit

    Full Text Available BACKGROUND: Cadmium (Cd is a major heavy metal pollutant which is highly toxic to plants and animals. Vast agricultural areas worldwide are contaminated with Cd. Plants take up Cd and through the food chain it reaches humans and causes toxicity. It is ideal to develop plants tolerant to Cd, without enhanced accumulation in the edible parts for human consumption. Glutathione transferases (GST are a family of multifunctional enzymes known to have important roles in combating oxidative stresses induced by various heavy metals including Cd. Some GSTs are also known to function as glutathione peroxidases. Overexpression/heterologous expression of GSTs is expected to result in plants tolerant to heavy metals such as Cd. RESULTS: Here, we report cloning of a glutathione transferase gene from Trichoderma virens, a biocontrol fungus and introducing it into Nicotiana tabacum plants by Agrobacterium-mediated gene transfer. Transgenic nature of the plants was confirmed by Southern blot hybridization and expression by reverse transcription PCR. Transgene (TvGST showed single gene Mendelian inheritance. When transgenic plants expressing TvGST gene were exposed to different concentrations of Cd, they were found to be more tolerant compared to wild type plants, with transgenic plants showing lower levels of lipid peroxidation. Levels of different antioxidant enzymes such as glutathione transferase, superoxide dismutase, ascorbate peroxidase, guiacol peroxidase and catalase showed enhanced levels in transgenic plants expressing TvGST compared to control plants, when exposed to Cd. Cadmium accumulation in the plant biomass in transgenic plants were similar or lower than wild-type plants. CONCLUSION: The results of the present study suggest that transgenic tobacco plants expressing a Trichoderma virens GST are more tolerant to Cd, without enhancing its accumulation in the plant biomass. It should be possible to extend the present results to crop plants for

  2. Cre/lox system to develop selectable marker free transgenic tobacco plants conferring resistance against sap sucking homopteran insect.

    Science.gov (United States)

    Chakraborti, Dipankar; Sarkar, Anindya; Mondal, Hossain A; Schuermann, David; Hohn, Barbara; Sarmah, Bidyut K; Das, Sampa

    2008-10-01

    A binary expression vector was constructed containing the insecticidal gene Allium sativum leaf agglutinin (ASAL), and a selectable nptII marker gene cassette, flanked by lox sites. Similarly, another binary vector was developed with the chimeric cre gene construct. Transformed tobacco plants were generated with these two independent vectors. Each of the T(0) lox plants was crossed with T(0) Cre plants. PCR analyses followed by the sequencing of the target T-DNA part of the hybrid T(1) plants demonstrated the excision of the nptII gene in highly precised manner in certain percentage of the T(1) hybrid lines. The frequency of such marker gene excision was calculated to be 19.2% in the hybrids. Marker free plants were able to express ASAL efficiently and reduce the survivability of Myzus persiceae, the deadly pest of tobacco significantly, compared to the control tobacco plants. Results of PCR and Southern blot analyses of some of the T(2) plants detected the absence of cre as well as nptII genes. Thus, the crossing strategy involving Cre/lox system for the excision of marker genes appears to be very effective and easy to execute. Documentation of such marker excision phenomenon in the transgenic plants expressing the important insecticidal protein for the first time has a great significance from agricultural and biotechnological points of view.

  3. Atypical scrapie prions from sheep and lack of disease in transgenic mice overexpressing human prion protein.

    Science.gov (United States)

    Wadsworth, Jonathan D F; Joiner, Susan; Linehan, Jacqueline M; Balkema-Buschmann, Anne; Spiropoulos, John; Simmons, Marion M; Griffiths, Peter C; Groschup, Martin H; Hope, James; Brandner, Sebastian; Asante, Emmanuel A; Collinge, John

    2013-11-01

    Public and animal health controls to limit human exposure to animal prions are focused on bovine spongiform encephalopathy (BSE), but other prion strains in ruminants may also have zoonotic potential. One example is atypical/Nor98 scrapie, which evaded statutory diagnostic methods worldwide until the early 2000s. To investigate whether sheep infected with scrapie prions could be another source of infection, we inoculated transgenic mice that overexpressed human prion protein with brain tissue from sheep with natural field cases of classical and atypical scrapie, sheep with experimental BSE, and cattle with BSE. We found that these mice were susceptible to BSE prions, but disease did not develop after prolonged postinoculation periods when mice were inoculated with classical or atypical scrapie prions. These data are consistent with the conclusion that prion disease is less likely to develop in humans after exposure to naturally occurring prions of sheep than after exposure to epizootic BSE prions of ruminants.

  4. Overexpression of plastidial thioredoxins f and m differentially alters photosynthetic activity and response to oxidative stress in tobacco plants

    Directory of Open Access Journals (Sweden)

    Pascal eREY

    2013-10-01

    Full Text Available Plants display a remarkable diversity of thioredoxins (Trxs, reductases controlling the thiol redox status of proteins. The physiological function of many of them remains elusive, particularly for plastidial Trxs f and m, which are presumed based on biochemical data to regulate photosynthetic reactions and carbon metabolism. Recent reports revealed that Trxs f and m participate in vivo in the control of starch metabolism and cyclic photosynthetic electron transfer around photosystem I, respectively. To further delineate their in planta function, we compared the photosynthetic characteristics, the level and/or activity of various Trx targets and the responses to oxidative stress in transplastomic tobacco plants overexpressing either Trx f or Trx m. We found that plants overexpressing Trx m specifically exhibit altered growth, reduced chlorophyll content, impaired photosynthetic linear electron transfer and decreased pools of glutathione and ascorbate. In both transplastomic lines, activities of two enzymes involved in carbon metabolism, NADP-malate dehydrogenase and NADP-glyceraldehyde-3-phosphate dehydrogenase are markedly and similarly altered. In contrast, plants overexpressing Trx m specifically display increased capacity for methionine sulfoxide reductases, enzymes repairing damaged proteins by regenerating methionine from oxidized methionine. Finally, we also observed that transplastomic plants exhibit distinct responses when exposed to oxidative stress conditions generated by methyl viologen or exposure to high light combined with low temperature, the plants overexpressing Trx m being notably more tolerant than Wt and those overexpressing Trx f. Altogether, these data indicate that Trxs f and m fulfill distinct physiological functions. They prompt us to propose that the m type is involved in key processes linking photosynthetic activity, redox homeostasis and antioxidant mechanisms in the chloroplast.

  5. The Metallothionein Gene, TaMT3, from Tamarix androssowii Confers Cd2+ Tolerance in Tobacco

    Directory of Open Access Journals (Sweden)

    Boru Zhou

    2014-06-01

    Full Text Available Cadmium (Cd is a nonessential microelement and low concentration Cd2+ has strong toxicity to plant growth. Plant metallothioneins, a class of low molecular, cystein(Cys-rich and heavy-metal binding proteins, play an important role in both metal chaperoning and scavenging of reactive oxygen species (ROS with their large number of cysteine residues and therefore, protect plants from oxidative damage. In this study, a metallothionein gene, TaMT3, isolated from Tamarix androssowii was transformed into tobacco (Nicotiana tobacum through Agrobacterium-mediated leaf disc method, and correctly expressed under the control of 35S promoter. Under Cd2+ stress, the transgenic tobacco showed significant increases of superoxide dismutase (SOD activity and chlorophyll concentration, but decreases of peroxidase (POD activity and malondialdehyde (MDA accumulation when compared to the non-transgenic tobacco. Vigorous growth of transgenic tobacco was observed at the early development stages, resulting in plant height and fresh weight were significantly larger than those of the non-transgenic tobacco under Cd2+ stress. These results demonstrated that the expression of the exogenous TaMT3 gene increased the ability of ROS cleaning-up, indicating a stronger tolerance to Cd2+ stress.

  6. The metallothionein gene, TaMT3, from Tamarix androssowii confers Cd2+ tolerance in tobacco.

    Science.gov (United States)

    Zhou, Boru; Yao, Wenjing; Wang, Shengji; Wang, Xinwang; Jiang, Tingbo

    2014-06-10

    Cadmium (Cd) is a nonessential microelement and low concentration Cd2+ has strong toxicity to plant growth. Plant metallothioneins, a class of low molecular, cystein(Cys)-rich and heavy-metal binding proteins, play an important role in both metal chaperoning and scavenging of reactive oxygen species (ROS) with their large number of cysteine residues and therefore, protect plants from oxidative damage. In this study, a metallothionein gene, TaMT3, isolated from Tamarix androssowii was transformed into tobacco (Nicotiana tobacum) through Agrobacterium-mediated leaf disc method, and correctly expressed under the control of 35S promoter. Under Cd2+ stress, the transgenic tobacco showed significant increases of superoxide dismutase (SOD) activity and chlorophyll concentration, but decreases of peroxidase (POD) activity and malondialdehyde (MDA) accumulation when compared to the non-transgenic tobacco. Vigorous growth of transgenic tobacco was observed at the early development stages, resulting in plant height and fresh weight were significantly larger than those of the non-transgenic tobacco under Cd2+ stress. These results demonstrated that the expression of the exogenous TaMT3 gene increased the ability of ROS cleaning-up, indicating a stronger tolerance to Cd2+ stress.

  7. Overexpression of CrtR-b2 (carotene beta hydroxylase 2) from S. lycopersicum L. differentially affects xanthophyll synthesis and accumulation in transgenic tomato plants.

    Science.gov (United States)

    D'Ambrosio, Caterina; Stigliani, Adriana Lucia; Giorio, Giovanni

    2011-02-01

    Plant chloroplasts are enriched in xanthophylls which participate in photosynthesis as light-absorbing pigments and as dissipaters of excess light. In comparison, chromoplasts have evolved the capacity to synthesize and store brightly coloured carotenoid pigments to give flowers and fruits the power to attract pollinators and fruit dispersers. The best performing accumulator of xanthophylls in tomato is the petal chromoplast in contrast to the fruit chromoplast which only seems able to store carotenes. We have generated genetically engineered tomato lines carrying the tomato CrtR-b2 transgene with the aim of forcing the fruit to accumulate beta-xanthophylls. Both chloroplast- and chromoplast-containing tissues of hemizygous transgenic plants were found to contain elevated xanthophyll contents as a direct consequence of the increased number of CrtR-b2 transcripts. Hemizygous transgenic leaves contained fourfold more violaxanthin than control leaves. Developing fruits were yellow instead of green since they lacked chlorophyll a, and their violaxanthin and neoxanthin contents were seven- and threefold higher, respectively, than those of the control. Ripe fruits of hemizygous transgenic plants contained free violaxanthin and significant amounts of esterified xanthophylls. Esterified xanthophylls were present also in ripe fruits of control and homozygous plants. However, in transgenic homozygous plants, we observed a reduction in transcript content in most tissues, particularly in petals, due to a post-transcriptional gene silencing process. These findings demonstrate that tomato fruit chromoplasts can accumulate xanthophylls with the same sequestration mechanism (esterification) as that exploited by chromoplasts of the tomato petal and pepper fruit. This study on transgenic plants overexpressing an important carotenoid gene (CrtR-b2) provides an interesting model for future investigations on perturbations in beta-carotene-derived xanthophyll synthesis which in turn may

  8. Over-expressing Salicornia europaea (SeNHX1) gene in tobacco ...

    African Journals Online (AJOL)

    An increase of salt tolerance in transgenics was observed both in vitro and in pot culture. When exposed to either 138 mM or 172 mM NaCl in vitro, the transgenic apical meristem and stem segments with buds exhibited stronger salt-tolerance than their wild-type counterparts. In pot soil with 10.2 mg g-1 DW Na+ stress, ...

  9. Hippocampal changes produced by overexpression of the human CHRNA5/A3/B4 gene cluster may underlie cognitive deficits rescued by nicotine in transgenic mice.

    Science.gov (United States)

    Molas, Susanna; Gener, Thomas; Güell, Jofre; Martín, Mairena; Ballesteros-Yáñez, Inmaculada; Sanchez-Vives, Maria V; Dierssen, Mara

    2014-11-11

    Addiction involves long-lasting maladaptive changes including development of disruptive drug-stimuli associations. Nicotine-induced neuroplasticity underlies the development of tobacco addiction but also, in regions such as the hippocampus, the ability of this drug to enhance cognitive capabilities. Here, we propose that the genetic locus of susceptibility to nicotine addiction, the CHRNA5/A3/B4 gene cluster, encoding the α5, α3 and β4 subunits of the nicotinic acetylcholine receptors (nAChRs), may influence nicotine-induced neuroadaptations. We have used transgenic mice overexpressing the human cluster (TgCHRNA5/A3/B4) to investigate hippocampal structure and function in genetically susceptible individuals. TgCHRNA5/A3/B4 mice presented a marked reduction in the dendrite complexity of CA1 hippocampal pyramidal neurons along with an increased dendritic spine density. In addition, TgCHRNA5/A3/B4 exhibited increased VGLUT1/VGAT ratio in the CA1 region, suggesting an excitatory/inhibitory imbalance. These hippocampal alterations were accompanied by a significant impairment in short-term novelty recognition memory. Interestingly, chronic infusion of nicotine (3.25 mg/kg/d for 7 d) was able to rescue the reduced dendritic complexity, the excitatory/inhibitory imbalance and the cognitive impairment in TgCHRNA5/A3/B4. Our results suggest that chronic nicotine treatment may represent a compensatory strategy in individuals with altered expression of the CHRNA5/A3/B4 region.

  10. Transgenic plants expressing the AaIT/GNA fusion protein show increased resistance and toxicity to both chewing and sucking pests.

    Science.gov (United States)

    Liu, Shu-Min; Li, Jie; Zhu, Jin-Qi; Wang, Xiao-Wei; Wang, Cheng-Shu; Liu, Shu-Sheng; Chen, Xue-Xin; Li, Sheng

    2016-04-01

    The adoption of pest-resistant transgenic plants to reduce yield losses and decrease pesticide use has been successful. To achieve the goal of controlling both chewing and sucking pests in a given transgenic plant, we generated transgenic tobacco, Arabidopsis, and rice plants expressing the fusion protein, AaIT/GNA, in which an insecticidal scorpion venom neurotoxin (Androctonus australis toxin, AaIT) is fused to snowdrop lectin (Galanthus nivalis agglutinin, GNA). Compared with transgenic tobacco and Arabidopsis plants expressing AaIT or GNA, transgenic plants expressing AaIT/GNA exhibited increased resistance and toxicity to one chewing pest, the cotton bollworm, Helicoverpa armigera. Transgenic tobacco and rice plants expressing AaIT/GNA showed increased resistance and toxicity to two sucking pests, the whitefly, Bemisia tabaci, and the rice brown planthopper, Nilaparvata lugens, respectively. Moreover, in the field, transgenic rice plants expressing AaIT/GNA exhibited a significant improvement in grain yield when infested with N. lugens. This study shows that expressing the AaIT/GNA fusion protein in transgenic plants can be a useful approach for controlling pests, particularly sucking pests which are not susceptible to the toxin in Bt crops. © 2015 Institute of Zoology, Chinese Academy of Sciences.

  11. Intestine-specific overexpression of IL-10 improves survival in polymicrobial sepsis.

    Science.gov (United States)

    Rajan, Saju; Vyas, Dinesh; Clark, Andrew T; Woolsey, Cheryl A; Clark, Jessica A; Hotchkiss, Richard S; Buchman, Timothy G; Coopersmith, Craig M

    2008-04-01

    Targeted IL-10 therapy improves survival in preclinical models of critical illness, and intestine-specific IL-10 decreases inflammation in models of chronic Inflammatory disease. We therefore sought to determine whether intestine-specific overexpression of IL-10 would improve survival in sepsis. Transgenic mice that overexpress IL-10 in their gut epithelium (Fabpi-IL-10 mice) and wild-type (WT) littermates (n = 127) were subjected to cecal ligation and puncture with a 27-gauge needle. The 7-day survival rate was 45% in transgenic animals and 30% in WT animals (P < or = 0.05). Systemic levels of IL-10 were undetectable in both groups of animals under basal conditions and were elevated to a similar degree in septic animals regardless of whether they expressed the transgene. Local parameter of injury, including gut epithelial apoptosis, intestinal permeability, peritoneal lavage cytokines, and stimulated cytokines from intraepithelial lymphocytes, were similar between transgenic and WT mice. However, in stimulated splenocytes, proinflammatory cytokines monocyte chemoattractant protein 1 (189 +/- 43 vs. 40 +/- 8 pg/mL) and IL-6 (116 +/- 28 vs. 34 +/- 9 pg/mL) were lower in Fabpi-IL-10 mice than WT littermates despite the intestine-specific nature of the transgene (P < 0.05). Cytokine levels were similar in blood and bronchoalveolar lavage fluid between the 2 groups, as were circulating LPS levels. Transgenic mice also had lower white blood cell counts associated with lower absolute neutrophil counts (0.5 +/- 0.1 vs. 1.0 +/- 0.2 10(3)/mm3; P < 0.05). These results indicate that gut-specific overexpression of IL-10 improves survival in a murine model of sepsis, and interactions between the intestinal epithelium and the systemic immune system may play a role in conferring this survival advantage.

  12. Establishment and evaluation of a transgenic mouse model of arthritis induced by overexpressing human tumor necrosis factor alpha

    Directory of Open Access Journals (Sweden)

    Ge Li

    2016-04-01

    Full Text Available Tumor necrosis factor alpha (TNFα plays a key role in the pathogenesis of rheumatoid arthritis (RA. Blockade of TNFα by monoclonal antibody has been widely used for the therapy of RA since the 1990s; however, its mechanism of efficacy, and potential safety concerns of the treatment are still not fully understood. This study sought to establish a transgenic arthritic mouse model by overexpressing human TNFα (hTNFα and to apply this model as a means to evaluate therapeutic consequences of TNFα inhibitors. The transgenic mouse line (TgTC with FVB background was generated by incorporating 3′-modified hTNFα gene sequences. A progressively erosive polyarthritis developed in the TgTC mice, with many characteristics observed in human rheumatoid arthritis, including polyarticular swelling, impairment of movement, synovial hyperplasia, and cartilage and bone erosion. Gene expression analysis demonstrated that hTNFα is not only expressed in hyperplastic synovial membrane, but also in tissues without lesions, including brain, lung and kidney. Treatment of the TgTC mice with anti-hTNFα monoclonal antibodies (mAb significantly decreased the level of hTNFα in the diseased joint and effectively prevented development of arthritis in a dose-dependent response fashion. Our results indicated that the TgTC mice represent a genetic model which can be used to comprehensively investigate the pathogenesis and therapeutics of TNFα-related diseases.

  13. Targeted Transgenic Overexpression of Mitochondrial Thymidine Kinase (TK2) Alters Mitochondrial DNA (mtDNA) and Mitochondrial Polypeptide Abundance

    Science.gov (United States)

    Hosseini, Seyed H.; Kohler, James J.; Haase, Chad P.; Tioleco, Nina; Stuart, Tami; Keebaugh, Erin; Ludaway, Tomika; Russ, Rodney; Green, Elgin; Long, Robert; Wang, Liya; Eriksson, Staffan; Lewis, William

    2007-01-01

    Mitochondrial toxicity limits nucleoside reverse transcriptase inhibitors (NRTIs) for acquired immune deficiency syndrome. NRTI triphosphates, the active moieties, inhibit human immunodeficiency virus reverse transcriptase and eukaryotic mitochondrial DNA polymerase pol-γ. NRTI phosphorylation seems to correlate with mitochondrial toxicity, but experimental evidence is lacking. Transgenic mice (TGs) with cardiac overexpression of thymidine kinase isoforms (mitochondrial TK2 and cytoplasmic TK1) were used to study NRTI mitochondrial toxicity. Echocardiography and nuclear magnetic resonance imaging defined cardiac performance and structure. TK gene copy and enzyme activity, mitochondrial (mt) DNA and polypeptide abundance, succinate dehydrogenase and cytochrome oxidase histochemistry, and electron microscopy correlated with transgenesis, mitochondrial structure, and biogenesis. Antiretroviral combinations simulated therapy. Untreated hTK1 or TK2 TGs exhibited normal left ventricle mass. In TK2 TGs, cardiac TK2 gene copy doubled, activity increased 300-fold, and mtDNA abundance doubled. Abundance of the 17-kd subunit of complex I, succinate dehydrogenase histochemical activity, and cristae density increased. NRTIs increased left ventricle mass 20% in TK2 TGs. TK activity increased 3 logs in hTK1 TGs, but no cardiac phenotype resulted. NRTIs abrogated functional effects of transgenically increased TK2 activity but had no effect on TK2 mtDNA abundance. Thus, NRTI mitochondrial phosphorylation by TK2 is integral to clinical NRTI mitochondrial toxicity. PMID:17322372

  14. Cloning and functional characterization of MusaVND1 using transgenic banana plants.

    Science.gov (United States)

    Negi, Sanjana; Tak, Himanshu; Ganapathi, T R

    2015-06-01

    Vascular related NAC (NAM, ATAF and CUC) domain-containing genes regulate secondary wall deposition and differentiation of xylem vessel elements. MusaVND1 is an ortholog of Arabidopsis VND1 and contains the highly conserved NAC domain. The expression of MusaVND1 is highest in developing corm and during lignification conditions, the increase in expression of MusaVND1 coincides with the expression of PAL, COMT and C4H genes. MusaVND1 encodes a nuclear localized protein as MusaVND1-GFP fusion protein gets localized to nucleus. Transient overexpression of MusaVND1 converts banana embryogenic cells to xylem vessel elements, with a final differentiation frequency of 33.54% at the end of tenth day. Transgenic banana plants overexpressing MusaVND1 showed stunted growth and were characterized by PCR and Southern blot analysis. Transgenic banana plants showed transdifferentiation of various types of cells into xylem vessel elements and ectopic deposition of lignin in cells of various plant organs such as leaf and corm. Tracheary element formation was seen in the cortical region of transgenic corm as well as in epidermal cells of leaves. Biochemical analysis indicates significantly higher levels of lignin and cellulose content in transgenic banana lines than control plants. MusaVND1 overexpressing transgenic banana plants showed elevated expression levels of genes involved in lignin and cellulose biosynthesis pathway. Further expression of different MYB transcription factors positively regulating secondary wall deposition was also up regulated in MusaVND1 transgenic lines.

  15. Upregulation of triglyceride synthesis in skeletal muscle overexpressing DGAT1

    OpenAIRE

    Yang, Feifei; Wei, Zhuying; Ding, Xiangbin; Liu, Xinfeng; Ge, Xiuguo; Song, Guimin; Li, Guangpeng; Guo, Hong

    2013-01-01

    The gene encoding diacylglycerol acyltransferase (DGAT1) is a functional and positional candidate gene for milk and intramuscular fat content. A bovine DGAT1 overexpression vector was constructed containing mouse MCK promoter and bovine DGAT1 cDNA. MCK-DGAT1 transgene in FVB mice was researched in present study. The transgene DGAT1 had a high level of expression in contrast to the endogenous DGAT1 in posterior tibial muscle of the transgenic mice, but a low expression level in the cardiac mus...

  16. Hyperlipidemia and cutaneous abnormalities in transgenic mice overexpressing human apolipoprotein C1

    NARCIS (Netherlands)

    Jong, M. C.; Gijbels, M. J.; Dahlmans, V. E.; Gorp, P. J.; Koopman, S. J.; Ponec, M.; Hofker, M. H.; Havekes, L. M.

    1998-01-01

    Transgenic mice were generated with different levels of human apolipoprotein C1 (APOC1) expression in liver and skin. At 2 mo of age, serum levels of cholesterol, triglycerides (TG), and FFA were strongly elevated in APOC1 transgenic mice compared with wild-type mice. These elevated levels of serum

  17. Assays of dioxins and dioxin-like compounds in actually contaminated soils using transgenic tobacco plants carrying a recombinant mouse aryl hydrocarbon receptor-mediated β-glucuronidase reporter gene expression system.

    Science.gov (United States)

    Inui, Hideyuki; Gion, Keiko; Utani, Yasushi; Wakai, Taketo; Kodama, Susumu; Eun, Heesoo; Kim, Yun-Seok; Ohkawa, Hideo

    2012-01-01

    The transgenic tobacco plant XD4V-26 carrying the recombinant mouse aryl hydrocarbon receptor XD4V-mediated β-glucuronidase (GUS) reporter gene expression system was used for assay of dioxins and dioxin-like compounds consisting of polychlorinated dibenzeno-p-dioxins, polychlorinated dibenzofurans, and coplanar polychlorinated biphenyls (Co-PCBs) in actually contaminated soils. The transgenic tobacco plant XD4V-26 showed a significant dose-dependent induced GUS activity when cultured on MS medium containing PCB126 [toxic equivalency factor (TEF) = 0.1]. In contrast, PCB169 and PCB180, which have 0.03 of TEF and unassigned TEF values, respectively, did not significantly induce GUS activity under the same conditions as with PCB126. When the tobacco plants were cultivated for up to 5 weeks on actually contaminated soils with dioxins and dioxin-like compounds collected from the periphery of an incinerator used for disposal of residential and industrial wastes, GUS activity in the leaves was dose-dependently increased. The plants clearly detected 360 pg-TEQ g(-1) of dioxins and dioxin-like compounds in this assay. There was a positive correlation between GUS activity and TEQ value of dioxins and dioxin-like compounds in the plants. This assay does not require any extraction and purification processes for the actually contaminated soil samples.

  18. Overexpression of the WOX gene STENOFOLIA improves biomass yield and sugar release in transgenic grasses and display altered cytokinin homeostasis.

    Directory of Open Access Journals (Sweden)

    Hui Wang

    2017-03-01

    Full Text Available Lignocellulosic biomass can be a significant source of renewable clean energy with continued improvement in biomass yield and bioconversion strategies. In higher plants, the leaf blade is the central energy convertor where solar energy and CO2 are assimilated to make the building blocks for biomass production. Here we report that introducing the leaf blade development regulator STENOFOLIA (STF, a WOX family transcription factor, into the biofuel crop switchgrass, significantly improves both biomass yield and sugar release. We found that STF overexpressing switchgrass plants produced approximately 2-fold more dry biomass and release approximately 1.8-fold more solubilized sugars without pretreatment compared to controls. The biomass increase was attributed mainly to increased leaf width and stem thickness, which was also consistent in STF transgenic rice and Brachypodium, and appeared to be caused by enhanced cell proliferation. STF directly binds to multiple regions in the promoters of some cytokinin oxidase/dehydrogenase (CKX genes and represses their expression in all three transgenic grasses. This repression was accompanied by a significant increase in active cytokinin content in transgenic rice leaves, suggesting that the increase in biomass productivity and sugar release could at least in part be associated with improved cytokinin levels caused by repression of cytokinin degrading enzymes. Our study provides a new tool for improving biomass feedstock yield in bioenergy crops, and uncovers a novel mechanistic insight in the function of STF, which may also apply to other repressive WOX genes that are master regulators of several key plant developmental programs.

  19. Insect-resistance and high-yield transgenic tobacco obtained by ...

    African Journals Online (AJOL)

    The modified synthesized VHb gene and insectidal gene (GFMcryIA) were transferred to tobacco plants by Agrobacterium-mediated transformation. The bivalent genes were inserted successfully into the tobacco genome and detected by PCR amplification. Southern blot and Western blot analyses showed that VHb gene ...

  20. Overexpression of CCS in G93A-SOD1 mice leads to accelerated neurological deficits with severe mitochondrial pathology.

    Science.gov (United States)

    Son, Marjatta; Puttaparthi, Krishna; Kawamata, Hibiki; Rajendran, Bhagya; Boyer, Philip J; Manfredi, Giovanni; Elliott, Jeffrey L

    2007-04-03

    Cu, Zn superoxide dismutase (SOD1) has been detected within spinal cord mitochondria of mutant SOD1 transgenic mice, a model of familial ALS. The copper chaperone for SOD1 (CCS) provides SOD1 with copper, facilitates the conversion of immature apo-SOD1 to a mature holoform, and influences in yeast the cytosolic/mitochondrial partitioning of SOD1. To determine how CCS affects G93A-SOD1-induced disease, we generated transgenic mice overexpressing CCS and crossed them to G93A-SOD1 or wild-type SOD1 transgenic mice. Both CCS transgenic mice and CCS/wild-type-SOD1 dual transgenic mice are neurologically normal. In contrast, CCS/G93A-SOD1 dual transgenic mice develop accelerated neurological deficits, with a mean survival of 36 days, compared with 242 days for G93A-SOD1 mice. Immuno-EM and subcellular fractionation studies on the spinal cord show that G93A-SOD1 is enriched within mitochondria in the presence of CCS overexpression. Our results indicate that CCS overexpression in G93A-SOD1 mice produces severe mitochondrial pathology and accelerates disease course.

  1. Tobacco expressing pap1 increases the responses to par and uv-a by enhancing soluble sugars and flavonoids and elevating plant protections

    International Nuclear Information System (INIS)

    Sompornpailin, K.; Kanthang, S.

    2015-01-01

    Five lines of transgenic tobacco over-expressing Production of Anthocyanin Pigment 1 (PAP1) cDNA were analysis of metabolic response against the radiation and their protection of the plant under tissue culture condition. PAP1 transgenic and wild type (WT) plants were treated with the radiations of photosynthetically activate radiation (PAR) or PAR combined with UV-A. All lines of transgenic significantly increased in amounts of p-coumaric acid, naringenin apigenin more than WT under both treatments. Additional UV-A radiating to plant rose up kaempferol content in WT plant (1.5 times) and in PAP1 transgenics (1.8 times). These transgenic plants treated under both conditions had also increased anthocyanin substances (pelargonidin) with significant value after compared to WT. Content of total soluble sugar (TSS) was related to the content of total flavonoids in transgenic. PAR combined with UV-A had a lower induction of the electrolyte leakage percentage and malondialdehyde (MDA) level in the transgenic leaf tissue compared to WT tissue. The metabolic substance levels were considered on its protection of plant cells. In transgenic tissue, the enhancement of apigenin level strongly diminished the increase level of electrolyte leakage while the levels of TSS, p-coumaric acid and naringinin less affected. Moreover, the increase levels of kaempferol and pelargonidin associated with the decrease level of MDA, while the TSS level reversely responded. The PAP1 transgenic increased response of light by adaptation of their metabolites (TSS, p-coumaric acid and flavonoids) consequently enhance parameter indicating protections of the cell. (author)

  2. Constitutive expression of CaPLA1 conferred enhanced growth and grain yield in transgenic rice plants.

    Science.gov (United States)

    Park, Ki Youl; Kim, Eun Yu; Seo, Young Sam; Kim, Woo Taek

    2016-03-01

    Phospholipids are not only important components of cell membranes, but participate in diverse processes in higher plants. In this study, we generated Capsicum annuum phospholipiase A1 (CaPLA1) overexpressing transgenic rice (Oryza sativa L.) plants under the control of the maize ubiquitin promoter. The T4 CaPLA1-overexpressing rice plants (Ubi:CaPLA1) had a higher root:shoot mass ratio than the wild-type plants in the vegetative stage. Leaf epidermal cells from transgenic plants had more cells than wild-type plants. Genes that code for cyclin and lipid metabolic enzymes were up-regulated in the transgenic lines. When grown under typical paddy field conditions, the transgenic plants produced more tillers, longer panicles and more branches per panicle than the wild-type plants, all of which resulted in greater grain yield. Microarray analysis suggests that gene expressions that are related with cell proliferation, lipid metabolism, and redox state were widely altered in CaPLA1-overexpressing transgenic rice plants. Ubi:CaPLA1 plants had a reduced membrane peroxidation state, as determined by malondialdehyde and conjugated diene levels and higher peroxidase activity than wild-type rice plants. Furthermore, three isoprenoid synthetic genes encoding terpenoid synthase, hydroxysteroid dehydrogenase and 3-hydroxy-3-methyl-glutaryl-CoA reductase were up-regulated in CaPLA1-overexpressing plants. We suggest that constitutive expression of CaPLA1 conferred increased grain yield with enhanced growth in transgenic rice plants by alteration of gene activities related with cell proliferation, lipid metabolism, membrane peroxidation state and isoprenoid biosynthesis.

  3. Tobacco, alcohol, and p53 overexpression in early colorectal neoplasia

    International Nuclear Information System (INIS)

    Terry, Mary Beth; Neugut, Alfred I; Mansukhani, Mahesh; Waye, Jerome; Harpaz, Noam; Hibshoosh, Hanina

    2003-01-01

    The p53 tumor suppressor gene is commonly mutated in colorectal cancer. While the effect of p53 mutations on colorectal cancer prognosis has been heavily studied, less is known about how epidemiologic risk factors relate to p53 status, particularly in early colorectal neoplasia prior to clinically invasive colorectal cancer (including adenomas, carcinoma in situ (CIS), and intramucosal carcinoma). We examined p53 status, as measured by protein overexpression, in 157 cases with early colorectal neoplasia selected from three New York City colonoscopy clinics. After collecting paraffin-embedded tissue blocks, immunohistochemistry was performed using an anti-p53 monoclonal mouse IgG 2 a [BP53-12-1] antibody. We analyzed whether p53 status was different for risk factors for colorectal neoplasia relative to a polyp-free control group (n = 508). p53 overexpression was found in 10.3%, 21.7%, and 34.9%, of adenomatous polyps, CIS, and intramucosal cases, respectively. Over 90% of the tumors with p53 overexpression were located in the distal colon and rectum. Heavy cigarette smoking (30+ years) was associated with cases not overexpressing p53 (OR = 1.8, 95% CI = 1.1–2.9) but not with those cases overexpressing p53 (OR = 1.0, 95% CI = 0.4–2.6). Heavy beer consumption (8+ bottles per week) was associated with cases overexpressing p53 (OR = 4.0, 95% CI = 1.3–12.0) but not with cases without p53 overexpression (OR = 1.6, 95% CI = 0.7–3.7). Our findings that p53 overexpression in early colorectal neoplasia may be positively associated with alcohol intake and inversely associated with cigarette smoking are consistent with those of several studies of p53 expression and invasive cancer, and suggest that there may be relationships of smoking and alcohol with p53 early in the adenoma to carcinoma sequence

  4. MsZEP, a novel zeaxanthin epoxidase gene from alfalfa (Medicago sativa), confers drought and salt tolerance in transgenic tobacco.

    Science.gov (United States)

    Zhang, Zhiqiang; Wang, Yafang; Chang, Leqin; Zhang, Tong; An, Jie; Liu, Yushi; Cao, Yuman; Zhao, Xia; Sha, Xuyang; Hu, Tianming; Yang, Peizhi

    2016-02-01

    The zeaxanthin epoxidase gene ( MsZEP ) was cloned and characterized from alfalfa and validated for its function of tolerance toward drought and salt stresses by heterologous expression in Nicotiana tabacum. Zeaxanthin epoxidase (ZEP) plays important roles in plant response to various environment stresses due to its functions in ABA biosynthetic and the xanthophyll cycle. To understand the expression characteristics and the biological functions of ZEP in alfalfa (Medicago sativa), a novel gene, designated as MsZEP (KM044311), was cloned, characterized and overexpressed in Nicotiana tabacum. The open reading frame of MsZEP contains 1992 bp nucleotides and encodes a 663-amino acid polypeptide. Amino acid sequence alignment indicated that deduced MsZEP protein was highly homologous to other plant ZEP sequences. Phylogenetic analysis showed that MsZEP was grouped into a branch with other legume plants. Real-time quantitative PCR revealed that MsZEP gene expression was clearly tissue-specific, and the expression levels were higher in green tissues (leaves and stems) than in roots. MsZEP expression decreased in shoots under drought, cold, heat and ABA treatment, while the expression levels in roots showed different trends. Besides, the results showed that nodules could up-regulate the MsZEP expression under non-stressful conditions and in the earlier stage of different abiotic stress. Heterologous expression of the MsZEP gene in N. tabacum could confer tolerance to drought and salt stress by affecting various physiological pathways, ABA levels and stress-responsive genes expression. Taken together, these results suggested that the MsZEP gene may be involved in alfalfa responses to different abiotic stresses and nodules, and could enhance drought and salt tolerance of transgenic tobacco by heterologous expression.

  5. Ectopic Overexpression of a Novel R2R3-MYB, NtMYB2 from Chinese Narcissus Represses Anthocyanin Biosynthesis in Tobacco

    Directory of Open Access Journals (Sweden)

    Muhammad Anwar

    2018-03-01

    Full Text Available R2R3 MYB transcription factors play key functions in the regulation of secondary metabolites. In the present study, a R2R3 MYB transcriptional factor NtMYB2 was identified from Chinese narcissus (Narcissus tazetta L. var. Chinensis Roem and functionally characterized. NtMYB2 belongs to subgroup 4 of the R2R3 MYB transcription factor family that are related to repressor MYBs involved in the regulation of anthocyanin and flavonoids. Transient expression confirmed that NtMYB2 strongly reduced the red pigmentation induced by MYB- anthocyanin activators in agro-infiltrated tobacco leaves. Ectopic expression of NtMYB2 in tobacco significantly reduced the pigmentation and altered the floral phenotypes in transgenic tobacco flowers. Gene expression analysis suggested that NtMYB2 repressed the transcript levels of structural genes involved in anthocyanin biosynthesis pathway, especially the UFGT gene. NtMYB2 gene is expressed in all examined narcissus tissues; the levels of transcription in petals and corona is higher than other tissues and the transcription level at the bud stage was highest. These results show that NtMYB2 is involved in the regulation of anthocyanin biosynthesis pathway and may act as a repressor by down regulating the transcripts of key enzyme genes in Chinese narcissus.

  6. Ectopic Overexpression of a Novel R2R3-MYB, NtMYB2 from Chinese Narcissus Represses Anthocyanin Biosynthesis in Tobacco.

    Science.gov (United States)

    Anwar, Muhammad; Wang, Guiqing; Wu, Jiacheng; Waheed, Saquib; Allan, Andrew C; Zeng, Lihui

    2018-03-28

    R2R3 MYB transcription factors play key functions in the regulation of secondary metabolites. In the present study, a R2R3 MYB transcriptional factor NtMYB2 was identified from Chinese narcissus ( Narcissus tazetta L. var. Chinensis Roem) and functionally characterized. NtMYB2 belongs to subgroup 4 of the R2R3 MYB transcription factor family that are related to repressor MYBs involved in the regulation of anthocyanin and flavonoids. Transient expression confirmed that NtMYB2 strongly reduced the red pigmentation induced by MYB- anthocyanin activators in agro-infiltrated tobacco leaves. Ectopic expression of NtMYB2 in tobacco significantly reduced the pigmentation and altered the floral phenotypes in transgenic tobacco flowers. Gene expression analysis suggested that NtMYB2 repressed the transcript levels of structural genes involved in anthocyanin biosynthesis pathway, especially the UFGT gene. NtMYB2 gene is expressed in all examined narcissus tissues; the levels of transcription in petals and corona is higher than other tissues and the transcription level at the bud stage was highest. These results show that NtMYB2 is involved in the regulation of anthocyanin biosynthesis pathway and may act as a repressor by down regulating the transcripts of key enzyme genes in Chinese narcissus.

  7. Overexpression of HARDY, an AP2/ERF gene from Arabidopsis, improves drought and salt tolerance by reducing transpiration and sodium uptake in transgenic Trifolium alexandrinum L.

    Science.gov (United States)

    Abogadallah, Gaber M; Nada, Reham M; Malinowski, Robert; Quick, Paul

    2011-06-01

    Trifolium alexandrinum L. was transformed with the Arabidopsis HARDY gene that belongs to the stress-related AP2/ERF (APETALA2/ethylene responsive element binding factors) superfamily of transcription factors. The fresh weights of the transgenic lines L2 and L3 were improved by 42 and 55% under drought stress and by 38 and 95% under salt stress compared to the wild type, respectively. The dry weights were similarly improved. Overexpression of HARDY improved the instantaneous water use efficiency (WUE) under drought stress by reducing transpiration (E) and under salt stress by improving photosynthesis (A), through reducing Na+ accumulation in leaves, and reducing E. However, HARDY improved the growth of drought-stressed transgenic plants as compared to the wild type by delaying water depletion from soil and preventing rapid decline in A. L2 and L3 had thicker stems and in case of L3, more xylem rows per vascular bundle, which may have made L3 more resistant to lodging in the field. Field performance of L2 and L3 under combined drought and salt stress was significantly better than that of the wild type in terms of fresh and dry weights (40%, 46% and 31%, 40%, respectively). The results provide further evidence for the efficiency of overexpression of a single gene in improving tolerance to abiotic stress under field conditions.

  8. DECOMPOSTION OF GENETICALLY ENGINEERED TOBACCO UNDER FIELD CONDITIONS: PERSISTENCE OF THE PROTEINASE INHIBITOR I PRODUCT AND EFFECTS OF SOIL MICROBIAL RESPIRATION AND PROTOZOA, NEMATODE AND MICROARTHR

    Science.gov (United States)

    1. To evaluate the potential effects of genetically engineered (transgenic) plants on soil ecosystems, litterbags containing leaves of non-engineered (parental) and transgenic tobacco plants were buried in field plots. The transgenic tobacco plants were genetically engineered to ...

  9. Characterization of amyloid beta peptides from brain extracts of transgenic mice overexpressing the London mutant of human amyloid precursor protein.

    Science.gov (United States)

    Pype, Stefan; Moechars, Dieder; Dillen, Lieve; Mercken, Marc

    2003-02-01

    Alzheimer's disease (AD) is marked by the presence of neurofibrillary tangles and amyloid plaques in the brain of patients. To study plaque formation, we report on further quantitative and qualitative analysis of human and mouse amyloid beta peptides (Abeta) from brain extracts of transgenic mice overexpressing the London mutant of human amyloid precursor protein (APP). Using enzyme-linked immunosorbant assays (ELISAs) specific for either human or rodent Abeta, we found that the peptides from both species aggregated to form plaques. The ratios of deposited Abeta1-42/1-40 were in the order of 2-3 for human and 8-9 for mouse peptides, indicating preferential deposition of Abeta42. We also determined the identity and relative levels of other Abeta variants present in protein extracts from soluble and insoluble brain fractions. This was done by combined immunoprecipitation and mass spectrometry (IP/MS). The most prominent peptides truncated either at the carboxyl- or the amino-terminus were Abeta1-38 and Abeta11-42, respectively, and the latter was strongly enriched in the extracts of deposited peptides. Taken together, our data indicate that plaques of APP-London transgenic mice consist of aggregates of multiple human and mouse Abeta variants, and the human variants that we identified were previously detected in brain extracts of AD patients.

  10. Recombinant Promoter (MUASCsV8CP) Driven Totiviral Killer Protein 4 (KP4) Imparts Resistance Against Fungal Pathogens in Transgenic Tobacco

    Science.gov (United States)

    Deb, Debasish; Shrestha, Ankita; Maiti, Indu B.; Dey, Nrisingha

    2018-01-01

    Development of disease-resistant plant varieties achieved by engineering anti-microbial transgenes under the control of strong promoters can suffice the inhibition of pathogen growth and simultaneously ensure enhanced crop production. For evaluating the prospect of such strong promoters, we comprehensively characterized the full-length transcript promoter of Cassava Vein Mosaic Virus (CsVMV; -565 to +166) and identified CsVMV8 (-215 to +166) as the highest expressing fragment in both transient and transgenic assays. Further, we designed a new chimeric promoter ‘MUASCsV8CP’ through inter-molecular hybridization among the upstream activation sequence (UAS) of Mirabilis Mosaic Virus (MMV; -297 to -38) and CsVMV8, as the core promoter (CP). The MUASCsV8CP was found to be ∼2.2 and ∼2.4 times stronger than the CsVMV8 and CaMV35S promoters, respectively, while its activity was found to be equivalent to that of the CaMV35S2 promoter. Furthermore, we generated transgenic tobacco plants expressing the totiviral ‘Killer protein KP4’ (KP4) under the control of the MUASCsV8CP promoter. Recombinant KP4 was found to accumulate both in the cytoplasm and apoplast of plant cells. The agar-based killing zone assays revealed enhanced resistance of plant-derived KP4 against two deuteromycetous foliar pathogenic fungi viz. Alternaria alternata and Phoma exigua var. exigua. Also, transgenic plants expressing KP4 inhibited the growth progression of these fungi and conferred significant fungal resistance in detached-leaf and whole plant assays. Taken together, we establish the potential of engineering “in-built” fungal stress-tolerance in plants by expressing KP4 under a novel chimeric caulimoviral promoter in a transgenic approach. PMID:29556246

  11. Overexpression of Poplar Pyrabactin Resistance-Like Abscisic Acid Receptors Promotes Abscisic Acid Sensitivity and Drought Resistance in Transgenic Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Jingling Yu

    Full Text Available Drought stress is an important environmental factor limiting productivity of plants, especially fast growing species with high water consumption like poplar. Abscisic acid (ABA is a phytohormone that positively regulates seed dormancy and drought resistance. The PYR1 (Pyrabactin Resistance 1/ PYRL (PYR-Like/ RCAR (Regulatory Component of ABA Receptor (PYR/PYL/RCAR ABA receptor family has been identified and widely characterized in Arabidopsis thaliana. However, their functions in poplars remain unknown. Here, we report that 2 of 14 PYR/PYL/RCAR orthologues in poplar (Populus trichocarpa (PtPYRLs function as a positive regulator of the ABA signal transduction pathway. The Arabidopsis transient expression and yeast two-hybrid assays showed the interaction among PtPYRL1 and PtPYRL5, a clade A protein phosphatase 2C, and a SnRK2, suggesting that a core signalling complex for ABA signaling pathway exists in poplars. Phenotypic analysis of PtPYRL1 and PtPYRL5 transgenic Arabidopsis showed that these two genes positively regulated the ABA responses during the seed germination. More importantly, the overexpression of PtPYRL1 and PtPYRL5 substantially improved ABA sensitivity and drought stress tolerance in transgenic plants. In summary, we comprehensively uncovered the properties of PtPYRL1 and PtPYRL5, which might be good target genes to genetically engineer drought-Resistant plants.

  12. Photosynthesis and protective mechanisms during ageing in transgenic tobacco leaves with over-expressed cytokinin oxidase/dehydrogenase and thus lowered cytokinin content

    Czech Academy of Sciences Publication Activity Database

    Mýtinová, Zuzana; Haisel, Daniel; Wilhelmová, Naděžda

    2006-01-01

    Roč. 44, č. 4 (2006), s. 599-605 ISSN 0300-3604 R&D Projects: GA ČR GA522/03/0312 Institutional research plan: CEZ:AV0Z50380511 Keywords : senescence * cytokinins * carotenoids * chlorophyll * xanthophylls cycle * photosynthetic efficiency * chlorophyll fluorescence * tobacco Subject RIV: CE - Biochemistry Impact factor: 0.782, year: 2006

  13. Overexpression of SmMYC2 Increases the Production of Phenolic Acids in Salvia miltiorrhiza

    Directory of Open Access Journals (Sweden)

    Na Yang

    2017-10-01

    Full Text Available MYC2 is a core transcription factor in the plant response to jasmonates. It also functions in secondary metabolism and various processes for growth and development. However, the knowledge about its role in Salvia miltiorrhiza is still very limited. We determined that the biosynthesis of salvianolic acid B (Sal B was strongly induced in 2-month-old transgenic plants that over-expressed SmMYC2. In the roots of transgenic line 12 that over-expressed SmMYC2 (OEM-12, the Sal B concentration was as high as 5.95 ± 0.07 mg g-1, a level that was 1.88-fold higher than that in control plants that had been transformed with an empty vector. Neither tanshinone IIA nor cryptotanshinone was detected by high-performance liquid chromatography in any of the genotypes. Global transcriptomic analysis using RNA sequencing revealed that most enzyme-encoding genes for the phenylpropanoid biosynthesis pathway were up-regulated in the overexpression lines. Furthermore, both the phenylalanine and tyrosine biosynthesis pathways were activated in those transgenics. Our data demonstrate that overexpression of SmMYC2 promotes the production of phenolic acids by simultaneously activating both primary and secondary pathways for metabolism in S. miltiorrhiza.

  14. [A novel gene (Aa-accA ) encoding acetyl-CoA carboxyltransferase alpha-subunit of Alkalimonas amylolytica N10 enhances salt and alkali tolerance of Escherichia coli and tobacco BY-2 cells].

    Science.gov (United States)

    Xian, Mingjie; Zhai, Lei; Zhong, Naiqin; Ma, Yiwei; Xue, Yanfen; Ma, Yanhe

    2013-08-04

    Acetyl-CoA carboxylase (ACC) catalyzes the first step of fatty acid synthesis. In most bacteria, ACC is composed of four subunits encoded by accA, accB, accC, and accD. Of them, accA encodes acetyl-CoA carboxyltransferase alpha-subunit. Our prior work on proteomics of Alkalimonas amylolytica N10 showed that the expression of the Aa-accA has a remarkable response to salt and alkali stress. This research aimed to find out the Aa-accA gene contributing to salt and alkali tolerance. The Aa-accA was amplified by PCR from A. amylolytica N10 and expressed in E. coli K12 host. The effects of Aa-accA expression on the growth of transgenic strains were examined under different NaCl concentration and pH conditions. Transgenic tobacco BY-2 cells harboring Aa-accA were also generated via Agrobacterium-mediated transformation. The viability of BY-2 cells was determined with FDA staining method after salt and alkali shock. The Aa-accA gene product has 318 amino acids and is homologous to the carboxyl transferase domain of acyl-CoA carboxylases. It showed 76% identity with AccA (acetyl-CoA carboxylase carboxyltransferase subunit alpha) from E. coli. Compared to the wild-type strains, transgenic E. coli K12 strain containing Aa-accA showed remarkable growth superiority when grown in increased NaCl concentrations and pH levels. The final cell density of the transgenic strains was 2.6 and 3.5 times higher than that of the control type when they were cultivated in LB medium containing 6% (W/V) NaCl and at pH 9, respectively. Complementary expression of Aa-accA in an accA-depletion E. coli can recover the tolerance of K12 delta accA to salt and alkali stresses to some extent. Similar to the transgenic E. coli, transgenic tobacco BY-2 cells showed higher percentages of viability compared to the wild BY-2 cells under the salt or alkali stress condition. We found that Aa-accA from A. amylolytica N10 overexpression enhances the tolerance of both transgenic E. coli and tobacco BY-2 cells to

  15. Overexpression of a natural chloroplast-encoded antisense RNA in tobacco destabilizes 5S rRNA and retards plant growth

    Directory of Open Access Journals (Sweden)

    Stern David B

    2010-09-01

    Full Text Available Abstract Background The roles of non-coding RNAs in regulating gene expression have been extensively studied in both prokaryotes and eukaryotes, however few reports exist as to their roles in organellar gene regulation. Evidence for accumulation of natural antisense RNAs (asRNAs in chloroplasts comes from the expressed sequence tag database and cDNA libraries, while functional data have been largely obtained from artificial asRNAs. In this study, we used Nicotiana tabacum to investigate the effect on sense strand transcripts of overexpressing a natural chloroplast asRNA, AS5, which is complementary to the region which encodes the 5S rRNA and tRNAArg. Results AS5-overexpressing (AS5ox plants obtained by chloroplast transformation exhibited slower growth and slightly pale green leaves. Analysis of AS5 transcripts revealed four distinct species in wild-type (WT and AS5ox plants, and additional AS5ox-specific products. Of the corresponding sense strand transcripts, tRNAArg overaccumulated several-fold in transgenic plants whereas 5S rRNA was unaffected. However, run-on transcription showed that the 5S-trnR region was transcribed four-fold more in the AS5ox plants compared to WT, indicating that overexpression of AS5 was associated with decreased stability of 5S rRNA. In addition, polysome analysis of the transformants showed less 5S rRNA and rbcL mRNA associated with ribosomes. Conclusions Our results suggest that AS5 can modulate 5S rRNA levels, giving it the potential to affect Chloroplast translation and plant growth. More globally, overexpression of asRNAs via chloroplast transformation may be a useful strategy for defining their functions.

  16. Neuroglobin Overexpression Inhibits AMPK Signaling and Promotes Cell Anabolism.

    Science.gov (United States)

    Cai, Bin; Li, Wenjun; Mao, XiaoOu; Winters, Ali; Ryou, Myoung-Gwi; Liu, Ran; Greenberg, David A; Wang, Ning; Jin, Kunlin; Yang, Shao-Hua

    2016-03-01

    Neuroglobin (Ngb) is a recently discovered globin with preferential localization to neurons. Growing evidence indicates that Ngb has distinct physiological functions separate from the oxygen storage and transport roles of other globins, such as hemoglobin and myoglobin. We found increased ATP production and decreased glycolysis in Ngb-overexpressing immortalized murine hippocampal cell line (HT-22), in parallel with inhibition of AMP-activated protein kinase (AMPK) signaling and activation of acetyl-CoA carboxylase (ACC). In addition, lipid and glycogen content was increased in Ngb-overexpressing HT-22 cells. AMPK signaling was also inhibited in the brain and heart from Ngb-overexpressing transgenic mice. Although Ngb overexpression did not change glycogen content in whole brain, glycogen synthase was activated in cortical neurons of Ngb-overexpressing mouse brain and Ngb overexpression primary neurons. Moreover, lipid and glycogen content was increased in hearts derived from Ngb-overexpressing mice. These findings suggest that Ngb functions as a metabolic regulator and enhances cellular anabolism through the inhibition of AMPK signaling.

  17. Transgenic Centipedegrass (Eremochloa ophiuroides [Munro] Hack. Overexpressing S-Adenosylmethionine Decarboxylase (SAMDC Gene for Improved Cold Tolerance Through Involvement of H2O2 and NO Signaling

    Directory of Open Access Journals (Sweden)

    Jianhao Luo

    2017-09-01

    Full Text Available Centipedegrass (Eremochloa ophiuroides [Munro] Hack. is an important warm-season turfgrass species. Transgenic centipedgrass plants overexpressing S-adenosylmethionine decarboxylase from bermudagrass (CdSAMDC1 that was induced in response to cold were generated in this study. Higher levels of CdSAMDC1 transcript and sperimidine (Spd and spermin (Spm concentrations and enhanced freezing and chilling tolerance were observed in transgenic plants as compared with the wild type (WT. Transgenic plants had higher levels of polyamine oxidase (PAO activity and H2O2 than WT, which were blocked by pretreatment with methylglyoxal bis (guanylhydrazone or MGBG, inhibitor of SAMDC, indicating that the increased PAO and H2O2 were a result of expression of CdSAMDC1. In addition, transgenic plants had higher levels of nitrate reductase (NR activity and nitric oxide (NO concentration. The increased NR activity were blocked by pretreatment with MGBG and ascorbic acid (AsA, scavenger of H2O2, while the increased NO level was blocked by MGBG, AsA, and inhibitors of NR, indicating that the enhanced NR-derived NO was dependent upon H2O2, as a result of expression CdSAMDC1. Elevated superoxide dismutase (SOD and catalase (CAT activities were observed in transgenic plants than in WT, which were blocked by pretreatment with MGBG, AsA, inhibitors of NR and scavenger of NO, indicating that the increased activities of SOD and CAT depends on expression of CdSAMDC1, H2O2, and NR-derived NO. Our results suggest that the elevated cold tolerance was associated with PAO catalyzed production of H2O2, which in turn led to NR-derived NO production and induced antioxidant enzyme activities in transgenic plants.

  18. Transgenic Centipedegrass (Eremochloa ophiuroides [Munro] Hack.) Overexpressing S-Adenosylmethionine Decarboxylase (SAMDC) Gene for Improved Cold Tolerance Through Involvement of H2O2 and NO Signaling.

    Science.gov (United States)

    Luo, Jianhao; Liu, Mingxi; Zhang, Chendong; Zhang, Peipei; Chen, Jingjing; Guo, Zhenfei; Lu, Shaoyun

    2017-01-01

    Centipedegrass ( Eremochloa ophiuroides [Munro] Hack.) is an important warm-season turfgrass species. Transgenic centipedgrass plants overexpressing S-adenosylmethionine decarboxylase from bermudagrass ( CdSAMDC1 ) that was induced in response to cold were generated in this study. Higher levels of CdSAMDC1 transcript and sperimidine (Spd) and spermin (Spm) concentrations and enhanced freezing and chilling tolerance were observed in transgenic plants as compared with the wild type (WT). Transgenic plants had higher levels of polyamine oxidase (PAO) activity and H 2 O 2 than WT, which were blocked by pretreatment with methylglyoxal bis (guanylhydrazone) or MGBG, inhibitor of SAMDC, indicating that the increased PAO and H 2 O 2 were a result of expression of CdSAMDC1 . In addition, transgenic plants had higher levels of nitrate reductase (NR) activity and nitric oxide (NO) concentration. The increased NR activity were blocked by pretreatment with MGBG and ascorbic acid (AsA), scavenger of H 2 O 2 , while the increased NO level was blocked by MGBG, AsA, and inhibitors of NR, indicating that the enhanced NR-derived NO was dependent upon H 2 O 2 , as a result of expression CdSAMDC1 . Elevated superoxide dismutase (SOD) and catalase (CAT) activities were observed in transgenic plants than in WT, which were blocked by pretreatment with MGBG, AsA, inhibitors of NR and scavenger of NO, indicating that the increased activities of SOD and CAT depends on expression of CdSAMDC1 , H 2 O 2 , and NR-derived NO. Our results suggest that the elevated cold tolerance was associated with PAO catalyzed production of H 2 O 2 , which in turn led to NR-derived NO production and induced antioxidant enzyme activities in transgenic plants.

  19. Enhancement of ginsenoside Rg(1) in Panax ginseng hairy root by overexpressing the α-L-rhamnosidase gene from Bifidobacterium breve.

    Science.gov (United States)

    Zhang, Ru; Zhang, Bian-Ling; Li, Gu-Cai; Xie, Tao; Hu, Teng; Luo, Zhi-Yong

    2015-10-01

    To improve the production of ginsenoside Rg1 in Panax ginseng. The α-L-rhamnosidase gene from Bifidobacterium breve (BbRha) was overexpressed into hairy root culture system using Agrobacterium rhizogenes A4. Ginsenoside Rg1 in hairy roots was obtained following transformation via overexpressed gene representing 2.2-fold higher than those of control lines. Several overexpression transgenic hairy root lines were obtained exhibiting markedly increased levels of the corresponding α-L-rhamnosidase enzymatic activity relative to control. Ginsenoside Rg1 levels in the transgenic lines were higher (2.2-fold) than those of control after following 30 days culturing, while ginsenoside Re contents in tested transgenic lines were found to be lower. The transgenic hairy roots harboring α-L-rhamnosidase gene improved the accumulation of ginsenoside Rg1 up to 3.6 mg g(-1) dry weight. BbRha gene selectively enhances the production of ginsenoside Rg1 in P. ginseng hairy roots.

  20. PKCepsilon overexpression, irrespective of genetic background, sensitizes skin to UVR-induced development of squamous-cell carcinomas.

    Science.gov (United States)

    Sand, Jordan M; Aziz, Moammir H; Dreckschmidt, Nancy E; Havighurst, Thomas C; Kim, KyungMann; Oberley, Terry D; Verma, Ajit K

    2010-01-01

    Chronic exposure to UVR is the major etiologic factor in the development of human skin cancers including squamous-cell carcinoma (SCC). We have previously shown that protein Kinase C epsilon (PKCepsilon) transgenic mice on FVB/N background, which overexpress PKCepsilon protein approximately eightfold over endogenous levels in epidermis, exhibit about threefold more sensitivity than wild-type littermates to UVR-induced development of SCC. To determine whether it is PKCepsilon and not the mouse genetic background that determines susceptibility to UVR carcinogenesis, we cross-bred PKCepsilon FVB/N transgenic mice with SKH-1 hairless mice to generate PKCepsilon-overexpressing SKH-1 hairless mice. To evaluate the susceptibility of PKCepsilon SKH-1 hairless transgenic mice to UVR carcinogenesis, the mice were exposed to UVR (1-2 KJ m(-2)) three times weekly from a bank of six kodacel-filtered FS40 sunlamps. As compared with the wild-type hairless mice, PKCepsilon overexpression in SKH-1 hairless mice decreased the latency (12 weeks), whereas it increased the incidence (twofold) and multiplicity (fourfold) of SCC. The SKH hairless transgenic mice were observed to be as sensitive as FVB/N transgenic mice to UVR-induced development of SCC and expression of proliferative markers (proliferating cell nuclear antigen, signal transducers and activators of transcription 3, and extracellular signal-regulated kinase 1/2). The results indicate that PKCepsilon level dictates susceptibility, irrespective of genetic background, to UVR carcinogenesis.

  1. Increased production of wax esters in transgenic tobacco plants by expression of a fatty acid reductase:wax synthase gene fusion.

    Science.gov (United States)

    Aslan, Selcuk; Hofvander, Per; Dutta, Paresh; Sun, Chuanxin; Sitbon, Folke

    2015-12-01

    Wax esters are hydrophobic lipids consisting of a fatty acid moiety linked to a fatty alcohol with an ester bond. Plant-derived wax esters are today of particular concern for their potential as cost-effective and sustainable sources of lubricants. However, this aspect is hampered by the fact that the level of wax esters in plants generally is too low to allow commercial exploitation. To investigate whether wax ester biosynthesis can be increased in plants using transgenic approaches, we have here exploited a fusion between two bacterial genes together encoding a single wax ester-forming enzyme, and targeted the resulting protein to chloroplasts in stably transformed tobacco (Nicotiana benthamiana) plants. Compared to wild-type controls, transgenic plants showed both in leaves and stems a significant increase in the total level of wax esters, being eight-fold at the whole plant level. The profiles of fatty acid methyl ester and fatty alcohol in wax esters were related, and C16 and C18 molecules constituted predominant forms. Strong transformants displayed certain developmental aberrations, such as stunted growth and chlorotic leaves and stems. These negative effects were associated with an accumulation of fatty alcohols, suggesting that an adequate balance between formation and esterification of fatty alcohols is crucial for a high wax ester production. The results show that wax ester engineering in transgenic plants is feasible, and suggest that higher yields may become achieved in the near future.

  2. Oxidized SOD1 alters proteasome activities in vitro and in the cortex of SOD1 overexpressing mice.

    Science.gov (United States)

    Le Pecheur, Marie; Bourdon, Emmanuel; Paly, Evelyne; Farout, Luc; Friguet, Bertrand; London, Jacqueline

    2005-07-04

    Premature ageing, one of the characteristics of Down syndrome (DS), may involve oxidative stress and impairment of proteasome activity. Transgenic mice overexpressing the human copper/zinc superoxide dismutase (SOD1) gene are one of the first murine models for DS and it has been shown that SOD1 overexpression might be either deleterious or beneficial. Here, we show a reduction in proteasome activities in the cortex of SOD1 transgenic mice and an associated increase in the content of oxidized SOD1 protein. As we demonstrate that in vitro oxidized SOD can inhibit purified proteasome peptidase activities, modified SOD1 might be partially responsible for proteasome inhibition shown in SOD1 transgenic mice.

  3. Overexpression of Arachis hypogaea AREB1 Gene Enhances Drought Tolerance by Modulating ROS Scavenging and Maintaining Endogenous ABA Content

    Directory of Open Access Journals (Sweden)

    Ling Li

    2013-06-01

    Full Text Available AhAREB1 (Arachis hypogaea Abscisic-acid Response Element Binding Protein 1 is a member of the basic domain leucine zipper (bZIP-type transcription factor in peanut. Previously, we found that expression of AhAREB1 was specifically induced by abscisic acid (ABA, dehydration and drought. To understand the drought defense mechanism regulated by AhAREB1, transgenic Arabidopsis overexpressing AhAREB1 was conducted in wild-type (WT, and a complementation experiment was employed to ABA non-sensitivity mutant abi5 (abscisic acid-insensitive 5. Constitutive expression of AhAREB1 confers water stress tolerance and is highly sensitive to exogenous ABA. Microarray and further real-time PCR analysis revealed that drought stress, reactive oxygen species (ROS scavenging, ABA synthesis/metabolism-related genes and others were regulated in transgenic Arabidopsis overexpressing AhAREB1. Accordingly, low level of ROS, but higher ABA content was detected in the transgenic Arabidopsis plants’ overexpression of AhAREB1. Taken together, it was concluded that AhAREB1 modulates ROS accumulation and endogenous ABA level to improve drought tolerance in transgenic Arabidopsis.

  4. Overexpressing both ATP sulfurylase and selenocysteine methyltransferase enhances selenium phytoremediation traits in Indian mustard

    International Nuclear Information System (INIS)

    LeDuc, Danika L.; AbdelSamie, Manal; Montes-Bayon, Maria; Wu, Carol P.; Reisinger, Sarah J.; Terry, Norman

    2006-01-01

    A major goal of our selenium (Se) phytoremediation research is to use genetic engineering to develop fast-growing plants with an increased ability to tolerate, accumulate, and volatilize Se. To this end we incorporated a gene (encoding selenocysteine methyltransferase, SMT) from the Se hyperaccumulator, Astragalus bisulcatus, into Indian mustard (LeDuc, D.L., Tarun, A.S., Montes-Bayon, M., Meija, J., Malit, M.F., Wu, C.P., AbdelSamie, M., Chiang, C.-Y., Tagmount, A., deSouza, M., Neuhierl, B., Boeck, A., Caruso, J., Terry, N., 2004. Overexpression of selenocysteine methyltransferase in Arabidopsis and Indian mustard increases selenium tolerance and accumulation Plant Physiol. 135, 377-383.). The resulting transgenic plants successfully enhanced Se phytoremediation in that the plants tolerated and accumulated Se from selenite significantly better than wild type. However, the advantage conferred by the SMT enzyme was much less when Se was supplied as selenate. In order to enhance the phytoremediation of selenate, we developed double transgenic plants that overexpressed the gene encoding ATP sulfurylase (APS) in addition to SMT, i.e., APS x SMT. The results showed that there was a substantial improvement in Se accumulation from selenate (4 to 9 times increase) in transgenic plants overexpressing both APS and SMT. - Simultaneous overexpression of APS and SMT genes in Indian mustard greatly increases ability to accumulate selenate

  5. Reduced Position Effect in Mature Transgenic Plants Conferred by the Chicken Lysozyme Matrix-Associated Region

    NARCIS (Netherlands)

    Mlynárová, Ľudmila; Loonen, Annelies; Heldens, Jos; Jansen, Ritsert C.; Keizer, Paul; Stiekema, Willem J.; Nap, Jan-Peter

    1994-01-01

    Matrix-associated regions may be useful for studying the role of chromatin architecture in transgene activity of transformed plants. The chicken lysozyme A element was shown to have specific affinity for tobacco nuclear matrices, and its influence on the variability of transgene expression in

  6. Transgenic plants expressing GLK1 and CCA1 having increased nitrogen assimilation capacity

    Science.gov (United States)

    Coruzzi, Gloria [New York, NY; Gutierrez, Rodrigo A [Santiago, CL; Nero, Damion C [Woodside, NY

    2012-04-10

    Provided herein are compositions and methods for producing transgenic plants. In specific embodiments, transgenic plants comprise a construct comprising a polynucleotide encoding CCA1, GLK1 or bZIP1, operably linked to a plant-specific promote, wherein the CCA1, GLK1 or bZIP1 is ectopically overexpressed in the transgenic plants, and wherein the promoter is optionally a constitutive or inducible promoter. In other embodiments, transgenic plants in which express a lower level of CCA1, GLK1 or bZIP1 are provided. Also provided herein are commercial products (e.g., pulp, paper, paper products, or lumber) derived from the transgenic plants (e.g., transgenic trees) produced using the methods provided herein.

  7. Overexpression of allene oxide cyclase improves the biosynthesis of artemisinin in Artemisia annua L.

    Directory of Open Access Journals (Sweden)

    Xu Lu

    Full Text Available Jasmonates (JAs are important signaling molecules in plants and play crucial roles in stress responses, secondary metabolites' regulation, plant growth and development. In this study, the promoter of AaAOC, which was the key gene of jasmonate biosynthetic pathway, had been cloned. GUS staining showed that AaAOC was expressed ubiquitiously in A. annua. AaAOC gene was overexpressed under control of 35S promoter. RT-Q-PCR showed that the expression levels of AaAOC were increased from 1.6- to 5.2-fold in AaAOC-overexpression transgenic A. annua. The results of GC-MS showed that the content of endogenous jasmonic acid (JA was 2- to 4.7-fold of the control level in AaAOC-overexpression plants. HPLC showed that the contents of artemisinin, dihydroartemisinic acid and artemisinic acid were increased significantly in AaAOC-overexpression plants. RT-Q-PCR showed that the expression levels of FPS (farnesyl diphosphate synthase, CYP71AV1 (cytochrome P450 dependent hydroxylase and DBR2 (double bond reductase 2 were increased significantly in AaAOC-overexpression plants. All data demonstrated that increased endogenous JA could significantly promote the biosynthesis of artemisinin in AaAOC-overexpression transgenic A. annua.

  8. Transgenic mice overexpressing glia maturation factor-β, an oxidative stress inducible gene, show premature aging due to Zmpste24 down-regulation.

    Science.gov (United States)

    Imai, Rika; Asai, Kanae; Hanai, Jun-ichi; Takenaka, Masaru

    2015-07-01

    Glia Maturation Factor-β (GMF), a brain specific protein, is induced by proteinuria in renal tubules. Ectopic GMF overexpression causes apoptosisin vitro via cellular vulnerability to oxidative stress. In order to examine the roles of GMF in non-brain tissue, we constructed transgenic mice overexpressing GMF (GMF-TG). The GMF-TG mice exhibited appearance phenotypes associated with premature aging. The GMF-TG mice also demonstrated short lifespans and reduced hair regrowth, suggesting an accelerated aging process. The production of an abnormal lamin A, a nuclear envelope protein, plays a causal role in both normal aging and accelerated aging diseases, known as laminopathies. Importantly, we identified the abnormal lamin A (prelamin A), accompanied by a down-regulation of a lamin A processing enzyme (Zmpste24) in the kidney of the GMF-TG mice. The GMF-TG mice showed accelerated aging in the kidney, compared with wild-type mice, showing increased TGF-β1, CTGF gene and serum creatinine. The gene expression of p21/waf1 was increased at an earlier stage of life, at 10 weeks, which was in turn down-regulated at a later stage, at 60 weeks. In conclusion, we propose that GMF-TG mice might be a novel mouse model of accelerated aging, due to the abnormal lamin A.

  9. Differential antioxidative responses in transgenic peanut bear no relationship to their superior transpiration efficiency under drought stress.

    Science.gov (United States)

    Bhatnagar-Mathur, Pooja; Devi, M Jyostna; Vadez, Vincent; Sharma, Kiran K

    2009-07-15

    To counter the effects of environmental stresses, the plants must undergo detoxification that is crucial to avoid the accumulation of damaging free oxygen radicals (ROI). Here, we detail the oxidative damage, the antioxidant composition, and the osmoprotection achieved in transgenic plants of peanut overexpressing the AtDREB1A transgene, driven by a stress-inducible promoter (Atrd29A) when exposed to progressive water stress conditions. This study explored the biochemical mechanisms where (i) the antioxidants such as superoxide dismutase (SOD), ascorbate peroxidase (APOX), and glutathione reductase (GR) accumulated in the transgenic plants at comparably higher levels than their untransformed counterparts under dry soil conditions, (ii) a significant increase in the proline levels in the transgenic plants was observed in dry soils, and (iii) a dramatic increase in the lipid peroxidation in the untransformed controls in drier soils. Most of the biochemical parameters related to the antioxidative machinery in the tested peanut transgenics were triggered by the overexpression of AtDREB1A that appeared to differ from the untransformed controls. The antioxidants showed a negative correlation with the fraction of transpirable soil water (FTSW) thresholds, where the normalized transpiration rate (NTR) started decreasing in the tested plants. However, no significant relationship was observed between any of these biochemical indicators and the higher transpiration efficiency (TE) values found in the transgenic events. Our results show that changes in the antioxidative machinery in these transgenic peanut plants (overexpressing the AtDREB1A transcription factor) under water-limiting conditions played no causative role in improved TE.

  10. Transgenic tobacco plants with improved cyanobacterial Rubisco expression but no extra assembly factors grow at near wild-type rates if provided with elevated CO2.

    Science.gov (United States)

    Occhialini, Alessandro; Lin, Myat T; Andralojc, P John; Hanson, Maureen R; Parry, Martin A J

    2016-01-01

    Introducing a carbon-concentrating mechanism and a faster Rubisco enzyme from cyanobacteria into higher plant chloroplasts may improve photosynthetic performance by increasing the rate of CO2 fixation while decreasing losses caused by photorespiration. We previously demonstrated that tobacco plants grow photoautotrophically using Rubisco from Synechococcus elongatus, although the plants exhibited considerably slower growth than wild-type and required supplementary CO2 . Because of concerns that vascular plant assembly factors may not be adequate for assembly of a cyanobacterial Rubisco, prior transgenic plants included the cyanobacterial chaperone RbcX or the carboxysomal protein CcmM35. Here we show that neither RbcX nor CcmM35 is needed for assembly of active cyanobacterial Rubisco. Furthermore, by altering the gene regulatory sequences on the Rubisco transgenes, cyanobacterial Rubisco expression was enhanced and the transgenic plants grew at near wild-type growth rates, although still requiring elevated CO2 . We performed detailed kinetic characterization of the enzymes produced with and without the RbcX and CcmM35 cyanobacterial proteins. These transgenic plants exhibit photosynthetic characteristics that confirm the predicted benefits of introduction of non-native forms of Rubisco with higher carboxylation rate constants in vascular plants and the potential nitrogen-use efficiency that may be achieved provided that adequate CO2 is available near the enzyme. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

  11. Ectopic Expression of Xylella fastidiosa rpfF Conferring Production of Diffusible Signal Factor in Transgenic Tobacco and Citrus Alters Pathogen Behavior and Reduces Disease Severity.

    Science.gov (United States)

    Caserta, R; Souza-Neto, R R; Takita, M A; Lindow, S E; De Souza, A A

    2017-11-01

    The pathogenicity of Xylella fastidiosa is associated with its ability to colonize the xylem of host plants. Expression of genes contributing to xylem colonization are suppressed, while those necessary for insect vector acquisition are increased with increasing concentrations of diffusible signal factor (DSF), whose production is dependent on RpfF. We previously demonstrated that transgenic citrus plants ectopically expressing rpfF from a citrus strain of X. fastidiosa subsp. pauca exhibited less susceptibility to Xanthomonas citri subsp. citri, another pathogen whose virulence is modulated by DSF accumulation. Here, we demonstrate that ectopic expression of rpfF in both transgenic tobacco and sweet orange also confers a reduction in disease severity incited by X. fastidiosa and reduces its colonization of those plants. Decreased disease severity in the transgenic plants was generally associated with increased expression of genes conferring adhesiveness to the pathogen and decreased expression of genes necessary for active motility, accounting for the reduced population sizes achieved in the plants, apparently by limiting pathogen dispersal through the plant. Plant-derived DSF signal molecules in a host plant can, therefore, be exploited to interfere with more than one pathogen whose virulence is controlled by DSF signaling.

  12. Overexpression of herbaceous peony miR156e-3p improves anthocyanin accumulation in transgenic Arabidopsis thaliana lateral branches.

    Science.gov (United States)

    Zhao, Daqiu; Xia, Xing; Wei, Mengran; Sun, Jing; Meng, Jiasong; Tao, Jun

    2017-12-01

    microRNAs (miRNAs) play critical regulatory roles in plant growth and development. In the present study, the function of herbaceous peony ( Paeonia lactiflora Pall.) miR156e-3p in the regulation of color formation has been investigated. Firstly, P. lactiflora miR156e-3p precursor sequence (pre-miR156e-3p) was isolated. Subsequently, the overexpression vector of pre-miR156e-3p was constructed and transformed into Arabidopsis thaliana . Moreover, the medium screening, GUS staining, polymerase chain reaction (PCR) of the GUS region and real-time quantitative PCR (qRT-PCR) of miR156e-3p all confirmed that the purpose gene had been successfully transferred into Arabidopsis plants and expressed, which resulted in apparent purple lateral branches. And this change in color was caused by the improved anthocyanin accumulation. In addition, expression analysis had shown that the level of miR156e-3p transcript was increased, while transcription level of target gene squamosa promoter binding protein-like gene ( SPL1 ), encoding SPL transcription factor that negatively regulated anthocyanin accumulation, was repressed in miR156e-3p-overexpressing transgenic plants, and its downstream gene dihydroflavonol 4-reductase gene ( DFR ) that was directly involved in anthocyanin biosynthesis was strongly expressed, which resulted in anthocyanin accumulation of Arabidopsis lateral branches. These findings would improve the understanding of miRNAs regulation of color formation in P. lactiflora .

  13. Changes in fatty acid content and composition between wild type and CsHMA3 overexpressing Camelina sativa under heavy-metal stress.

    Science.gov (United States)

    Park, Won; Feng, Yufeng; Kim, Hyojin; Suh, Mi Chung; Ahn, Sung-Ju

    2015-09-01

    Under heavy-metal stress, CsHMA3 overexpressing transgenic Camelina plants displayed not only a better quality, but also a higher quantity of unsaturated fatty acids in their seeds compared with wild type. Camelina sativa L. belongs to the Brassicaceae family and is frequently used as a natural vegetable oil source, as its seeds contain a high content of fatty acids. In this study, we observed that, when subjected to heavy metals (Cd, Co, Zn and Pb), the seeds of CsHMA3 (Heavy-Metal P1B-ATPase 3) transgenic lines retained their original golden yellow color and smooth outline, unlike wild-type seeds. Furthermore, we investigated the fatty acids content and composition of wild type and CsHMA3 transgenic lines after heavy metal treatments compared to the control. The results showed higher total fatty acid amounts in seeds of CsHMA3 transgenic lines compared with those in wild-type seeds under heavy-metal stresses. In addition, the compositions of unsaturated fatty acids-especially 18:1 (oleic acid), 18:2 (linoleic acid; only in case of Co treatment), 18:3 (linolenic acid) and 20:1 (eicosenoic acid)-in CsHMA3 overexpressing transgenic lines treated with heavy metals were higher than those of wild-type seeds under the same conditions. Furthermore, reactive oxygen species (ROS) contents in wild-type leaves and roots when treated with heavy metal were higher than in CsHMA3 overexpressing transgenic lines. These results indicate that overexpression of CsHMA3 affects fatty acid composition and content-factors that are responsible for the fuel properties of biodiesel-and can alleviate ROS accumulation caused by heavy-metal stresses in Camelina. Due to these factors, we propose that CsHMA3 transgenic Camelina can be used for phytoremediation of metal-contaminated soil as well as for oil production.

  14. A Novel mouse model of enhanced proteostasis: Full-length human heat shock factor 1 transgenic mice

    International Nuclear Information System (INIS)

    Pierce, Anson; Wei, Rochelle; Halade, Dipti; Yoo, Si-Eun; Ran, Qitao; Richardson, Arlan

    2010-01-01

    Research highlights: → Development of mouse overexpressing native human HSF1 in all tissues including CNS. → HSF1 overexpression enhances heat shock response at whole-animal and cellular level. → HSF1 overexpression protects from polyglutamine toxicity and favors aggresomes. → HSF1 overexpression enhances proteostasis at the whole-animal and cellular level. -- Abstract: The heat shock response (HSR) is controlled by the master transcriptional regulator heat shock factor 1 (HSF1). HSF1 maintains proteostasis and resistance to stress through production of heat shock proteins (HSPs). No transgenic model exists that overexpresses HSF1 in tissues of the central nervous system (CNS). We generated a transgenic mouse overexpressing full-length non-mutant HSF1 and observed a 2-4-fold increase in HSF1 mRNA and protein expression in all tissues studied of HSF1 transgenic (HSF1 +/0 ) mice compared to wild type (WT) littermates, including several regions of the CNS. Basal expression of HSP70 and 90 showed only mild tissue-specific changes; however, in response to forced exercise, the skeletal muscle HSR was more elevated in HSF1 +/0 mice compared to WT littermates and in fibroblasts following heat shock, as indicated by levels of inducible HSP70 mRNA and protein. HSF1 +/0 cells elicited a significantly more robust HSR in response to expression of the 82 repeat polyglutamine-YFP fusion construct (Q82YFP) and maintained proteasome-dependent processing of Q82YFP compared to WT fibroblasts. Overexpression of HSF1 was associated with fewer, but larger Q82YFP aggregates resembling aggresomes in HSF1 +/0 cells, and increased viability. Therefore, our data demonstrate that tissues and cells from mice overexpressing full-length non-mutant HSF1 exhibit enhanced proteostasis.

  15. Inhibition of G1-phase arrest induced by ionizing radiation in hematopoietic cells by overexpression of genes involved in the G1/S-phase transition

    International Nuclear Information System (INIS)

    Epperly, M.; Berry, L.; Halloran, A.; Greenberger, J.S.

    1995-01-01

    D-type cyclins and cyclin-dependent kinase (cdk-4) are likely involved in regulating passage of cells through the G 1 phase of the cell cycle. A decrease in the proportion of cells in G 1 , a relatively radiation-sensitive phase of the cell cycle, should result in increased resistance to ionizing radiation; however, the effect of such overexpression on X-ray-induced G 1 -phase arrest is not known. Radiation survival curves were obtained at a dose rate of either 8 cGy/min or 1 Gy/min for subclones of the IL-3-dependent hematopoietic progenitor cell line 32D cl 3 expressing transgenes for either cyclin-D1, D2 or D3 or cdk-4. We compared the results to those with overexpression of the transgene for Bcl-2, whose expression enhances radiation survival and delays apoptosis. Cells overexpressing transgenes for each D-type cyclin or Bcl-2 had an increased number of cells in S phase compared to parent line 32D cl 3; however, overexpression of cdk-4 had no effect on cell cycle distribution. Cell death resulting from withdrawal of IL-3 was not affected by overexpression of D2, cdk-4 or Bcl-2. Flow cytometry 24 h after 5 Gy irradiation demonstrated that overexpression of each G 1 -phase regulatory transgene decreased the proportion of cells at the G 1 /S-phase border. Western analysis revealed induction of cyclin-D protein levels by irradiation, but no change in the D O , but a significant increase in the rvec n for cyclin-D or cdk-4 transgene-overexpressing clones at 1 Gy/min (P 1 /S-phase arrest. 31 refs., 4 figs., 4 tabs

  16. Bean Yellow Dwarf Virus replicons for high-level transgene expression in transgenic plants and cell cultures.

    Science.gov (United States)

    Zhang, Xiuren; Mason, Hugh

    2006-02-05

    A novel stable transgenic plant expression system was developed using elements of the replication machinery of Bean Yellow Dwarf Virus (BeYDV). The system contains two transgenes: 1) The BeYDV replicon vector with an expression cassette flanked by cis-acting DNA elements of BeYDV, and 2) The viral replication initiator protein (Rep) controlled by an alcohol-inducible promoter. When Rep expression was triggered by treatment with ethanol, it induced release of the BeYDV replicon from stably integrated T-DNA and episomal replication to high copy number. Replicon amplification resulted in substantially increased transgene mRNA levels (up to 80-fold) and translation products (up to 10-fold) after induction of Rep expression by ethanol treatment in tobacco NT1 cells and leaves of whole potato plants. Thus, the BeYDV stable transformant replicon system is a powerful tool for plant-based production of recombinant proteins. (c) 2005 Wiley Periodicals, Inc.

  17. Overexpression of Poplar PtrWRKY89 in Transgenic Arabidopsis Leads to a Reduction of Disease Resistance by Regulating Defense-Related Genes in Salicylate- and Jasmonate-Dependent Signaling.

    Directory of Open Access Journals (Sweden)

    Yuanzhong Jiang

    Full Text Available The plant hormones jasmonic acid (JA and salicylic acid (SA play key roles in plant defenses against pathogens and several WRKY transcription factors have been shown to have a role in SA/JA crosstalk. In a previous study, overexpression of the poplar WRKY gene PtrWRKY89 enhanced resistance to pathogens in transgenic poplars. In this study, the promoter of PtrWRKY89 (ProPtrWRKY89 was isolated and used to drive GUS reporter gene. High GUS activity was observed in old leaves of transgenic Arabidopsis containing ProPtrWRKY89-GUS construct and GUS expression was extremely induced by SA solution and SA+MeJA mixture but not by MeJA treatment. Subcellular localization and transactivation assays showed that PtrWRKY89 acted as a transcription activator in the nucleus. Constitutive expression of PtrWRKY89 in Arabidopsis resulted in more susceptible to Pseudomonas syringae and Botrytis cinerea compared to wild-type plants. Quantitative real-time PCR (qRT-PCR analysis confirmed that marker genes of SA and JA pathways were down-regulated in transgenic Arabidopsis after pathogen inoculations. Overall, our results indicated that PtrWRKY89 modulates a cross talk in resistance to P. syringe and B. cinerea by negatively regulating both SA and JA pathways in Arabidopsis.

  18. Overexpression of Poplar PtrWRKY89 in Transgenic Arabidopsis Leads to a Reduction of Disease Resistance by Regulating Defense-Related Genes in Salicylate- and Jasmonate-Dependent Signaling.

    Science.gov (United States)

    Jiang, Yuanzhong; Guo, Li; Liu, Rui; Jiao, Bo; Zhao, Xin; Ling, Zhengyi; Luo, Keming

    2016-01-01

    The plant hormones jasmonic acid (JA) and salicylic acid (SA) play key roles in plant defenses against pathogens and several WRKY transcription factors have been shown to have a role in SA/JA crosstalk. In a previous study, overexpression of the poplar WRKY gene PtrWRKY89 enhanced resistance to pathogens in transgenic poplars. In this study, the promoter of PtrWRKY89 (ProPtrWRKY89) was isolated and used to drive GUS reporter gene. High GUS activity was observed in old leaves of transgenic Arabidopsis containing ProPtrWRKY89-GUS construct and GUS expression was extremely induced by SA solution and SA+MeJA mixture but not by MeJA treatment. Subcellular localization and transactivation assays showed that PtrWRKY89 acted as a transcription activator in the nucleus. Constitutive expression of PtrWRKY89 in Arabidopsis resulted in more susceptible to Pseudomonas syringae and Botrytis cinerea compared to wild-type plants. Quantitative real-time PCR (qRT-PCR) analysis confirmed that marker genes of SA and JA pathways were down-regulated in transgenic Arabidopsis after pathogen inoculations. Overall, our results indicated that PtrWRKY89 modulates a cross talk in resistance to P. syringe and B. cinerea by negatively regulating both SA and JA pathways in Arabidopsis.

  19. Expression Patterns of OsPIL11, a Phytochrome-Interacting Factor in Rice, and Preliminary Analysis of Its Roles in Light Signal Transduction

    Directory of Open Access Journals (Sweden)

    Li LI

    2012-12-01

    Full Text Available The expression patterns of OsPIL11, one of six putative phytochrome-interacting factors, were analyzed in different organs of transgenic tobacco (Nicotiana tabacum. The expression of OsPIL11 was organ-specific and was regulated by leaf development, abscisic acid (ABA, jasmonic acid (JA and salicylic acid (SA. To further explore the role of OsPIL11 in plant light signal transduction, a plant expression vector of OsPIL11 was constructed and introduced into tobacco. When grown under continuous red light, OsPIL11-overexpressed transgenic tobacco exhibited shorter hypocotyls and larger cotyledons and leaves compared to wild-type seedlings. When grown under continuous far-red light, however, transgenic and wild-type seedlings showed similar phenotypes. These results indicate that OsPIL11 is involved in red light induced de-etiolation, but not in far-red light induced de-etiolation in transgenic tobacco, which lays the foundation for dissecting the function of OsPIL11 in phytochrome-mediated light signal transduction in rice.

  20. Over-expression of Trxo1 increases the viability of tobacco BY-2 cells under H2O2 treatment.

    Science.gov (United States)

    Ortiz-Espín, Ana; Locato, Vittoria; Camejo, Daymi; Schiermeyer, Andreas; De Gara, Laura; Sevilla, Francisca; Jiménez, Ana

    2015-09-01

    Reactive oxygen species (ROS), especially hydrogen peroxide, play a critical role in the regulation of plant development and in the induction of plant defence responses during stress adaptation, as well as in plant cell death. The antioxidant system is responsible for controlling ROS levels in these processes but redox homeostasis is also a key factor in plant cell metabolism under normal and stress situations. Thioredoxins (Trxs) are ubiquitous small proteins found in different cell compartments, including mitochondria and nuclei (Trxo1), and are involved in the regulation of target proteins through reduction of disulphide bonds, although their role under oxidative stress has been less well studied. This study describes over-expression of a Trxo1 for the first time, using a cell-culture model subjected to an oxidative treatment provoked by H2O2. Control and over-expressing PsTrxo1 tobacco (Nicotiana tabacum) BY-2 cells were treated with 35 mm H2O2 and the effects were analysed by studying the growth dynamics of the cultures together with oxidative stress parameters, as well as several components of the antioxidant systems involved in the metabolism of H2O2. Analysis of different hallmarks of programmed cell death was also carried out. Over-expression of PsTrxo1 caused significant differences in the response of TBY-2 cells to high concentrations of H2O2, namely higher and maintained viability in over-expressing cells, whilst the control line presented a severe decrease in viability and marked indications of oxidative stress, with generalized cell death after 3 d of treatment. In over-expressing cells, an increase in catalase activity, decreases in H2O2 and nitric oxide contents and maintenance of the glutathione redox state were observed. A decreased content of endogenous H2O2 may be responsible in part for the delayed cell death found in over-expressing cells, in which changes in oxidative parameters and antioxidants were less extended after the oxidative

  1. Generation of bi-transgenic pigs overexpressing human lactoferrin and lysozyme in milk.

    Science.gov (United States)

    Cui, Dan; Li, Jia; Zhang, Linlin; Liu, Shen; Wen, Xiao; Li, Qiuyan; Zhao, Yaofeng; Hu, Xiaoxiang; Zhang, Ran; Li, Ning

    2015-04-01

    Intensive swine production industry uses antibiotics to treat diseases and improve pig growth. This can not only cause antibiotic resistance, but can also pollute the environment or eventually affect human public health. To date, human lactoferrin (hLF) and human lysozyme (hLZ) have been known as non-adaptive but interactive antimicrobial members and could act in concert against bacteria, which contribute to host defense. Therefore, their expression in pigs might be an alternative strategy for replacing antibiotics in the pig production industry. In our study, we produced hLF and hLZ bi-transgenic pigs and assessed the milk's antibacterial ability. Integration of both transgenes was confirmed by PCR and southern blot. Both the hLF and hLZ were expressed in the mammary gland of bi-transgenic pigs, as detected by western blotting. The expression amounts were 6.5 g/L for hLF and 1.1 mg/L for hLZ using ELISA. Interestingly, pig milk containing hLF and hLZ had synergistic antimicrobial activity. Our results suggest an alternative approach for avoiding the use of antibiotics in the pig industry, which would be of great benefit to the commercial swine production.

  2. Constitutive expression of a fungus-inducible carboxylesterase improves disease resistance in transgenic pepper plants.

    Science.gov (United States)

    Ko, Moonkyung; Cho, Jung Hyun; Seo, Hyo-Hyoun; Lee, Hyun-Hwa; Kang, Ha-Young; Nguyen, Thai Son; Soh, Hyun Cheol; Kim, Young Soon; Kim, Jeong-Il

    2016-08-01

    Resistance against anthracnose fungi was enhanced in transgenic pepper plants that accumulated high levels of a carboxylesterase, PepEST in anthracnose-susceptible fruits, with a concurrent induction of antioxidant enzymes and SA-dependent PR proteins. A pepper esterase gene (PepEST) is highly expressed during the incompatible interaction between ripe fruits of pepper (Capsicum annuum L.) and a hemibiotrophic anthracnose fungus (Colletotrichum gloeosporioides). In this study, we found that exogenous application of recombinant PepEST protein on the surface of the unripe pepper fruits led to a potentiated state for disease resistance in the fruits, including generation of hydrogen peroxide and expression of pathogenesis-related (PR) genes that encode mostly small proteins with antimicrobial activity. To elucidate the role of PepEST in plant defense, we further developed transgenic pepper plants overexpressing PepEST under the control of CaMV 35S promoter. Molecular analysis confirmed the establishment of three independent transgenic lines carrying single copy of transgenes. The level of PepEST protein was estimated to be approximately 0.002 % of total soluble protein in transgenic fruits. In response to the anthracnose fungus, the transgenic fruits displayed higher expression of PR genes, PR3, PR5, PR10, and PepThi, than non-transgenic control fruits did. Moreover, immunolocalization results showed concurrent localization of ascorbate peroxidase (APX) and PR3 proteins, along with the PepEST protein, in the infected region of transgenic fruits. Disease rate analysis revealed significantly low occurrence of anthracnose disease in the transgenic fruits, approximately 30 % of that in non-transgenic fruits. Furthermore, the transgenic plants also exhibited resistance against C. acutatum and C. coccodes. Collectively, our results suggest that overexpression of PepEST in pepper confers enhanced resistance against the anthracnose fungi by activating the defense signaling

  3. Embryo-specific expression of soybean oleosin altered oil body morphogenesis and increased lipid content in transgenic rice seeds.

    Science.gov (United States)

    Liu, Wen Xian; Liu, Hua Liang; Qu, Le Qing

    2013-09-01

    Oleosin is the most abundant protein in the oil bodies of plant seeds, playing an important role in regulating oil body formation and lipid accumulation. To investigate whether lipid accumulation in transgenic rice seeds depends on the expression level of oleosin, we introduced two soybean oleosin genes encoding 24 kDa proteins into rice under the control of an embryo-specific rice promoter REG-2. Overexpression of soybean oleosin in transgenic rice leads to an increase of seed lipid content up to 36.93 and 46.06 % higher than that of the non-transgenic control, respectively, while the overall fatty acid profiles of triacylglycerols remained unchanged. The overexpression of soybean oleosin in transgenic rice seeds resulted in more numerous and smaller oil bodies compared with wild type, suggesting that an inverse relationship exists between oil body size and the total oleosin level. The increase in lipid content is accompanied by a reduction in the accumulation of total seed protein. Our results suggest that it is possible to increase rice seed oil content for food use and for use as a low-cost feedstock for biodiesel by overexpressing oleosin in rice seeds.

  4. A method for the production and expedient screening of CRISPR/Cas9-mediated non-transgenic mutant plants.

    Science.gov (United States)

    Chen, Longzheng; Li, Wei; Katin-Grazzini, Lorenzo; Ding, Jing; Gu, Xianbin; Li, Yanjun; Gu, Tingting; Wang, Ren; Lin, Xinchun; Deng, Ziniu; McAvoy, Richard J; Gmitter, Frederick G; Deng, Zhanao; Zhao, Yunde; Li, Yi

    2018-01-01

    Developing CRISPR/Cas9-mediated non-transgenic mutants in asexually propagated perennial crop plants is challenging but highly desirable. Here, we report a highly useful method using an Agrobacterium -mediated transient CRISPR/Cas9 gene expression system to create non-transgenic mutant plants without the need for sexual segregation. We have also developed a rapid, cost-effective, and high-throughput mutant screening protocol based on Illumina sequencing followed by high-resolution melting (HRM) analysis. Using tetraploid tobacco as a model species and the phytoene desaturase ( PDS ) gene as a target, we successfully created and expediently identified mutant plants, which were verified as tetra-allelic mutants. We produced pds mutant shoots at a rate of 47.5% from tobacco leaf explants, without the use of antibiotic selection. Among these pds plants, 17.2% were confirmed to be non-transgenic, for an overall non-transgenic mutation rate of 8.2%. Our method is reliable and effective in creating non-transgenic mutant plants without the need to segregate out transgenes through sexual reproduction. This method should be applicable to many economically important, heterozygous, perennial crop species that are more difficult to regenerate.

  5. Pancreatic beta-cell lipotoxicity induced by overexpression of hormone-sensitive lipase

    DEFF Research Database (Denmark)

    Winzell, Maria Sörhede; Svensson, Håkan; Enerbäck, Sven

    2003-01-01

    Lipid perturbations associated with triglyceride overstorage in beta-cells impair insulin secretion, a process termed lipotoxicity. To assess the role of hormone-sensitive lipase, which is expressed and enzymatically active in beta-cells, in the development of lipotoxicity, we generated transgenic...... mice overexpressing hormone-sensitive lipase specifically in beta-cells. Transgenic mice developed glucose intolerance and severely blunted glucose-stimulated insulin secretion when challenged with a high-fat diet. As expected, both lipase activity and forskolin-stimulated lipolysis was increased...

  6. Knock-out and Transgenic Strategies to Improve Neural Transplantation Therapy for Parkinson's Disease

    National Research Council Canada - National Science Library

    Isacson, Ole

    2000-01-01

    .... In our second objective of enhancing axonal growth leading to optimal functional recovery by neuronal transplants, we employed transgenic bcl-2 overexpressing donor cells and similar molecules...

  7. Overexpression of the ped/pea-15 Gene Causes Diabetes by Impairing Glucose-Stimulated Insulin Secretion in Addition to Insulin Action

    OpenAIRE

    Vigliotta, Giovanni; Miele, Claudia; Santopietro, Stefania; Portella, Giuseppe; Perfetti, Anna; Maitan, Maria Alessandra; Cassese, Angela; Oriente, Francesco; Trencia, Alessandra; Fiory, Francesca; Romano, Chiara; Tiveron, Cecilia; Tatangelo, Laura; Troncone, Giancarlo; Formisano, Pietro

    2004-01-01

    Overexpression of the ped/pea-15 gene is a common feature of type 2 diabetes. In the present work, we show that transgenic mice ubiquitously overexpressing ped/pea-15 exhibited mildly elevated random-fed blood glucose levels and decreased glucose tolerance. Treatment with a 60% fat diet led ped/pea-15 transgenic mice to develop diabetes. Consistent with insulin resistance in these mice, insulin administration reduced glucose levels by only 35% after 45 min, compared to 70% in control mice. In...

  8. Overexpression of a 9-cis-epoxycarotenoid dioxygenase gene in Nicotiana plumbaginifolia increases abscisic acid and phaseic acid levels and enhances drought tolerance.

    Science.gov (United States)

    Qin, Xiaoqiong; Zeevaart, Jan A D

    2002-02-01

    The plant hormone abscisic acid (ABA) plays important roles in seed maturation and dormancy and in adaptation to a variety of environmental stresses. An effort to engineer plants with elevated ABA levels and subsequent stress tolerance is focused on the genetic manipulation of the cleavage reaction. It has been shown in bean (Phaseolus vulgaris) that the gene encoding the cleavage enzyme (PvNCED1) is up-regulated by water stress, preceding accumulation of ABA. Transgenic wild tobacco (Nicotiana plumbaginifolia Viv.) plants were produced that overexpress the PvNCED1 gene either constitutively or in an inducible manner. The constitutive expression of PvNCED1 resulted in an increase in ABA and its catabolite, phaseic acid (PA). When the PvNCED1 gene was driven by the dexamethasone (DEX)-inducible promoter, a transient induction of PvNCED1 message and accumulation of ABA and PA were observed in different lines after application of DEX. Accumulation of ABA started to level off after 6 h, whereas the PA level continued to increase. In the presence of DEX, seeds from homozygous transgenic line TN1 showed a 4-d delay in germination. After spraying with DEX, the detached leaves from line TN1 had a drastic decrease in their water loss relative to control leaves. These plants also showed a marked increase in their tolerance to drought stress. These results indicate that it is possible to manipulate ABA levels in plants by overexpressing the key regulatory gene in ABA biosynthesis and that stress tolerance can be improved by increasing ABA levels.

  9. Overexpression of TiERF1 enhances resistance to sharp eyespot in transgenic wheat

    OpenAIRE

    Chen, Liang; Zhang, ZengYan; Liang, HongXia; Liu, HongXia; Du, LiPu; Xu, Huijun; Xin, Zhiyong

    2008-01-01

    Wheat sharp eyespot, primarily caused by a soil-borne fungus Rhizoctonia cerealis, has become one of the most serious diseases of wheat in China. In this study, an ethylene response factor (ERF) gene from a wheat relative Thinopyrum intermedium, TiERF1, was characterized further, transgenic wheat lines expressing TiERF1 were developed, and the resistance of the transgenic wheat lines against R. cerealis was investigated. Southern blotting analysis indicated that at least two copies of the TiE...

  10. Knock-Out and Transgenic Strategies to Improve Neural Transplantation Therapy for Parkinson's Disease

    National Research Council Canada - National Science Library

    Isacson, Ole

    2001-01-01

    .... To enhance axonal growth leading to optimal functional recovery by neuronal transplants, we employed transgenic bcl-2 overexpressing donor cells and similar molecules influencing the growth of axons...

  11. Regulatory dephosphorylation of CDK at G₂/M in plants: yeast mitotic phosphatase cdc25 induces cytokinin-like effects in transgenic tobacco morphogenesis.

    Science.gov (United States)

    Lipavská, Helena; Masková, Petra; Vojvodová, Petra

    2011-05-01

    During the last three decades, the cell cycle and its control by cyclin-dependent kinases (CDKs) have been extensively studied in eukaryotes. This endeavour has produced an overall picture that basic mechanisms seem to be largely conserved among all eukaryotes. The intricate regulation of CDK activities includes, among others, CDK activation by CDC25 phosphatase at G₂/M. In plants, however, studies of this regulation have lagged behind as a plant Cdc25 homologue or other unrelated phosphatase active at G₂/M have not yet been identified. Failure to identify a plant mitotic CDK activatory phosphatase led to characterization of the effects of alien cdc25 gene expression in plants. Tobacco, expressing the Schizosaccharomyces pombe mitotic activator gene, Spcdc25, exhibited morphological, developmental and biochemical changes when compared with wild type (WT) and, importantly, increased CDK dephosphorylation at G₂/M. Besides changes in leaf shape, internode length and root development, in day-neutral tobacco there was dramatically earlier onset of flowering with a disturbed acropetal floral capacity gradient typical of WT. In vitro, de novo organ formation revealed substantially earlier and more abundant formation of shoot primordia on Spcdc25 tobacco stem segments grown on shoot-inducing media when compared with WT. Moreover, in contrast to WT, stem segments from transgenic plants formed shoots even without application of exogenous growth regulator. Spcdc25-expressing BY-2 cells exhibited a reduced mitotic cell size due to a shortening of the G₂ phase together with high activity of cyclin-dependent kinase, NtCDKB1, in early S-phase, S/G₂ and early M-phase. Spcdc25-expressing tobacco ('Samsun') cell suspension cultures showed a clustered, more circular, cell phenotype compared with chains of elongated WT cells, and increased content of starch and soluble sugars. Taken together, Spcdc25 expression had cytokinin-like effects on the characteristics studied

  12. Overexpression of a 9-cis-Epoxycarotenoid Dioxygenase Gene in Nicotiana plumbaginifolia Increases Abscisic Acid and Phaseic Acid Levels and Enhances Drought Tolerance1

    Science.gov (United States)

    Qin, Xiaoqiong; Zeevaart, Jan A.D.

    2002-01-01

    The plant hormone abscisic acid (ABA) plays important roles in seed maturation and dormancy and in adaptation to a variety of environmental stresses. An effort to engineer plants with elevated ABA levels and subsequent stress tolerance is focused on the genetic manipulation of the cleavage reaction. It has been shown in bean (Phaseolus vulgaris) that the gene encoding the cleavage enzyme (PvNCED1) is up-regulated by water stress, preceding accumulation of ABA. Transgenic wild tobacco (Nicotiana plumbaginifolia Viv.) plants were produced that overexpress the PvNCED1 gene either constitutively or in an inducible manner. The constitutive expression of PvNCED1 resulted in an increase in ABA and its catabolite, phaseic acid (PA). When the PvNCED1 gene was driven by the dexamethasone (DEX)-inducible promoter, a transient induction of PvNCED1 message and accumulation of ABA and PA were observed in different lines after application of DEX. Accumulation of ABA started to level off after 6 h, whereas the PA level continued to increase. In the presence of DEX, seeds from homozygous transgenic line TN1 showed a 4-d delay in germination. After spraying with DEX, the detached leaves from line TN1 had a drastic decrease in their water loss relative to control leaves. These plants also showed a marked increase in their tolerance to drought stress. These results indicate that it is possible to manipulate ABA levels in plants by overexpressing the key regulatory gene in ABA biosynthesis and that stress tolerance can be improved by increasing ABA levels. PMID:11842158

  13. Production of soybean isoflavone genistein in non-legume plants via genetically modified secondary metabolism pathway.

    Science.gov (United States)

    Liu, Rongrong; Hu, Yuanlei; Li, Jialin; Lin, Zhongping

    2007-01-01

    Genetic modification of secondary metabolic pathways to produce desirable natural products is an attractive approach in plant biotechnology. In our study, we attempted to produce a typical soybean isoflavone genistein, a well-known health-promoting metabolite, in non-legume plants via genetic engineering. Both overexpression and antisense suppression strategies were used to manipulate the expression of several genes encoding key enzymes in the flavonoids/isoflavonoids pathway in transgenic tobacco, lettuce, and petunia. Introducing soybean isoflavone synthase (IFS) into these plants, which naturally do not produce isoflavonoids due to a lack of this leguminous enzyme, resulted in genistein biosynthesis in tobacco petals, petunia leaves and petals, and lettuce leaves. In tobacco, when flavanone 3-hydroxylase (F3H) expression was suppressed by its antisense gene while soybean IFS was overexpressed at the same time, genistein yield increased prominently. In addition, overexpression of phenylalanine ammonia-lyase (PAL) also led to an enhanced genistein production in tobacco petals and lettuce leaves in the presence of IFS than in the plants that overexpressed only IFS.

  14. Evaluating water deficit and glyphosate treatment on the accumulation of phenolic compounds and photosynthesis rate in transgenic Codonopsis lanceolata (Siebold & Zucc.) Trautv. over-expressing γ-tocopherol methyltransferase (γ-tmt) gene.

    Science.gov (United States)

    Ghimire, Bimal Kumar; Son, Na-Young; Kim, Seung-Hyun; Yu, Chang Yeon; Chung, Ill-Min

    2017-07-01

    The effect of water stress and herbicide treatment on the phenolic compound concentration and photosynthesis rate in transgenic Codonopsis lanceolata plants over-expressing the γ-tmt gene was investigated and compared to that in control non-transgenic C. lanceolata plants. The total phenolic compound content was investigated using high-performance liquid chromatography combined with diode array detection in C. lanceolata seedlings 3 weeks after water stress and treatment with glyphosate. Changes in the composition of phenolic compounds were observed in leaf and root extracts from transformed C. lanceolata plants following water stress and treatment with glyphosate. The total concentration of phenolic compounds in the leaf extracts of transgenic samples after water stress ranged from 3455.13 ± 40.48 to 8695.00 ± 45.44 µg g -1 dry weight (DW), whereas the total concentration phenolic compound in the leaf extracts of non-transgenic control samples was 5630.83 ± 45.91 µg g -1  DW. The predominant phenolic compounds that increased after the water stress in the transgenic leaf were (+) catechin, benzoic acid, chlorogenic acid, ferulic acid, gallic acid, rutin, vanillic acid, and veratric acid. The total concentration of phenolic compounds in the leaf extracts of transgenic samples after glyphosate treatment ranged from 4744.37 ± 81.81 to 12,051.02 ± 75.00 µg g -1 DW, whereas the total concentration of the leaf extracts of non-transgenic control samples after glyphosate treatment was 3778.28 ± 59.73 µg g -1 DW. Major phenolic compounds that increased in the transgenic C. lanceolata plants after glyphosate treatment included kaempherol, gallic acid, myricetin, p-hydroxybenzjoic acid, quercetin, salicylic acid, t-cinnamic acid, catechin, benzoicacid, ferulic acid, protocatechuic acid, veratric acid, and vanillic acid. Among these, vanillic acid showed the greatest increase in both leaf and root extracts from transgenic plants relative to

  15. L-type calcium channel CaV 1.2 in transgenic mice overexpressing human AbetaPP751 with the London (V717I) and Swedish (K670M/N671L) mutations.

    Science.gov (United States)

    Willis, Michael; Kaufmann, Walter A; Wietzorrek, Georg; Hutter-Paier, Birgit; Moosmang, Sven; Humpel, Christian; Hofmann, Franz; Windisch, Manfred; Knaus, Hans-Günther; Marksteiner, Josef

    2010-01-01

    Cumulative evidence indicates that amyloid-beta peptides exert some of their neurodegenerative effects through modulation of L-type voltage gated calcium channels, which play key roles in a diverse range of CNS functions. In this study we examined the expression of CaV1.2 L-type voltage gated calcium channels in transgenic mice overexpressing human AbetaPP751 with the London (V717I) and Swedish (K670M/N671L) mutations by immunohistochemistry in light and electron microscopy. In hippocampal layers of wild type and transgenic mice, CaV1.2 channels were predominantly localized to somato-dendritic domains of neurons, and to astrocytic profiles with an age-dependent increase in labeling density. In transgenic animals, CaV1.2-like immunoreactive clusters were found in neuronal profiles in association with amyloid-beta plaques. Both the number and density of these clusters depended upon age of animals and number of plaques. The most striking difference between wild type and transgenic mice was the age-dependent expression of CaV1.2 channels in reactive astrocytes. At the age of 6 month, CaV1.2 channels were rarely detected in reactive astrocytes of transgenic mice, but an incremental number of CaV1.2 expressing reactive astrocytes was found with increasing age of animals and number of amyloid-beta plaques. This study demonstrates that CaV1.2 channels are highly expressed in reactive astrocytes of 12-months of age transgenic mice, which might be a consequence of the increasing amyloid burden. Further studies should clarify which functional implications are associated with the higher availability of CaV1.2 channels in late stage Alzheimer's disease.

  16. The aux1 gene of the Ri plasmid is sufficient to confer auxin autotrophy in tobacco BY-2 cells.

    Science.gov (United States)

    Nemoto, Keiichirou; Hara, Masamitsu; Goto, Shingo; Kasai, Kouji; Seki, Hikaru; Suzuki, Masashi; Oka, Atsuhiro; Muranaka, Toshiya; Mano, Yoshihiro

    2009-05-01

    Tobacco (Nicotiana tabacum) Bright Yellow-2 (BY-2) cells are rapidly proliferating meristematic cells that require auxin for culture in vitro. We have established several transgenic BY-2 cell lines that carry the T-DNA of Agrobacterium rhizogenes 15834, which harbors an agropine-type root-inducing (Ri) plasmid. Two of these lines, BYHR-3 and BYHR-7, were used to test the role of auxin in the proliferation of plant cells. The lines grew rapidly in Linsmaier-Skoog (LS) medium lacking auxin and other phytohormones. The TR-DNA, containing the aux1 (tryptophan monooxygenase) and aux2 (indoleacetamide hydrolase) genes, was present in the genomes of both transgenic lines, whereas the TL-DNA, containing the rolA, B, C and D genes, was present in the genome of BYHR-7 but not BYHR-3. Since the introduction of the rolABCD genes alone did not affect the auxin requirement of BY-2 cells, the aux1 and aux2 genes, but not the rolABCD genes, appear to be relevant to the auxin autotrophy of these transgenic lines. Furthermore, the overexpression of aux1 allowed BY-2 cells to grow rapidly in the absence of auxin, suggesting the existence in plant cells of an unidentified gene whose product is functionally equivalent or similar to that of aux2 of the Ri plasmid.

  17. Targeted Modification of Homogalacturonan by Transgenic Expression of a Fungal Polygalacturonase Alters Plant Growth1

    Science.gov (United States)

    Capodicasa, Cristina; Vairo, Donatella; Zabotina, Olga; McCartney, Lesley; Caprari, Claudio; Mattei, Benedetta; Manfredini, Cinzia; Aracri, Benedetto; Benen, Jacques; Knox, J. Paul; De Lorenzo, Giulia; Cervone, Felice

    2004-01-01

    Pectins are a highly complex family of cell wall polysaccharides comprised of homogalacturonan (HGA), rhamnogalacturonan I and rhamnogalacturonan II. We have specifically modified HGA in both tobacco (Nicotiana tabacum) and Arabidopsis by expressing the endopolygalacturonase II of Aspergillus niger (AnPGII). Cell walls of transgenic tobacco plants showed a 25% reduction in GalUA content as compared with the wild type and a reduced content of deesterified HGA as detected by antibody labeling. Neutral sugars remained unchanged apart from a slight increase of Rha, Ara, and Gal. Both transgenic tobacco and Arabidopsis were dwarfed, indicating that unesterified HGA is a critical factor for plant cell growth. The dwarf phenotypes were associated with AnPGII activity as demonstrated by the observation that the mutant phenotype of tobacco was completely reverted by crossing the dwarfed plants with plants expressing PGIP2, a strong inhibitor of AnPGII. The mutant phenotype in Arabidopsis did not appear when transformation was performed with a gene encoding AnPGII inactivated by site directed mutagenesis. PMID:15247378

  18. Global Overexpression of ET-1 Decreases Blood Pressure - A Systematic Review and Meta-Analysis of ET-1 Transgenic Mice.

    Science.gov (United States)

    Lu, Yong Ping; Tsuprykov, Oleg; Vignon-Zellweger, Nicolas; Heiden, Susi; Hocher, Berthold

    2016-01-01

    ET-1 has independent effects on blood pressure regulation in vivo, it is involved in tubular water and salt excretion, promotes constriction of smooth muscle cells, modulates sympathetic nerve activity, and activates the liberation of nitric oxide. To determine the net effect of these partially counteracting mechanisms on blood pressure, a systematic meta-analysis was performed. Based on the principles of Cochrane systematic reviews, we searched in major literature databases - MEDLINE (PubMed), Embase, Google Scholar, and the China Biological Medicine Database (CBM-disc) - for articles relevant to the topic of the blood pressure phenotype of endothelin-1 transgenic (ET-1+/+) mice from January 1, 1988 to March 31, 2016. Review Manager Version 5.0 (Rev-Man 5.0) software was applied for statistical analysis. In total thirteen studies reported blood pressure data. The meta-analysis of blood pressure data showed that homozygous ET-1 transgenic mice (ET-1+/+ mice) had a significantly lower blood pressure as compared to WT mice (mean difference: -2.57 mmHg, 95% CI: -4.98∼ -0.16, P = 0.04), with minimal heterogeneity (P = 0.86). A subgroup analysis of mice older than 6 months revealed that the blood pressure difference between ET-1+/+ mice and WT mice was even more pronounced (mean difference: -6.19 mmHg, 95% CI: -10.76∼ -1.62, P = 0.008), with minimal heterogeneity (P = 0.91). This meta-analysis provides robust evidence that global ET-1 overexpression in mice lowers blood pressure in an age-dependent manner. Older ET-1+/+ mice have a somewhat more pronounced reduction of blood pressure. © 2016 The Author(s) Published by S. Karger AG, Basel.

  19. OVER-EXPRESSION OF GENE ENCODING FATTY ACID METABOLIC ENZYMES IN FISH

    Directory of Open Access Journals (Sweden)

    Alimuddin Alimuddin

    2008-12-01

    Full Text Available Eicosapentaenoic acid (EPA, 20:5n-3 and docosahexaenoic acid (DHA, 22:6n-3 have important nutritional benefits in humans. EPA and DHA are mainly derived from fish, but the decline in the stocks of major marine capture fishes could result in these fatty acids being consumed less. Farmed fish could serve as promising sources of EPA and DHA, but they need these fatty acids in their diets. Generation of fish strains that are capable of synthesizing enough amounts of EPA/DHA from the conversion of α-linolenic acid (LNA, 18:3n-3 rich oils can supply a new EPA/DHA source. This may be achieved by over-expression of genes encoding enzymes involved in HUFA biosynthesis. In aquaculture, the successful of this technique would open the possibility to reduce the enrichment of live food with fish oils for marine fish larvae, and to completely substitute fish oils with plant oils without reducing the quality of flesh in terms of EPA and DHA contents. Here, three genes, i.e. Δ6-desaturase-like (OmΔ6FAD, Δ5-desaturase-like (OmΔ5FAD and elongase-like (MELO encoding EPA/DHA metabolic enzymes derived from masu salmon (Oncorhynchus masou were individually transferred into zebrafish (Danio rerio as a model to increase its ability for synthesizing EPA and DHA. Fatty acid analysis showed that EPA content in whole body of the second transgenic fish generation over-expressing OmΔ6FAD gene was 1.4 fold and that of DHA was 2.1 fold higher (P<0.05 than those in non-transgenic fish. The EPA content in whole body of transgenic fish over-expressing OmΔ5FAD gene was 1.21-fold, and that of DHA was 1.24-fold higher (P<0.05 than those in nontransgenic fish. The same patterns were obtained in transgenic fish over-expressing MELO gene. EPA content was increased by 1.30-fold and DHA content by 1.33-fold higher (P<0.05 than those in non-transgenic fish. The results of studies demonstrated that fatty acid content of fish can be enhanced by over-expressing

  20. Overexpression of hepatoma-derived growth factor in melanocytes does not lead to oncogenic transformation

    International Nuclear Information System (INIS)

    Sedlmaier, Angela; Wernert, Nicolas; Gallitzendörfer, Rainer; Abouzied, Mekky M; Gieselmann, Volkmar; Franken, Sebastian

    2011-01-01

    HDGF is a growth factor which is overexpressed in a wide range of tumors. Importantly, expression levels were identified as a prognostic marker in some types of cancer such as melanoma. To investigate the presumed oncogenic/transforming capacity of HDGF, we generated transgenic mice overexpressing HDGF in melanocytes. These mice were bred with mice heterozygous for a defective copy of the Ink4a tumor suppressor gene and were exposed to UV light to increase the risk for tumor development both genetically and physiochemically. Mice were analyzed by immunohistochemistry and Western blotting. Furthermore, primary melanocytes were isolated from different strains created. Transgenic animals overexpressed HDGF in hair follicle melanocytes. Interestingly, primary melanocytes isolated from transgenic animals were not able to differentiate in vitro whereas cells isolated from wild type and HDGF-deficient animals were. Although, HDGF -/- /Ink4a +/- mice displayed an increased number of epidermoid cysts after exposure to UV light, no melanomas or premelanocytic alterations could be detected in this mouse model. The results therefore provide no evidence that HDGF has a transforming capacity in tumor development. Our results in combination with previous findings point to a possible role in cell differentiation and suggest that HDGF promotes tumor progression after secondary upregulation and may represent another protein fitting into the concept of non-oncogene addiction of tumor tissue

  1. Localization and expression of substance P in transgenic mice overexpressing human APP751 with the London (V717I) and Swedish (K670M/N671L) mutations.

    Science.gov (United States)

    Willis, Michael; Hutter-Paier, Birgit; Wietzorrek, Georg; Windisch, Manfred; Humpel, Christian; Knaus, Hans Günther; Marksteiner, Josef

    2007-04-27

    Substance P-like immunoreactivity (-LI) is found in neuritic plaques, and is reduced in patients suffering from Alzheimer disease (AD). In this study, we examined the distribution and expression of substance P in transgenic mice overexpressing human amyloid precursor protein (hAPP) APP751 with the London (V717I) and Swedish (K670M/N671L) mutations. Immunohistochemistry was performed to localize substance P- and glial fibrillary acidic protein-LI by confocal microscopy. In hAPP transgenic mice, the number of beta-amyloid plaques significantly increased from 6 to 12 months. About 5% of beta-amyloid plaques were substance P-immunoreactive. In transgenic mice, the morphology of substance P-immunoreactive structures changed by consisting of swollen and dystrophic neurites mostly associated with beta-amyloid plaques. The overall localization and the relative substance P densities were not different between wild type and transgenic mice at 6 and 12 months. At month 12, a dramatic change in the distribution pattern of substance P-LI was observed as it was now expressed in a high number of reactive astrocytes. This expression of substance P in astrocytes was mainly found in the hippocampal formation and thalamic nuclei with a preferential association with beta-amyloid plaques, whereas in cortical regions only faintly substance P-immunoreactive astrocytes were observed. This study indicates that substance P undergoes complex changes in this animal Alzheimer disease model. Future experiments including substance P antagonists are necessary to further explore the interaction between beta-amyloid deposits and substance P.

  2. Overexpression of TIMP-1 under the MMP-9 promoter interferes with wound healing in transgenic mice

    OpenAIRE

    Salonurmi, T.; Parikka, M.; Kontusaari, S.; Pirila, E.; Munaut, Carine; Salo, T.; Tryggvason, K.

    2004-01-01

    We have generated transgenic mice harboring the murine matrix metalloproteinase 9 (MMP-9) promoter cloned in front of human TIMP-1 cDNA. The transgenic mice were viable and fertile and exhibited normal growth and general development. During wound healing the mice were shown to express human TIMP-1 in keratinocytes that normally express MMP-9. However, the healing of skin wounds was significantly retarded with slow migration of keratinocytes over the wound in transgenic mice. In situ zymograph...

  3. Overexpression of homologous phytochrome genes in tomato: exploring the limits in photoperception

    NARCIS (Netherlands)

    Husaineid, S.H.; Kok, R.A.; Schreuder, M.E.L.; Plas, van der L.H.W.; Krol, van der A.R.

    2007-01-01

    Transgenic tomato [Lycopersicon esculentum (=Solanum lycopersicum)] lines overexpressing tomato PHYA, PHYB1, or PHYB2, under control of the constitutive double-35S promoter from cauliflower mosaic virus (CaMV) have been generated to test the level of saturation in individual phytochrome-signalling

  4. Functional characterization of CCR in birch (Betula platyphylla × Betula pendula) through overexpression and suppression analysis.

    Science.gov (United States)

    Zhang, Wenbo; Wei, Rui; Chen, Su; Jiang, Jing; Li, Huiyu; Huang, Haijiao; Yang, Guang; Wang, Shuo; Wei, Hairong; Liu, Guifeng

    2015-06-01

    We cloned a Cinnamoyl-CoA Reductase gene (BpCCR1) from an apical meristem and first internode of Betula platyphylla and characterized its functions in lignin biosynthesis, wood formation and tree growth through transgenic approaches. We generated overexpression and suppression transgenic lines and analyzed them in comparison with the wild-type in terms of lignin content, anatomical characteristics, height and biomass. We found that BpCCR1 overexpression could increase lignin content up to 14.6%, and its underexpression decreased lignin content by 6.3%. Surprisingly, modification of BpCCR1 expression led to conspicuous changes in wood characteristics, including xylem vessel number and arrangement, and secondary wall thickness. The growth of transgenic trees in terms of height was also significantly influenced by the modification of BpCCR1 genes. We discuss the functions of BpCCR1 in the context of a phylogenetic tree built with CCR genes from multiple species. © 2014 Scandinavian Plant Physiology Society.

  5. Starch characteristics of transgenic wheat (Triticum aestivum L.) overexpressing the Dx5 high molecular weight glutenin subunit are substantially equivalent to those in nonmodified wheat.

    Science.gov (United States)

    Beckles, Diane M; Tananuwong, Kanitha; Shoemaker, Charles F

    2012-04-01

    The effects of engineering higher levels of the High Molecular Weight Glutenin Dx5 subunit on starch characteristics in transgenic wheat (Triticum aestivum L.) grain were evaluated. This is important because of the interrelationship between starch and protein accumulation in grain, the strong biotechnological interest in modulating Dx5 levels and the increasing likelihood that transgenic wheat will be commercialized in the U.S. Unintended effects of Dx5 overexpression on starch could affect wheat marketability and therefore should be examined. Two controls with native levels of Dx5 were used: (i) the nontransformed Bobwhite cultivar, and (ii) a transgenic line (Bar-D) expressing a herbicide resistant (bar) gene, and they were compared with 2 transgenic lines (Dx5G and Dx5J) containing bar and additional copies of Dx5. There were few changes between Bar-D and Dx5G compared to Bobwhite. However, Dx5J, the line with the highest Dx5 protein (×3.5) accumulated 140% more hexose, 25% less starch and the starch had a higher frequency of longer amylopectin chains. These differences were not of sufficient magnitude to influence starch functionality, because granule morphology, crystallinity, amylose-to-amylopectin ratio, and the enthalpy of starch gelatinization and the amylose-lipid complex melting were similar to the control (P > 0.05). This overall similarity was borne out by Partial Least Squares-Discriminant Function Analysis, which could not distinguish among genotypes. Collectively our data imply that higher Dx5 can affect starch accumulation and some aspects of starch molecular structure but that the starches of the Dx5 transgenic wheat lines are substantially equivalent to the controls. Transgenic manipulation of biochemical pathways is an effective way to enhance food sensory quality, but it can also lead to unintended effects. These spurious changes are a concern to Government Regulatory Agencies and to those Industries that market the product. In this study we

  6. Cytoprotective effect of recombinant human erythropoietin produced in transgenic tobacco plants.

    Directory of Open Access Journals (Sweden)

    Farooqahmed S Kittur

    Full Text Available Asialo-erythropoietin, a desialylated form of human erythropoietin (EPO lacking hematopoietic activity, is receiving increased attention because of its broader protective effects in preclinical models of tissue injury. However, attempts to translate its protective effects into clinical practice is hampered by unavailability of suitable expression system and its costly and limit production from expensive mammalian cell-made EPO (rhuEPO(M by enzymatic desialylation. In the current study, we took advantage of a plant-based expression system lacking sialylating capacity but possessing an ability to synthesize complex N-glycans to produce cytoprotective recombinant human asialo-rhuEPO. Transgenic tobacco plants expressing asialo-rhuEPO were generated by stably co-expressing human EPO and β1,4-galactosyltransferase (GalT genes under the control of double CaMV 35S and glyceraldehyde-3-phosphate gene (GapC promoters, respectively. Plant-produced asialo-rhuEPO (asialo-rhuEPO(P was purified by immunoaffinity chromatography. Detailed N-glycan analysis using NSI-FTMS and MS/MS revealed that asialo-rhuEPO(P bears paucimannosidic, high mannose-type and complex N-glycans. In vitro cytoprotection assays showed that the asialo-rhuEPO(P (20 U/ml provides 2-fold better cytoprotection (44% to neuronal-like mouse neuroblastoma cells from staurosporine-induced cell death than rhuEPO(M (21%. The cytoprotective effect of the asialo-rhuEPO(P was found to be mediated by receptor-initiated phosphorylation of Janus kinase 2 (JAK2 and suppression of caspase 3 activation. Altogether, these findings demonstrate that plants are a suitable host for producing cytoprotective rhuEPO derivative. In addition, the general advantages of plant-based expression system can be exploited to address the cost and scalability issues related to its production.

  7. Transgenic mouse models of hormonal mammary carcinogenesis: advantages and limitations.

    Science.gov (United States)

    Kirma, Nameer B; Tekmal, Rajeshwar R

    2012-09-01

    Mouse models of breast cancer, especially transgenic and knockout mice, have been established as valuable tools in shedding light on factors involved in preneoplastic changes, tumor development and malignant progression. The majority of mouse transgenic models develop estrogen receptor (ER) negative tumors. This is seen as a drawback because the majority of human breast cancers present an ER positive phenotype. On the other hand, several transgenic mouse models have been developed that produce ER positive mammary tumors. These include mice over-expressing aromatase, ERα, PELP-1 and AIB-1. In this review, we will discuss the value of these models as physiologically relevant in vivo systems to understand breast cancer as well as some of the pitfalls involving these models. In all, we argue that the use of transgenic models has improved our understanding of the molecular aspects and biology of breast cancer. Copyright © 2011 Elsevier Ltd. All rights reserved.

  8. Thioredoxin (Trxo1) interacts with proliferating cell nuclear antigen (PCNA) and its overexpression affects the growth of tobacco cell culture.

    Science.gov (United States)

    Calderón, Aingeru; Ortiz-Espín, Ana; Iglesias-Fernández, Raquel; Carbonero, Pilar; Pallardó, Federico Vicente; Sevilla, Francisca; Jiménez, Ana

    2017-04-01

    Thioredoxins (Trxs), key components of cellular redox regulation, act by controlling the redox status of many target proteins, and have been shown to play an essential role in cell survival and growth. The presence of a Trx system in the nucleus has received little attention in plants, and the nuclear targets of plant Trxs have not been conclusively identified. Thus, very little is known about the function of Trxs in this cellular compartment. Previously, we studied the intracellular localization of PsTrxo1 and confirmed its presence in mitochondria and, interestingly, in the nucleus under standard growth conditions. In investigating the nuclear function of PsTrxo1 we identified proliferating cellular nuclear antigen (PCNA) as a PsTrxo1 target by means of affinity chromatography techniques using purified nuclei from pea leaves. Such protein-protein interaction was corroborated by dot-blot and bimolecular fluorescence complementation (BiFC) assays, which showed that both proteins interact in the nucleus. Moreover, PsTrxo1 showed disulfide reductase activity on previously oxidized recombinant PCNA protein. In parallel, we studied the effects of PsTrxo1 overexpression on Tobacco Bright Yellow-2 (TBY-2) cell cultures. Microscopy and flow-cytometry analysis showed that PsTrxo1 overexpression increases the rate of cell proliferation in the transformed lines, with a higher percentage of the S phase of the cell cycle at the beginning of the cell culture (days 1 and 3) and at the G2/M phase after longer times of culture (day 9), coinciding with an upregulation of PCNA protein. Furthermore, in PsTrxo1 overexpressed cells there is a decrease in the total cellular glutathione content but maintained nuclear GSH accumulation, especially at the end of the culture, which is accompanied by a higher mitotic index, unlike non-overexpressing cells. These results suggest that Trxo1 is involved in the cell cycle progression of TBY-2 cultures, possibly through its link with cellular PCNA

  9. Developmental and environmental regulation of the Nicotiana plumbaginifolia cytosolic Cu/Zn-superoxide dismutase promoter in transgenic tobacco.

    Science.gov (United States)

    Hérouart, D; Van Montagu, M; Inzé, D

    1994-03-01

    Superoxide dismutases (SODs) play a key role in the cellular defense against reactive oxygen species. To study the transcriptional regulation at the cellular level, the promoter of the Nicotiana plumbaginifolia cytosolic gene encoding Cu/ZnSOD (SODCc) was fused to the beta-glucuronidase (GUS) reporter gene (gusA) and analyzed in transgenic tobacco plants. The promoter was highly active in vascular bundles of leaves and stems, where it is confined to phloem cells. In flowers, GUS activity was detected in ovules and pollen grains, in pigmented tissues of petals, and in vascular tissue of ovaries and anthers. In response to treatment with the superoxide-generating herbicide paraquat, very strong GUS staining was observed in photosynthetically active cells of leaves and in some epidermal root cells of seedlings. The expression of the SODCc-gusA was also induced in seedlings after heat shock and chilling and after treatment with sulfhydryl antioxidants such as reduced glutathione and cysteine. It is postulated that SODCc expression is directly linked to a cell-specific production of excess superoxide radicals in the cytosol.

  10. Generating high temperature tolerant transgenic plants: Achievements and challenges.

    Science.gov (United States)

    Grover, Anil; Mittal, Dheeraj; Negi, Manisha; Lavania, Dhruv

    2013-05-01

    Production of plants tolerant to high temperature stress is of immense significance in the light of global warming and climate change. Plant cells respond to high temperature stress by re-programming their genetic machinery for survival and reproduction. High temperature tolerance in transgenic plants has largely been achieved either by over-expressing heat shock protein genes or by altering levels of heat shock factors that regulate expression of heat shock and non-heat shock genes. Apart from heat shock factors, over-expression of other trans-acting factors like DREB2A, bZIP28 and WRKY proteins has proven useful in imparting high temperature tolerance. Besides these, elevating the genetic levels of proteins involved in osmotic adjustment, reactive oxygen species removal, saturation of membrane-associated lipids, photosynthetic reactions, production of polyamines and protein biosynthesis process have yielded positive results in equipping transgenic plants with high temperature tolerance. Cyclic nucleotide gated calcium channel proteins that regulate calcium influxes across the cell membrane have recently been shown to be the key players in induction of high temperature tolerance. The involvement of calmodulins and kinases in activation of heat shock factors has been implicated as an important event in governing high temperature tolerance. Unfilled gaps limiting the production of high temperature tolerant transgenic plants for field level cultivation are discussed. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  11. Overexpression of hepatocyte growth factor in SBMA model mice has an additive effect on combination therapy with castration

    International Nuclear Information System (INIS)

    Ding, Ying; Adachi, Hiroaki; Katsuno, Masahisa; Huang, Zhe; Jiang, Yue-Mei; Kondo, Naohide; Iida, Madoka; Tohnai, Genki; Nakatsuji, Hideaki; Funakoshi, Hiroshi; Nakamura, Toshikazu; Sobue, Gen

    2015-01-01

    Spinal and bulbar muscular atrophy (SBMA) is an inherited motor neuron disease caused by the expansion of a polyglutamine (polyQ)-encoding tract within the androgen receptor (AR) gene. The pathologic features of SBMA are motor neuron loss in the spinal cord and brainstem and diffuse nuclear accumulation and nuclear inclusions of mutant AR in residual motor neurons and certain visceral organs. Hepatocyte growth factor (HGF) is a polypeptide growth factor which has neuroprotective properties. To investigate whether HGF overexpression can affect disease progression in a mouse model of SBMA, we crossed SBMA transgenic model mice expressing an AR gene with an expanded CAG repeat with mice overexpressing HGF. Here, we report that high expression of HGF induces Akt phosphorylation and modestly ameliorated motor symptoms in an SBMA transgenic mouse model treated with or without castration. These findings suggest that HGF overexpression can provide a potential therapeutic avenue as a combination therapy with disease-modifying therapies in SBMA. - Highlights: • HGF overexpression ameliorates the motor phenotypes of the SBMA mouse model. • HGF overexpression induces Akt phosphorylation in the SBMA mouse model. • This is the first report of combination therapy in a mouse model of polyQ diseases.

  12. Overexpression of hepatocyte growth factor in SBMA model mice has an additive effect on combination therapy with castration

    Energy Technology Data Exchange (ETDEWEB)

    Ding, Ying [Department of Neurology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Adachi, Hiroaki, E-mail: hadachi-ns@umin.org [Department of Neurology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Department of Neurology, University of Occupational and Environmental Health School of Medicine, 1-1 Iseigaoka, Yahata-nishi-ku, Kitakyushu 807-8555 (Japan); Katsuno, Masahisa [Department of Neurology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Huang, Zhe [Department of Neurology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Department of Neurology, University of Occupational and Environmental Health School of Medicine, 1-1 Iseigaoka, Yahata-nishi-ku, Kitakyushu 807-8555 (Japan); Jiang, Yue-Mei; Kondo, Naohide; Iida, Madoka; Tohnai, Genki; Nakatsuji, Hideaki [Department of Neurology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Funakoshi, Hiroshi [Center for Advanced Research and Education, Asahikawa Medical University, 1-1-1- Higashinijo Midorigaoka, Asahikawa 078-8510 (Japan); Nakamura, Toshikazu [Neurogen Inc., 1-1-52-201 Nakahozumi, Ibaraki 567-0034 (Japan); Sobue, Gen, E-mail: sobueg@med.nagoya-u.ac.jp [Department of Neurology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan); Research Division of Dementia and Neurodegenerative Disease, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550 (Japan)

    2015-12-25

    Spinal and bulbar muscular atrophy (SBMA) is an inherited motor neuron disease caused by the expansion of a polyglutamine (polyQ)-encoding tract within the androgen receptor (AR) gene. The pathologic features of SBMA are motor neuron loss in the spinal cord and brainstem and diffuse nuclear accumulation and nuclear inclusions of mutant AR in residual motor neurons and certain visceral organs. Hepatocyte growth factor (HGF) is a polypeptide growth factor which has neuroprotective properties. To investigate whether HGF overexpression can affect disease progression in a mouse model of SBMA, we crossed SBMA transgenic model mice expressing an AR gene with an expanded CAG repeat with mice overexpressing HGF. Here, we report that high expression of HGF induces Akt phosphorylation and modestly ameliorated motor symptoms in an SBMA transgenic mouse model treated with or without castration. These findings suggest that HGF overexpression can provide a potential therapeutic avenue as a combination therapy with disease-modifying therapies in SBMA. - Highlights: • HGF overexpression ameliorates the motor phenotypes of the SBMA mouse model. • HGF overexpression induces Akt phosphorylation in the SBMA mouse model. • This is the first report of combination therapy in a mouse model of polyQ diseases.

  13. Prothymosin α overexpression contributes to the development of pulmonary emphysema

    Science.gov (United States)

    Su, Bing-Hua; Tseng, Yau-Lin; Shieh, Gia-Shing; Chen, Yi-Cheng; Shiang, Ya-Chieh; Wu, Pensee; Li, Kuo-Jung; Yen, Te-Hsin; Shiau, Ai-Li; Wu, Chao-Liang

    2013-01-01

    Emphysema is one of the disease conditions that comprise chronic obstructive pulmonary disease. Prothymosin α transgenic mice exhibit an emphysema phenotype, but the pathophysiological role of prothymosin α in emphysema remains unclear. Here we show that prothymosin α contributes to the pathogenesis of emphysema by increasing acetylation of histones and nuclear factor-kappaB, particularly upon cigarette smoke exposure. We find a positive correlation between prothymosin α levels and the severity of emphysema in prothymosin α transgenic mice and emphysema patients. Prothymosin α overexpression increases susceptibility to cigarette smoke-induced emphysema, and cigarette smoke exposure further enhances prothymosin α expression. We show that prothymosin α inhibits the association of histone deacetylases with histones and nuclear factor-kappaB, and that prothymosin α overexpression increases expression of nuclear factor-kappaB-dependent matrix metalloproteinase 2 and matrix metalloproteinase 9, which are found in the lungs of patients with chronic obstructive pulmonary disease. These results demonstrate the clinical relevance of prothymosin α in regulating acetylation events during the pathogenesis of emphysema. PMID:23695700

  14. Overexpression of Jatropha Gibberellin 2-oxidase 6 (JcGA2ox6 Induces Dwarfism and Smaller Leaves, Flowers and Fruits in Arabidopsis and Jatropha

    Directory of Open Access Journals (Sweden)

    Ying-Xiong Hu

    2017-12-01

    Full Text Available Gibberellins (GAs are plant hormones that play fundamental roles in plant growth and development. Gibberellin 2-oxidase (GA2ox plays a direct role in determining the levels of bioactive GAs by catalyzing bioactive GAs or their immediate precursors to inactive forms. In this study, a GA2ox gene, designated JcGA2ox6, was isolated from Jatropha curcas. JcGA2ox6 is expressed in all tissues of adult Jatropha, with the highest expression level in male flowers and the lowest expression level in young leaves. Overexpression of JcGA2ox6 in Arabidopsis resulted in a typical dwarf phenotype, along with late flowering, smaller leaves and flowers, shorter siliques and smaller seeds. Similarly, when JcGA2ox6 was overexpressed in Jatropha, the transgenic plants exhibited a dwarf phenotype with dark-green leaves and smaller inflorescences, flowers, fruits and seeds. However, the flowering time of Jatropha was not affected by overexpression of JcGA2ox6, unlike that in the transgenic Arabidopsis. Moreover, the number of flowers per inflorescence, the weight of 10 seeds and the seed oil content were significantly decreased in transgenic Jatropha. The results indicated that overexpression of JcGA2ox6 had a great impact on the vegetative and reproductive growth of transgenic Jatropha. Furthermore, we found that the dwarf phenotype of transgenic Jatropha was caused by a decrease in endogenous bioactive GA4, which was correlated with the degree of dwarfism.

  15. Overexpression of dimethylarginine dimethylaminohydrolase 1 attenuates airway inflammation in a mouse model of asthma.

    Directory of Open Access Journals (Sweden)

    Kayla G Kinker

    Full Text Available Levels of asymmetric dimethylarginine (ADMA, an endogenous inhibitor of nitric oxide synthase, are increased in lung, sputum, exhaled breath condensate and plasma samples from asthma patients. ADMA is metabolized primarily by dimethylarginine dimethylaminohydrolase 1 (DDAH1 and DDAH2. We determined the effect of DDAH1 overexpression on development of allergic inflammation in a mouse model of asthma. The expression of DDAH1 and DDAH2 in mouse lungs was determined by RT-quantitative PCR (qPCR. ADMA levels in bronchoalveolar lavage fluid (BALF and serum samples were determined by mass spectrometry. Wild type and DDAH1-transgenic mice were intratracheally challenged with PBS or house dust mite (HDM. Airway inflammation was assessed by bronchoalveolar lavage (BAL total and differential cell counts. The levels of IgE and IgG1 in BALF and serum samples were determined by ELISA. Gene expression in lungs was determined by RNA-Seq and RT-qPCR. Our data showed that the expression of DDAH1 and DDAH2 was decreased in the lungs of mice following HDM exposure, which correlated with increased ADMA levels in BALF and serum. Transgenic overexpression of DDAH1 resulted in decreased BAL total cell and eosinophil numbers following HDM exposure. Total IgE levels in BALF and serum were decreased in HDM-exposed DDAH1-transgenic mice compared to HDM-exposed wild type mice. RNA-Seq results showed downregulation of genes in the inducible nitric oxide synthase (iNOS signaling pathway in PBS-treated DDAH1-transgenic mice versus PBS-treated wild type mice and downregulation of genes in IL-13/FOXA2 signaling pathway in HDM-treated DDAH1-transgenic mice versus HDM-treated wild type mice. Our findings suggest that decreased expression of DDAH1 and DDAH2 in the lungs may contribute to allergic asthma and overexpression of DDAH1 attenuates allergen-induced airway inflammation through modulation of Th2 responses.

  16. Expression of an (E-β-farnesene synthase gene from Asian peppermint in tobacco affected aphid infestation

    Directory of Open Access Journals (Sweden)

    Xiudao Yu

    2013-10-01

    Full Text Available Aphids are major agricultural pests that cause significant yield losses in crop plants each year. (E-β-farnesene (EβF is the main or only component of an alarm pheromone involved in chemical communication within aphid species and particularly in the avoidance of predation. EβF also occurs in the essential oil of some plant species, and is catalyzed by EβF synthase. By using oligonucleotide primers designed from the known sequence of an EβF synthase gene from black peppermint (Mentha × piperita, two cDNA sequences, MaβFS1 and MaβFS2, were isolated from Asian peppermint (Mentha asiatica. Expression pattern analysis showed that the MaβFS1 gene exhibited higher expression in flowers than in roots, stems and leaves at the transcriptional level. Overexpression of MaβFS1 in tobacco plants resulted in emission of pure EβF ranging from 2.62 to 4.85 ng d− 1 g− 1 of fresh tissue. Tritrophic interactions involving peach aphids (Myzus persicae, and predatory lacewing (Chrysopa septempunctata larvae demonstrated that transgenic tobacco expressing MaβFS1 had lower aphid infestation. This result suggested that the EβF synthase gene from Asian peppermint could be a good candidate for genetic engineering of agriculturally important crop plants.

  17. Overexpression of a Pathogenesis-Related Protein 10 Enhances Biotic and Abiotic Stress Tolerance in Rice

    Directory of Open Access Journals (Sweden)

    Jingni Wu

    2016-12-01

    Full Text Available Pathogenesis-related proteins play multiple roles in plant development and biotic and abiotic stress tolerance. Here, we characterize a rice defense related gene named “jasmonic acid inducible pathogenesis-related class 10” (JIOsPR10 to gain an insight into its functional properties. Semi-quantitative RT-PCR analysis showed up-regulation of JIOsPR10 under salt and drought stress conditions. Constitutive over-expression JIOsPR10 in rice promoted shoot and root development in transgenic plants, however, their productivity was unaltered. Further experiments exhibited that the transgenic plants showed reduced susceptibility to rice blast fungus, and enhanced salt and drought stress tolerance as compared to the wild type. A comparative proteomic profiling of wild type and transgenic plants showed that overexpression of JIOsPR10 led to the differential modulation of several proteins mainly related with oxidative stresses, carbohydrate metabolism, and plant defense. Taken together, our findings suggest that JIOsPR10 plays important roles in biotic and abiotic stresses tolerance probably by activation of stress related proteins.

  18. A novel 5-enolpyruvylshikimate-3-phosphate synthase shows high glyphosate tolerance in Escherichia coli and tobacco plants.

    Directory of Open Access Journals (Sweden)

    Gaoyi Cao

    Full Text Available A key enzyme in the shikimate pathway, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS is the primary target of the broad-spectrum herbicide glyphosate. Identification of new aroA genes coding for EPSPS with a high level of glyphosate tolerance is essential for the development of glyphosate-tolerant crops. In the present study, the glyphosate tolerance of five bacterial aroA genes was evaluated in the E. coli aroA-defective strain ER2799 and in transgenic tobacco plants. All five aroA genes could complement the aroA-defective strain ER2799, and AM79 aroA showed the highest glyphosate tolerance. Although glyphosate treatment inhibited the growth of both WT and transgenic tobacco plants, transgenic plants expressing AM79 aroA tolerated higher concentration of glyphosate and had a higher fresh weight and survival rate than plants expressing other aroA genes. When treated with high concentration of glyphosate, lower shikimate content was detected in the leaves of transgenic plants expressing AM79 aroA than transgenic plants expressing other aroA genes. These results suggest that AM79 aroA could be a good candidate for the development of transgenic glyphosate-tolerant crops.

  19. Female-to-male sex reversal in mice caused by transgenic overexpression of Dmrt1

    DEFF Research Database (Denmark)

    Zhao, Liang; Svingen, Terje; Ting Ng, Ee

    2015-01-01

    for primary sex determination and instead maintains Sertoli cell phenotype in postnatal testes. Here, we report that enforced expression of Dmrt1 in XX mouse fetal gonads using a Wt1-BAC transgene system is sufficient to drive testicular differentiation and male secondary sex development. XX transgenic fetal...... into testicular cell types, including steroidogenic fetal Leydig cells and non-meiotic germ cells. As a consequence, male internal and external reproductive organs developed postnatally, with an absence of female reproductive tissues. These results reveal that Dmrt1 has retained its ability to act as the primary...

  20. Overexpression of an Arabidopsis heterogeneous nuclear ribonucleoprotein gene, AtRNP1, affects plant growth and reduces plant tolerance to drought and salt stresses

    International Nuclear Information System (INIS)

    Wang, Zhenyu; Zhao, Xiuyang; Wang, Bing; Liu, Erlong; Chen, Ni; Zhang, Wei; Liu, Heng

    2016-01-01

    Heterogeneous nuclear ribonucleoproteins (hnRNPs) participate in diverse regulations of plant growth and environmental stress responses. In this work, an Arabidopsis hnRNP of unknown function, AtRNP1, was investigated. We found that AtRNP1 gene is highly expressed in rosette and cauline leaves, and slightly induced under drought, salt, osmotic and ABA stresses. AtRNP1 protein is localized to both the nucleus and cytoplasm. We performed homologous overexpression of AtRNP1 and found that the transgenic plants showed shortened root length and plant height, and accelerated flowering. In addition, the transgenic plants also showed reduced tolerance to drought, salt, osmotic and ABA stresses. Further studies revealed that under both normal and stress conditions, the proline contents in the transgenic plants are markedly decreased, associated with reduced expression levels of a proline synthase gene and several stress-responsive genes. These results suggested that the overexpression of AtRNP1 negatively affects plant growth and abiotic stress tolerance. - Highlights: • AtRNP1 is a widely expressed gene and its expression is slightly induced under abiotic stresses. • AtRNP1 protein is localized to both the nucleus and cytoplasm. • Overexpression of AtRNP1 affects plant growth. • Overexpression of AtRNP1 reduces plant tolerance to drought and salt stresses. • AtRNP1 overexpression plants show decreased proline accumulation and stress-responsive gene expressions.

  1. Overexpression of an Arabidopsis heterogeneous nuclear ribonucleoprotein gene, AtRNP1, affects plant growth and reduces plant tolerance to drought and salt stresses

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Zhenyu, E-mail: wzy72609@163.com [Ministry of Education Key Laboratory of Cell Activities and Stress Adaptations, School of Life Sciences, Lanzhou University, Lanzhou 730030 (China); Zhao, Xiuyang, E-mail: xiuzh@psb.vib-ugent.be [Ministry of Education Key Laboratory of Cell Activities and Stress Adaptations, School of Life Sciences, Lanzhou University, Lanzhou 730030 (China); Wang, Bing, E-mail: wangbing@ibcas.ac.cn [Ministry of Education Key Laboratory of Cell Activities and Stress Adaptations, School of Life Sciences, Lanzhou University, Lanzhou 730030 (China); Liu, Erlong, E-mail: liuel14@lzu.edu.cn [Ministry of Education Key Laboratory of Cell Activities and Stress Adaptations, School of Life Sciences, Lanzhou University, Lanzhou 730030 (China); Chen, Ni, E-mail: 63710156@qq.com [Ministry of Education Key Laboratory of Cell Activities and Stress Adaptations, School of Life Sciences, Lanzhou University, Lanzhou 730030 (China); Zhang, Wei, E-mail: wzhang1216@yahoo.com [Shanghai Key Laboratory of Bio-Energy Crops, School of Life Sciences, Shanghai University, Shanghai 200444 (China); Liu, Heng, E-mail: hengliu@lzu.edu.cn [Ministry of Education Key Laboratory of Cell Activities and Stress Adaptations, School of Life Sciences, Lanzhou University, Lanzhou 730030 (China)

    2016-04-01

    Heterogeneous nuclear ribonucleoproteins (hnRNPs) participate in diverse regulations of plant growth and environmental stress responses. In this work, an Arabidopsis hnRNP of unknown function, AtRNP1, was investigated. We found that AtRNP1 gene is highly expressed in rosette and cauline leaves, and slightly induced under drought, salt, osmotic and ABA stresses. AtRNP1 protein is localized to both the nucleus and cytoplasm. We performed homologous overexpression of AtRNP1 and found that the transgenic plants showed shortened root length and plant height, and accelerated flowering. In addition, the transgenic plants also showed reduced tolerance to drought, salt, osmotic and ABA stresses. Further studies revealed that under both normal and stress conditions, the proline contents in the transgenic plants are markedly decreased, associated with reduced expression levels of a proline synthase gene and several stress-responsive genes. These results suggested that the overexpression of AtRNP1 negatively affects plant growth and abiotic stress tolerance. - Highlights: • AtRNP1 is a widely expressed gene and its expression is slightly induced under abiotic stresses. • AtRNP1 protein is localized to both the nucleus and cytoplasm. • Overexpression of AtRNP1 affects plant growth. • Overexpression of AtRNP1 reduces plant tolerance to drought and salt stresses. • AtRNP1 overexpression plants show decreased proline accumulation and stress-responsive gene expressions.

  2. Ectopic expression of class 1 KNOX genes induce and adventitious shoot regeneration and alter growth and development of tobacco (Nicotiana tabacum L) and European plum (Prunus domestica L)

    Science.gov (United States)

    Transgenic plants of tobacco (Nicotiana tabacum L) and plum (Prunus domestica L) were produced by transforming with apple class 1 KNOX genes (MdKN1 and MdKN2) or corn KN1 gene. Transgenic tobacco plants were regenerated in vitro from transformed leaf discs cultured in a tissue medium lacking cytoki...

  3. Ectopic over-expression of peroxisomal ascorbate peroxidase (SbpAPX) gene confers salt stress tolerance in transgenic peanut (Arachis hypogaea).

    Science.gov (United States)

    Singh, Natwar; Mishra, Avinash; Jha, Bhavanath

    2014-08-15

    Peroxisomal ascorbate peroxidase gene (SbpAPX) of an extreme halophyte Salicornia brachiata imparts abiotic stress endurance and plays a key role in the protection against oxidative stress. The cloned SbpAPX gene was transformed to local variety of peanut and about 100 transgenic plants were developed using optimized in vitro regeneration and Agrobacterium mediated genetic transformation method. The T0 transgenic plants were confirmed for the gene integration; grown under controlled condition in containment green house facility; seeds were harvested and T1 plants were raised. Transgenic plants (T1) were further confirmed by PCR using gene specific primers and histochemical GUS assay. About 40 transgenic plants (T1) were selected randomly and subjected for salt stress tolerance study. Transgenic plants remained green however non-transgenic plants showed bleaching and yellowish leaves under salt stress conditions. Under stress condition, transgenic plants continued normal growth and completed their life cycle. Transgenic peanut plants exhibited adequate tolerance under salt stress condition and thus could be explored for the cultivation in salt affected areas for the sustainable agriculture. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Alterations in Adiposity and Glucose Homeostasis in Adult Gasp-1 Overexpressing Mice

    Directory of Open Access Journals (Sweden)

    Luce Périè

    2017-12-01

    Full Text Available Background/Aims: Myostatin is known as a powerful negative regulator of muscle growth playing a key role in skeletal muscle homeostasis. Recent studies revealed that myostatin-deficient mice lead to an increase of insulin sensitivity, a decrease of adiposity and a resistance to obesity, showing that myostatin can also impact on metabolism. Thus, myostatin appeared as a potential therapeutic target to treat insulin resistance. Methods: We generated transgenic mice overexpressing Gasp-1, a myostatin inhibitor. Results: Surprisingly, we found that these mice gained weight with age due to an increase in fat mass associated with ectopic fat accumulation. In addition, these mice developed an adipocyte hypertrophy, hyperglycemia, hyperinsulinemia, muscle and hepatic insulin resistance. Understanding the molecular networks controlling this insulin resistance responsiveness in overexpressing Gasp-1 mice is essential. Molecular analyses revealed a deregulation of adipokines and muscle cytokines expression, but also an increase in plasma myostatin levels. The increase in myostatin bioactivity by a positive feedback mechanism in the Tg(Gasp-1 transgenic mice could lead to this combination of phenotypes. Conclusion: Altogether, these data suggested that overexpressing Gasp-1 mice develop most of the symptoms associated with metabolic syndrome and could be a relevant model for the study of obesity or type 2 diabetes.

  5. Selection of Housekeeping Genes for Transgene Expression Analysis in Eucommia ulmoides Oliver Using Real-Time RT-PCR

    Directory of Open Access Journals (Sweden)

    Ren Chen

    2010-01-01

    Full Text Available In order to select appropriate housekeeping genes for accurate calibration of experimental variations in real-time (RT- PCR results in transgene expression analysis, particularly with respect to the influence of transgene on stability of endogenous housekeeping gene expression in transgenic plants, we outline a reliable strategy to identify the optimal housekeeping genes from a set of candidates by combining statistical analyses of their (RT- PCR amplification efficiency, gene expression stability, and transgene influences. We used the strategy to select two genes, ACTα and EF1α, from 10 candidate housekeeping genes, as the optimal housekeeping genes to evaluate transgenic Eucommia ulmoides Oliver root lines overexpressing IPPI or FPPS1 genes, which are involved in isoprenoid biosynthesis.

  6. Overexpression of a modiifed AM79 aroA gene in transgenic maize confers high tolerance to glyphosate

    Institute of Scientific and Technical Information of China (English)

    REN Zhen-jing; CAO Gao-yi; ZHANG Yu-wen; LIU Yan; LIU Yun-jun

    2015-01-01

    It has previously been shown that a bacterial 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) encoding gene AM79 aroA can be a candidate gene to develop glyphosate-tolerant transgenic crops (Cao et al. 2012). In this study, AM79 aroA was redesigned using the plant biased codons and eliminating the motifs which would lead to the instability of mRNA, to create a synthetic gene that would be expressed highly in plant cel s. The redesigned and artiifcial y synthesized gene, named as mAM79, was cloned into plant expression vector pM3301UbiSpAM79, where mAM79 is fused with signal peptide sequence of pea rib-1,5-bisphospate carboxylase (rbcS) smal subunit and control ed by ubiquitin promoter. The plasmid was transformed into maize (Zea mays) immature embryos using Agrobacterium-mediated transformation method. Total 74 regenerated plants were obtained and PCR analysis showed that these transgenic plants had the integration of mAM79. Southern blot analysis was performed on the genomic DNA from four transgenic lines, and the result showed that one or two copies of mAM79 were integrated into maize genome. RT-PCR analysis result indicated that mAM79 was highly transcribed in transgenic maize plants. When sprayed with glyphosate, transgenic maize line AM85 and AM72 could tolerate 4-fold of commercial usage of glyphosate;however, al the non-transgenic maize plants were kil ed by glyphosate. The results in this study conifrmed that mAM79 could be used to develop glyphosate-tolerant maize, and the obtained transgenic maize lines could be used for the breeding of glyphosate-tolerant maize.

  7. Over-expression of Sub1 A, a submergence tolerance gene from ...

    African Journals Online (AJOL)

    Sub1A, an ethylene-response-factor-like (ERE-like) gene, mediates the extinguished submergence tolerance of rice. To gain further insight into the function of Sub1A in other species, we transformed tobacco plants with the gene under the control of the ubiquitin promoter. Compared to the wild-type plants, transgenic plants ...

  8. Variation Analysis of Physiological Traits in Betula platyphylla Overexpressing TaLEA-ThbZIP Gene under Salt Stress.

    Directory of Open Access Journals (Sweden)

    Xiyang Zhao

    Full Text Available The aim of this study was to determine whether transgenic birch (Betula platyphylla ectopic overexpressing a late embryogenesis abundant (LEA gene and a basic leucine zipper (bZIP gene from the salt-tolerant genus Tamarix (salt cedar show increased tolerance to salt (NaCl stress. Co-transfer of TaLEA and ThbZIP in birch under the control of two independent CaMV 35S promoters significantly enhanced salt stress. PCR and northern blot analyses indicated that the two genes were ectopically overexpressed in several dual-gene transgenic birch lines. We compared the effects of salt stress among three transgenic birch lines (L-4, L-5, and L-8 and wild type (WT. In all lines, the net photosynthesis values were higher before salt stress treatment than afterwards. After the salt stress treatment, the transgenic lines L-4 and L-8 showed higher values for photosynthetic traits, chlorophyll fluorescence, peroxidase and superoxide dismutase activities, and lower malondialdehyde and Na+ contents, compared with those in WT and L-5. These different responses to salt stress suggested that the transcriptional level of the TaLEA and ThbZIP genes differed among the transgenic lines, resulting in a variety of genetic and phenotypic effects. The results of this research can provide a theoretical basis for the genetic engineering of salt-tolerant trees.

  9. Tubular overexpression of gremlin induces renal damage susceptibility in mice.

    Directory of Open Access Journals (Sweden)

    Alejandra Droguett

    Full Text Available A growing number of patients are recognized worldwide to have chronic kidney disease. Glomerular and interstitial fibrosis are hallmarks of renal progression. However, fibrosis of the kidney remains an unresolved challenge, and its molecular mechanisms are still not fully understood. Gremlin is an embryogenic gene that has been shown to play a key role in nephrogenesis, and its expression is generally low in the normal adult kidney. However, gremlin expression is elevated in many human renal diseases, including diabetic nephropathy, pauci-immune glomerulonephritis and chronic allograft nephropathy. Several studies have proposed that gremlin may be involved in renal damage by acting as a downstream mediator of TGF-β. To examine the in vivo role of gremlin in kidney pathophysiology, we generated seven viable transgenic mouse lines expressing human gremlin (GREM1 specifically in renal proximal tubular epithelial cells under the control of an androgen-regulated promoter. These lines demonstrated 1.2- to 200-fold increased GREM1 expression. GREM1 transgenic mice presented a normal phenotype and were without proteinuria and renal function involvement. In response to the acute renal damage cause by folic acid nephrotoxicity, tubule-specific GREM1 transgenic mice developed increased proteinuria after 7 and 14 days compared with wild-type treated mice. At 14 days tubular lesions, such as dilatation, epithelium flattening and hyaline casts, with interstitial cell infiltration and mild fibrosis were significantly more prominent in transgenic mice than wild-type mice. Tubular GREM1 overexpression was correlated with the renal upregulation of profibrotic factors, such as TGF-β and αSMA, and with increased numbers of monocytes/macrophages and lymphocytes compared to wild-type mice. Taken together, our results suggest that GREM1-overexpressing mice have an increased susceptibility to renal damage, supporting the involvement of gremlin in renal damage

  10. A rice chloroplast transit peptide sequence does not alter the cytoplasmic localization of sheep serotonin N-acetyltransferase expressed in transgenic rice plants.

    Science.gov (United States)

    Byeon, Yeong; Lee, Hyoung Yool; Lee, Kyungjin; Back, Kyoungwhan

    2014-09-01

    Ectopic overexpression of melatonin biosynthetic genes of animal origin has been used to generate melatonin-rich transgenic plants to examine the functional roles of melatonin in plants. However, the subcellular localization of these proteins expressed in the transgenic plants remains unknown. We studied the localization of sheep (Ovis aries) serotonin N-acetyltransferase (OaSNAT) and a translational fusion of a rice SNAT transit peptide to OaSNAT (TS:OaSNAT) in plants. Laser confocal microscopy analysis revealed that both OaSNAT and TS:OaSNAT proteins were localized to the cytoplasm even with the addition of the transit sequence to OaSNAT. Transgenic rice plants overexpressing the TS:OaSNAT fusion transgene exhibited high SNAT enzyme activity relative to untransformed wild-type plants, but lower activity than transgenic rice plants expressing the wild-type OaSNAT gene. Melatonin levels in both types of transgenic rice plant corresponded well with SNAT enzyme activity levels. The TS:OaSNAT transgenic lines exhibited increased seminal root growth relative to wild-type plants, but less than in the OaSNAT transgenic lines, confirming that melatonin promotes root growth. Seed-specific OaSNAT expression under the control of a rice prolamin promoter did not confer high levels of melatonin production in transgenic rice seeds compared with seeds from transgenic plants expressing OaSNAT under the control of the constitutive maize ubiquitin promoter. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  11. Overexpression of the transporters AtZIP1 and AtMTP1 in cassava changes zinc accumulation and partitioning

    Directory of Open Access Journals (Sweden)

    Eliana eGaitan-Solis

    2015-07-01

    Full Text Available Zinc deficiency in humans is a serious problem worldwide with an estimated one third of populations at risk for insufficient zinc in diet which leads to impairment of cognitive abilities and immune system function. The goal of this research was to increase the bioavailable zinc in the edible portion of cassava roots to improve the overall zinc nutrition of populations that rely on cassava as a dietary staple. To increase zinc concentrations, two A. thaliana genes coding for ZIP1 and MTP1 were overexpressed with a tuber-specific or constitutive promoter. Eighteen transgenic events from four constructs, out of a total of 73 events generated, showed significantly higher zinc concentrations in the edible portion of the storage root compared to the non-transgenic controls. The zinc content in the transgenic lines ranged from 4 - 73 mg/Kg Dry Weight (DW as compared to the non-transgenic control which contained 8 mg/Kg. Striking changes in whole plant phenotype such as smaller plant size and chlorotic leaves were observed in transgenic lines that over accumulated zinc. In a confined field trial five transgenic events grown for 12 months showed a range of zinc concentrations from 18 – 217 mg/Kg DW. Although the overexpression of zinc transporters was successful in increasing the zinc concentrations in 25% of the transgenic lines generated, it also resulted in a decrease in plant and tuber size and overall yield due to what appears to be zinc deficiency in the aerial parts of the plant.

  12. The use of transgenic animals to study lipoprotein metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Rubin, E.M.; Plump, A.S.

    1993-12-01

    The application of transgenic technology to lipoprotein metabolism and atherosclerosis was first reported in 1988. Today, a large percentage of the genes involved in lipoprotein metabolism have been overexpressed in mice, and a substantial number of these same genes have been disrupted by homologous recombination in embryonic stem (ES) cells. The utility of animal models of lipoprotein metabolism and atherosclerosis is far-reaching given the complex nature of these systems. There are at least 17 known genes directly involved in lipoprotein metabolism and likely dozens more may be involved. This massive network of interacting factors has necessitated the development of in vivo systems which can be subject to genetic manipulation. The power of overexpression is obvious: elucidating function in a relatively controlled genetic environment in which the whole system is present and operational. The not-so-obvious problem with transgenics is ``background,`` or for purposes of the current discussion, the mouse`s own lipoprotein system. With the advent of gene knockout, we have been given the ability to overcome ``background.`` By recreating the genetic complement of the mouse we can alter a system in essentially any manner desired. As unique tools, and in combination with one another, the overexpression of foreign genes and the targeted disruption or alteration of endogenous genes has already and will continue to offer a wealth of information on the biology of lipoprotein metabolism and its effect on atherosclerosis susceptibility.

  13. Decreased nuclear β-catenin, tau hyperphosphorylation and neurodegeneration in GSK-3β conditional transgenic mice

    OpenAIRE

    Lucas, José J.; Hernández, Félix; Gómez-Ramos, Pilar; Morán, María A.; Hen, René; Avila, Jesús

    2001-01-01

    Glycogen synthase kinase-3β (GSK-3β) has been postulated to mediate Alzheimer’s disease tau hyperphosphorylation, β-amyloid-induced neurotoxicity and presenilin-1 mutation pathogenic effects. By using the tet-regulated system we have produced conditional transgenic mice overexpressing GSK-3β in the brain during adulthood while avoiding perinatal lethality due to embryonic transgene expression. These mice show decreased levels of nuclear β-catenin and hyperphosphorylation of tau in hippocampal...

  14. Overexpression of plasma membrane H+-ATPase in guard cells promotes light-induced stomatal opening and enhances plant growth.

    Science.gov (United States)

    Wang, Yin; Noguchi, Ko; Ono, Natsuko; Inoue, Shin-ichiro; Terashima, Ichiro; Kinoshita, Toshinori

    2014-01-07

    Stomatal pores surrounded by a pair of guard cells in the plant epidermis control gas exchange between plants and the atmosphere in response to light, CO2, and the plant hormone abscisic acid. Light-induced stomatal opening is mediated by at least three key components: the blue light receptor phototropin (phot1 and phot2), plasma membrane H(+)-ATPase, and plasma membrane inward-rectifying K(+) channels. Very few attempts have been made to enhance stomatal opening with the goal of increasing photosynthesis and plant growth, even though stomatal resistance is thought to be the major limiting factor for CO2 uptake by plants. Here, we show that transgenic Arabidopsis plants overexpressing H(+)-ATPase using the strong guard cell promoter GC1 showed enhanced light-induced stomatal opening, photosynthesis, and plant growth. The transgenic plants produced larger and increased numbers of rosette leaves, with ∼42-63% greater fresh and dry weights than the wild type in the first 25 d of growth. The dry weights of total flowering stems of 45-d-old transgenic plants, including seeds, siliques, and flowers, were ∼36-41% greater than those of the wild type. In addition, stomata in the transgenic plants closed normally in response to darkness and abscisic acid. In contrast, the overexpression of phototropin or inward-rectifying K(+) channels in guard cells had no effect on these phenotypes. These results demonstrate that stomatal aperture is a limiting factor in photosynthesis and plant growth, and that manipulation of stomatal opening by overexpressing H(+)-ATPase in guard cells is useful for the promotion of plant growth.

  15. Unwinding after high salinity stress: Pea DNA helicase 45 over- expression in tobacco confers high salinity tolerance without affecting yield (abstract)

    International Nuclear Information System (INIS)

    Tuteja, N.

    2005-01-01

    Soil salinity is an increasing threat for agriculture and is a major factor in reducing plant productivity; therefore, it is necessary to obtain salinity-tolerant varieties. A typical characteristic of soil salinity is the induction of multiple stress- inducible genes. Some of the genes encoding osmolytes, ion channels or enzymes are able to confer salinity-tolerant phenotypes when transferred to sensitive plants. As salinity stress affects the cellular gene-expression machinery, it is evident that molecules involved in nucleic acid processing including helicases, are likely to be affected as well. DNA helicases unwind duplex DNA and are involved in replication, repair, recombination and transcription while RNA helicases unfold the secondary structures in RNA and are involved in transcription, ribosome biogenesis and translation initiation. We have earlier reported the isolation of a pea DNA helicase 45 (PDH45) that exhibits striking homology with eIF-4A (Plant J. 24:219-230,2000). Here we report that PDH45 mRNA is induced in pea seedlings in response to high salt and its over- expression driven by a constitutive CAMV-355-promoter in tobacco plants confers salinity tolerance, thus suggesting a new pathway for manipulating stress tolerance in crop plants. The T0 transgenic plants showed high-levels of PDH45 protein in normal and stress conditions, as compared to wild type (WT) plants. The T0 transgenics also showed tolerance to high salinity as tested by a leaf disc senescence assay. The T1 transgenics were able to grow to maturity and set normal viable seeds under continuous salinity stress, without any reduction in plant yield, in terms of seed weight. Measurement of Na/sup +/ ions in different parts of the plant showed higher accumulation in the old leaves and negligible in seeds of T1 transgenic lines as compared with the WT plants. The possible mechanism of salinity tolerance will be discussed. Over-expression of PDH45 provides a possible example of the

  16. Human plasma phospholipid transfer protein increases the antiatherogenic potential of high density lipoproteins in transgenic mice

    NARCIS (Netherlands)

    M.J. van Haperen (Rien); A. van Tol (Arie); P. Vermeulen; M. Jauhiainen; T. van Gent (Teus); P.M. van den Berg (Paul); S. Ehnholm (Sonja); A.W.M. van der Kamp (Arthur); M.P.G. de Crom (Rini); F.G. Grosveld (Frank)

    2000-01-01

    textabstractPlasma phospholipid transfer protein (PLTP) transfers phospholipids between lipoprotein particles and alters high density lipoprotein (HDL) subfraction patterns in vitro, but its physiological function is poorly understood. Transgenic mice that overexpress

  17. Pathogenesis of axonal dystrophy and demyelination in alphaA-crystallin-expressing transgenic mice.

    NARCIS (Netherlands)

    Rijk, A. van; Sweers, M.A.; Merkx, G.F.M.; Lammens, M.M.Y.; Bloemendal, H.

    2003-01-01

    We recently described a transgenic mouse strain overexpressing hamster alphaA-crystallin, a small heat shock protein, under direction of the hamster vimentin promoter. As a result myelin was degraded and axonal dystrophy in both central nervous system (especially spinal cord) and peripheral nervous

  18. ThNAC13, a NAC Transcription Factor from Tamarix hispida, Confers Salt and Osmotic Stress Tolerance to Transgenic Tamarix and Arabidopsis

    Science.gov (United States)

    Wang, Liuqiang; Li, Zhen; Lu, Mengzhu; Wang, Yucheng

    2017-01-01

    NAC (NAM, ATAF1/2, and CUC2) proteins play critical roles in many plant biological processes and environmental stress. However, NAC proteins from Tamarix hispida have not been functionally characterized. Here, we studied a NAC gene from T. hispida, ThNAC13, in response to salt and osmotic stresses. ThNAC13 is a nuclear protein with a C-terminal transactivation domain. ThNAC13 can bind to NAC recognized sites and calmodulin-binding NAC (CBNAC) binding element. Overexpression of ThNAC13 in Arabidopsis improved seed germination rate and increased root growth and fresh weight gain under salt or osmotic stress. Transgenic T. hispida plants transiently overexpressing ThNAC13 and with RNAi-silenced ThNAC13 were generated for gain- and loss-of-function experiments. Following exposure to salt or osmotic stress, overexpression of ThNAC13 induced superoxide dismutase (SOD) and peroxidase (POD) activities, chlorophyll and proline contents; decreased the reactive oxygen species (ROS) and malondialdehyde levels; and reduced electrolyte leakage rates in both transgenic Tamarix and Arabidopsis plants. In contrast, RNAi-silenced ThNAC13 showed the opposite results in transgenic Tamarix. Furthermore, ThNAC13 induced the expression of SODs and PODs in transgenic Arabidopsis. These results suggest that ThNAC13 improves salt and osmotic tolerance by enhancing the ROS-scavenging capability and adjusting osmotic potential. PMID:28491072

  19. ThNAC13, a NAC Transcription Factor from Tamarix hispida, Confers Salt and Osmotic Stress Tolerance to Transgenic Tamarix and Arabidopsis

    Directory of Open Access Journals (Sweden)

    Mengzhu Lu

    2017-04-01

    Full Text Available NAC (NAM, ATAF1/2, and CUC2 proteins play critical roles in many plant biological processes and environmental stress. However, NAC proteins from Tamarix hispida have not been functionally characterized. Here, we studied a NAC gene from T. hispida, ThNAC13, in response to salt and osmotic stresses. ThNAC13 is a nuclear protein with a C-terminal transactivation domain. ThNAC13 can bind to NAC recognized sites and calmodulin-binding NAC (CBNAC binding element. Overexpression of ThNAC13 in Arabidopsis improved seed germination rate and increased root growth and fresh weight gain under salt or osmotic stress. Transgenic T. hispida plants transiently overexpressing ThNAC13 and with RNAi-silenced ThNAC13 were generated for gain- and loss-of-function experiments. Following exposure to salt or osmotic stress, overexpression of ThNAC13 induced superoxide dismutase (SOD and peroxidase (POD activities, chlorophyll and proline contents; decreased the reactive oxygen species (ROS and malondialdehyde levels; and reduced electrolyte leakage rates in both transgenic Tamarix and Arabidopsis plants. In contrast, RNAi-silenced ThNAC13 showed the opposite results in transgenic Tamarix. Furthermore, ThNAC13 induced the expression of SODs and PODs in transgenic Arabidopsis. These results suggest that ThNAC13 improves salt and osmotic tolerance by enhancing the ROS-scavenging capability and adjusting osmotic potential.

  20. High level expression of Acidothermus cellulolyticus β-1, 4-endoglucanase in transgenic rice enhances the hydrolysis of its straw by cultured cow gastric fluid

    Energy Technology Data Exchange (ETDEWEB)

    Chou, Hong L.; Dai, Ziyu; Hsieh, Chia W.; Ku, Maurice S.

    2011-12-10

    Large-scale production of effective cellulose hydrolytic enzymes is the key to the bioconversion of agricultural residues to ethanol. The goal of this study was to develop a rice plant as a bioreactor for the large-scale production of cellulose hydrolytic enzymes via genetic transformation, and to simultaneously improve rice straw as an efficient biomass feedstock for conversion of cellulose to glucose. In this study, the cellulose hydrolytic enzyme {beta}-1, 4-endoglucanase (E1) from the thermophilic bacterium Acidothermus cellulolyticus was overexpressed in rice through Agrobacterium-mediated transformation. The expression of the bacterial gene in rice was driven by the constitutive Mac promoter, a hybrid promoter of Ti plasmid mannopine synthetase promoter and cauliflower mosaic virus 35S promoter enhancer with the signal peptide of tobacco pathogenesis-related protein for targeting the protein to the apoplastic compartment for storage. A total of 52 transgenic rice plants from six independent lines expressing the bacterial enzyme were obtained, which expressed the gene at high levels with a normal phenotype. The specific activities of E1 in the leaves of the highest expressing transgenic rice lines were about 20 fold higher than those of various transgenic plants obtained in previous studies and the protein amounts accounted for up to 6.1% of the total leaf soluble protein. Zymogram and temperature-dependent activity analyses demonstrated the thermostability of the enzyme and its substrate specificity against cellulose, and a simple heat treatment can be used to purify the protein. In addition, hydrolysis of transgenic rice straw with cultured cow gastric fluid yielded almost twice more reducing sugars than wild type straw. Taken together, these data suggest that transgenic rice can effectively serve as a bioreactor for large-scale production of active, thermostable cellulose hydrolytic enzymes. As a feedstock, direct expression of large amount of cellulases in

  1. Seed-specific increased expression of 2S albumin promoter of sesame qualifies it as a useful genetic tool for fatty acid metabolic engineering and related transgenic intervention in sesame and other oil seed crops.

    Science.gov (United States)

    Bhunia, Rupam Kumar; Chakraborty, Anirban; Kaur, Ranjeet; Gayatri, T; Bhattacharyya, Jagannath; Basu, Asitava; Maiti, Mrinal K; Sen, Soumitra Kumar

    2014-11-01

    The sesame 2S albumin (2Salb) promoter was evaluated for its capacity to express the reporter gusA gene encoding β-glucuronidase in transgenic tobacco seeds relative to the soybean fad3C gene promoter element. Results revealed increased expression of gusA gene in tobacco seed tissue when driven by sesame 2S albumin promoter. Prediction based deletion analysis of both the promoter elements confirmed the necessary cis-acting regulatory elements as well as the minimal promoter element for optimal expression in each case. The results also revealed that cis-regulatory elements might have been responsible for high level expression as well as spatio-temporal regulation of the sesame 2S albumin promoter. Transgenic over-expression of a fatty acid desaturase (fad3C) gene of soybean driven by 2S albumin promoter resulted in seed-specific enhanced level of α-linolenic acid in sesame. The present study, for the first time helped to identify that the sesame 2S albumin promoter is a promising endogenous genetic element in genetic engineering approaches requiring spatio-temporal regulation of gene(s) of interest in sesame and can also be useful as a heterologous genetic element in other important oil seed crop plants in general for which seed oil is the harvested product. The study also established the feasibility of fatty acid metabolic engineering strategy undertaken to improve quality of edible seed oil in sesame using the 2S albumin promoter as regulatory element.

  2. Overexpression of HvIcy6 in Barley Enhances Resistance against Tetranychus urticae and Entails Partial Transcriptomic Reprogramming

    Directory of Open Access Journals (Sweden)

    M. Estrella Santamaria

    2018-03-01

    Full Text Available Cystatins have been largely used for pest control against phytophagous species. However, cystatins have not been commonly overexpressed in its cognate plant species to test their pesticide capacity. Since the inhibitory role of barley HvCPI-6 cystatin against the phytophagous mite Tetranychus urticae has been previously demonstrated, the purpose of our study was to determine if barley transgenic lines overexpressing its own HvIcy6 gene were more resistant against this phytophagous infestation. Besides, a transcriptomic analysis was done to find differential expressed genes among wild-type and transformed barley plants. Barley plants overexpressing HvIcy6 cystatin gene remained less susceptible to T. urticae attack when compared to wild-type plants, with a significant lesser foliar damaged area and a lower presence of the mite. Transcriptomic analysis revealed a certain reprogramming of cellular metabolism and a lower expression of several genes related to photosynthetic activity. Therefore, although caution should be taken to discard potential deleterious pleiotropic effects, cystatins may be used as transgenes with impact on agricultural crops by conferring enhanced levels of resistance to phytophagous pests.

  3. Enhanced Androgen Signaling With Androgen Receptor Overexpression in the Osteoblast Lineage Controls Skeletal Turnover, Matrix Quality and Bone Architecture

    National Research Council Canada - National Science Library

    Wiren, Kristine M; Jepsen, Karl

    2006-01-01

    .... We genetically engineered transgenic mice in which androgen receptor (AR) overexpression is skeletally targeted in two separate models to better understand the role of androgen signaling directly in bone...

  4. Dek overexpression in murine epithelia increases overt esophageal squamous cell carcinoma incidence

    Science.gov (United States)

    Cimperman, Katherine A.; Haas, Sarah R.; Guasch, Geraldine; Waclaw, Ronald R.; Komurov, Kakajan; Lane, Adam; Wikenheiser-Brokamp, Kathryn A.

    2018-01-01

    Esophageal cancer occurs as either squamous cell carcinoma (ESCC) or adenocarcinoma. ESCCs comprise almost 90% of cases worldwide, and recur with a less than 15% five-year survival rate despite available treatments. The identification of new ESCC drivers and therapeutic targets is critical for improving outcomes. Here we report that expression of the human DEK oncogene is strongly upregulated in esophageal SCC based on data in the cancer genome atlas (TCGA). DEK is a chromatin-associated protein with important roles in several nuclear processes including gene transcription, epigenetics, and DNA repair. Our previous data have utilized a murine knockout model to demonstrate that Dek expression is required for oral and esophageal SCC growth. Also, DEK overexpression in human keratinocytes, the cell of origin for SCC, was sufficient to cause hyperplasia in 3D organotypic raft cultures that mimic human skin, thus linking high DEK expression in keratinocytes to oncogenic phenotypes. However, the role of DEK over-expression in ESCC development remains unknown in human cells or genetic mouse models. To define the consequences of Dek overexpression in vivo, we generated and validated a tetracycline responsive Dek transgenic mouse model referred to as Bi-L-Dek. Dek overexpression was induced in the basal keratinocytes of stratified squamous epithelium by crossing Bi-L-Dek mice to keratin 5 tetracycline transactivator (K5-tTA) mice. Conditional transgene expression was validated in the resulting Bi-L-Dek_K5-tTA mice and was suppressed with doxycycline treatment in the tetracycline-off system. The mice were subjected to an established HNSCC and esophageal carcinogenesis protocol using the chemical carcinogen 4-nitroquinoline 1-oxide (4NQO). Dek overexpression stimulated gross esophageal tumor development, when compared to doxycycline treated control mice. Furthermore, high Dek expression caused a trend toward esophageal hyperplasia in 4NQO treated mice. Taken together, these

  5. Increased Melanoma Growth and Metastasis Spreading in Mice Overexpressing Placenta Growth Factor

    Science.gov (United States)

    Marcellini, Marcella; De Luca, Naomi; Riccioni, Teresa; Ciucci, Alessandro; Orecchia, Angela; Lacal, Pedro Miguel; Ruffini, Federica; Pesce, Maurizio; Cianfarani, Francesca; Zambruno, Giovanna; Orlandi, Augusto; Failla, Cristina Maria

    2006-01-01

    Placenta growth factor (PlGF), a member of the vascular endothelial growth factor family, plays an important role in adult pathological angiogenesis. To further investigate PlGF functions in tumor growth and metastasis formation, we used transgenic mice overexpressing PlGF in the skin under the control of the keratin 14 promoter. These animals showed a hypervascularized phenotype of the skin and increased levels of circulating PlGF with respect to their wild-type littermates. Transgenic mice and controls were inoculated intradermally with B16-BL6 melanoma cells. The tumor growth rate was fivefold increased in transgenic animals compared to wild-type mice, in the presence of a similar percentage of tumor necrotic tissue. Tumor vessel area was increased in transgenic mice as compared to controls. Augmented mobilization of endothelial and hematopoietic stem cells from the bone marrow was observed in transgenic animals, possibly contributing to tumor vascularization. The number and size of pulmonary metastases were significantly higher in transgenic mice compared to wild-type littermates. Finally, PlGF promoted tumor cell invasion of the extracellular matrix and increased the activity of selected matrix metalloproteinases. These findings indicate that PlGF, in addition to enhancing tumor angiogenesis and favoring tumor growth, may directly influence melanoma dissemination. PMID:16877362

  6. A Mutation of the Prdm9 Mouse Hybrid Sterility Gene Carried by a Transgene.

    Science.gov (United States)

    Mihola, O; Trachtulec, Z

    2017-01-01

    PRDM9 is a protein with histone-3-methyltransferase activity, which specifies the sites of meiotic recombination in mammals. Deficiency of the Prdm9 gene in the laboratory mouse results in complete arrest of the meiotic prophase of both sexes. Moreover, the combination of certain PRDM9 alleles from different mouse subspecies causes hybrid sterility, e.g., the male-specific meiotic arrest found in the (PWD/Ph × C57BL/6J)F1 animals. The fertility of all these mice can be rescued using a Prdm9-containing transgene. Here we characterized a transgene made from the clone RP24-346I22 that was expected to encompass the entire Prdm9 gene. Both (PWD/Ph × C57BL/6J)F1 intersubspecific hybrid males and Prdm9-deficient laboratory mice of both sexes carrying this transgene remained sterile, suggesting that Prdm9 inactivation occurred in the Tg(RP24-346I22) transgenics. Indeed, comparative qRT-PCR analysis of testicular RNAs from transgene-positive versus negative animals revealed similar expression levels of Prdm9 mRNAs from the exons encoding the C-terminal part of the protein but elevated expression from the regions coding for the N-terminus of PRDM9, indicating that the transgenic carries a new null Prdm9 allele. Two naturally occurring alternative Prdm9 mRNA isoforms were overexpressed in Tg(RP24-346I22), one formed via splicing to a 3'-terminal exon consisting of short interspersed element B2 and one isoform including an alternative internal exon of 28 base pairs. However, the overexpression of these alternative transcripts was apparently insufficient for Prdm9 function or for increasing the fertility of the hybrid males.

  7. Overexpression of human and fly frataxins in Drosophila provokes deleterious effects at biochemical, physiological and developmental levels.

    Directory of Open Access Journals (Sweden)

    Juan A Navarro

    Full Text Available BACKGROUND: Friedreich's ataxia (FA, the most frequent form of inherited ataxias in the Caucasian population, is caused by a reduced expression of frataxin, a highly conserved protein. Model organisms have contributed greatly in the efforts to decipher the function of frataxin; however, the precise function of this protein remains elusive. Overexpression studies are a useful approach to investigate the mechanistic actions of frataxin; however, the existing literature reports contradictory results. To further investigate the effect of frataxin overexpression, we analyzed the consequences of overexpressing human (FXN and fly (FH frataxins in Drosophila. METHODOLOGY/PRINCIPAL FINDINGS: We obtained transgenic flies that overexpressed human or fly frataxins in a general pattern and in different tissues using the UAS-GAL4 system. For both frataxins, we observed deleterious effects at the biochemical, histological and behavioral levels. Oxidative stress is a relevant factor in the frataxin overexpression phenotypes. Systemic frataxin overexpression reduces Drosophila viability and impairs the normal embryonic development of muscle and the peripheral nervous system. A reduction in the level of aconitase activity and a decrease in the level of NDUF3 were also observed in the transgenic flies that overexpressed frataxin. Frataxin overexpression in the nervous system reduces life span, impairs locomotor ability and causes brain degeneration. Frataxin aggregation and a misfolding of this protein have been shown not to be the mechanism that is responsible for the phenotypes that have been observed. Nevertheless, the expression of human frataxin rescues the aconitase activity in the fh knockdown mutant. CONCLUSION/SIGNIFICANCE: Our results provide in vivo evidence of a functional equivalence for human and fly frataxins and indicate that the control of frataxin expression is important for treatments that aim to increase frataxin levels.

  8. Overexpression of mouse TTF-2 gene causes cleft palate

    Science.gov (United States)

    Meng, Tian; Shi, Jia-Yu; Wu, Min; Wang, Yan; Li, Ling; Liu, Yan; Zheng, Qian; Huang, Lei; Shi, Bing

    2012-01-01

    In humans, mutations of the gene encoding for thyroid transcription factor-2 (TTF-2 or FOXE1) result in Bamforth syndrome. Bamforth syndrome is characterized by agenesis, cleft palate, spiky hair and choanal atresia. TTF-2 null mice (TTF-2−/−) also exhibit cleft palate, suggesting its involvement in the palatogenesis. However, the molecular pathology and genetic regulation by TTF2 remain largely unknown. In the present study, the recombinant expression vector pBROAD3-TTF-2 containing the promoter of the mouse ROSA26 gene was created to form the structural gene of mouse TTF-2 and was microinjected into the male pronuclei of fertilized ova. Sequence analysis confirmed that the TTF-2 transgenic mouse model was established successfully. The transgenic mice displayed a phenotype of cleft palate. In addition, we found that TTF-2 was highly expressed in the medial edge epithelium (MEE) from the embryonic day 12.5 (E12.5) to E14.5 in TTF-2 transgenic mice. These observations suggest that overexpression of TTF-2 during palatogenesis may contribute to formation of cleft palate. PMID:22304410

  9. Inducible transgenics. New lessons on events governing the induction and commitment in mammary tumorigenesis

    International Nuclear Information System (INIS)

    Hulit, James; Di Vizio, Dolores; Pestell, Richard G

    2001-01-01

    Breast cancer arises from multiple genetic events that together contribute to the established, irreversible malignant phenotype. The development of inducible tissue-specific transgenics has allowed a careful dissection of the events required for induction and subsequent maintenance of tumorigenesis. Mammary gland targeted expression of oncogenic Ras or c-Myc is sufficient for the induction of mammary gland tumorigenesis in the rodent, and when overexpressed together the rate of tumor onset is substantially enhanced. In an exciting recent finding, D'Cruz et al discovered tetracycline-regulated c-Myc overexpression in the mammary gland induced invasive mammary tumors that regressed upon withdrawal of c-Myc expression. Almost one-half of the c-Myc-induced tumors harbored K-ras or N-ras gene point mutations, correlating with tumor persistence on withdrawal of c-Myc transgene expression. These findings suggest maintenance of tumorigenesis may involve a second mutation within the Ras pathway

  10. Epidermal dysplasia and abnormal hair follicles in transgenic mice overexpressing homeobox gene MSX-2.

    Science.gov (United States)

    Jiang, T X; Liu, Y H; Widelitz, R B; Kundu, R K; Maxson, R E; Chuong, C M

    1999-08-01

    The homeobox gene Msx-2 is expressed specifically in sites of skin appendage formation. To explore its part in skin morphogenesis, we produced transgenic mice expressing Msx-2 under the control of the cytomegalovirus promoter. The skin of these transgenic mice was flaky, exhibiting desquamation and shorter hairs. Histologic analysis showed thickened epidermis with hyperproliferation, which was restricted to the basal layer. Hyperkeratosis was also evident. A wide zone of suprabasal cells were misaligned and coexpressed keratins 14 and 10. There was reduced expression of integrin beta 1 and DCC in the basal layer. Hair follicles were misaligned with a shrunken matrix region. The dermis showed increased cellularity and empty vacuoles. We suggest that Msx-2 is involved in the growth control of skin and skin appendages.

  11. The role of the F-box gene TaFBA1 from wheat (Triticum aestivum L.) in drought tolerance.

    Science.gov (United States)

    Zhou, Shumei; Sun, Xiudong; Yin, Suhong; Kong, Xiangzhu; Zhou, Shan; Xu, Ying; Luo, Yin; Wang, Wei

    2014-11-01

    Drought is one of the most important factors limiting plant growth and development. We identified a gene in wheat (Triticum aestivum L.) under drought stress named TaFBA1. TaFBA1 encodes a putative 325-amino-acid F-box protein with a conserved N-terminal F-box domain and a C-terminal AMN1 domain. Real-time RT-PCR analysis revealed that TaFBA1 transcript accumulation was upregulated by high-salinity, water stress, and abscisic acid (ABA) treatment. To evaluate the functions of TaFBA1 in the regulation of drought stress responses, we produced transgenic tobacco lines overexpressing TaFBA1. Under water stress conditions, the transgenic tobacco plants had a higher germination rate, higher relative water content, net photosynthesis rate (Pn), less chlorophyll loss, and less growth inhibition than WT. These results demonstrate the high tolerance of the transgenic plants to drought stress compared to the WT. The enhanced oxidative stress tolerance of these plants, which may be involved in their drought tolerance, was indicated by their lower levels of reactive oxygen species (ROS) accumulation, MDA content, and cell membrane damage under drought stress compared to WT. The antioxidant enzyme activities were higher in the transgenic plants than in WT, which may be related to the upregulated expression of some antioxidant genes via overexpression of TaFBA1. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  12. Analysis of transgenic wheat (Triticum aestivum L.) harboring a maize (Zea mays L.) gene for plastid EF-Tu: segregation pattern, expression and effects of the transgene.

    Science.gov (United States)

    Fu, Jianming; Ristic, Zoran

    2010-06-01

    We previously reported that transgenic wheat (Triticum aestivum L.) carrying a maize (Zea mays L.) gene (Zmeftu1) for chloroplast protein synthesis elongation factor, EF-Tu, displays reduced thermal aggregation of leaf proteins, reduced injury to photosynthetic membranes (thylakoids), and enhanced rate of CO(2) fixation following exposure to heat stress (18 h at 45 degrees C) [Fu et al. in Plant Mol Biol 68:277-288, 2008]. In the current study, we investigated the segregation pattern and expression of the transgene Zmeftu1 and determined the grain yield of transgenic plants after exposure to a brief heat stress (18 h at 45 degrees C). We also assessed thermal aggregation of soluble leaf proteins in transgenic plants, testing the hypothesis that increased levels of EF-Tu will lead to a non-specific protection of leaf proteins against thermal aggregation. The transgenic wheat displayed a single-gene pattern of segregation of Zmeftu1. Zmeftu1 was expressed, and the transgenic plants synthesized and accumulated three anti-EF-Tu cross-reacting polypeptides of similar molecular mass but different pI, suggesting the possibility of posttranslational modification of this protein. The transgenic plants also showed better grain yield after exposure to heat stress compared with their non-transgenic counterparts. Soluble leaf proteins of various molecular masses displayed lower thermal aggregation in transgenic than in non-transgenic wheat. The results suggest that overexpression of chloroplast EF-Tu can be beneficial to wheat tolerance to heat stress. Moreover, the results also support the hypothesis that EF-Tu contributes to heat tolerance by acting as a molecular chaperone and protecting heat-labile proteins from thermal aggregation in a non-specific manner.

  13. Construction and Quality Analysis of Transgenic Rehmannia glutinosa Containing TMV and CMV Coat Protein

    Directory of Open Access Journals (Sweden)

    Zhongqiu Teng

    2016-08-01

    Full Text Available Plant viruses, especially tobacco mosaic virus (TMV and cucumber mosaic virus (CMV are serious threats to Rehmannia glutinosa which is a “top grade” herb in China. In the present study, TMV- and CMV-resistant Rehmannia glutinosa Libosch. plants were constructed by transforming the protein (CP genes of TMV and CMV into Rehmannia glutinosa via a modified procedure of Agrobacterium tumefaciens-mediated transformation. Integration and expression of TMV CP and CMV CP transgenes in 2 lines, LBA-1 and LBA-2, were confirmed by PCR, Southern blot and RT-PCR. Both LBA-1 and LBA-2 were resistant to infection of homologous TMV and CMV strains. The quality of transgenic Rehmanniae Radix was evaluated based on fingerprint analysis and components quantitative analysis comparing with control root tubes. These results showed that chemical composition of transgenic Rehmanniae Radix were similar to non-transgenic ones, which demonstrated that the medical quality and biosafety of transgenic Rehmanniae Radix were equivalent to non-transgenic material when consumed as traditional Chinese medicinal (TCM.

  14. A wheat calreticulin gene (TaCRT1) contributes to drought tolerance in transgenic arabidopsis

    International Nuclear Information System (INIS)

    Xiang, V.; Du, C.; Jia, H.; Song, M.; Wang, Y.; Ma, Z.

    2018-01-01

    The TaCRT1 gene is a member of calreticulin (CRT) family in wheat. In our previous study, we showed that transgenic tobacco lines over expressing wheat TaCRT1 showed enhanced tolerance to salt stress. This study aimed to determine whether TaCRT1 over expression would increase drought tolerance in transgenic Arabidopsis. Over expression of TaCRT1 in Arabidopsis plants enhances tolerance to drought stress. However, the transgenic line was found to retard the growth. Moreover, the transgenic line showed decreased water loss but higher sensitivity to exogenous abscisic acid (ABA) compared with the wild type (Col-0). Meanwhile, the transgenic line had the elevated endogenous ABA level. The semi-quantitative RT-PCR (sqRT-PCR) analysis showed that transcription levels of ABA-biosynthesizing gene (NCED3) and ABA-responsive gene (ABF3) were higher in the transgenic line than that in the Col-0 under normal condition. The above results implied that the TaCRT1 might be able to used as a potential target to improve the drought tolerance in crops. (author)

  15. Astroglial c-Myc overexpression predisposes mice to primary malignant gliomas

    DEFF Research Database (Denmark)

    Jensen, Niels Aagaard; Pedersen, Karen-Marie; Lihme, Frederikke

    2003-01-01

    Malignant astrocytomas are common human primary brain tumors that result from neoplastic transformation of astroglia or their progenitors. Here we show that deregulation of the c-Myc pathway in developing astroglia predisposes mice to malignant astrocytomas within 2-3 weeks of age. The genetically...... engineered murine (GEM) gliomas harbor a molecular signature resembling that of human primary glioblastoma multiforme, including up-regulation of epidermal growth factor receptor and Mdm2. The GEM gliomas seem to originate in an abnormal population of glial fibrillary acidic protein-expressing cells...... the neoplastic process, presumably by inducing the sustained growth of early astroglial cells. This is in contrast to most other transgenic studies in which c-Myc overexpression requires co-operating transgenes for rapid tumor induction....

  16. Cardiac cell therapy: overexpression of connexin43 in skeletal myoblasts and prevention of ventricular arrhythmias

    NARCIS (Netherlands)

    Fernandes, Sarah; van Rijen, Harold V. M.; Forest, Virginie; Evain, Stéphane; Leblond, Anne-Laure; Mérot, Jean; Charpentier, Flavien; de Bakker, Jacques M. T.; Lemarchand, Patricia

    2009-01-01

    Cell-based therapies have great potential for the treatment of cardiovascular diseases. Recently, using a transgenic mouse model Roell et al. reported that cardiac engraftment of connexin43 (Cx43)-overexpressing myoblasts in vivo prevents post-infarct arrhythmia, a common cause of death in patients

  17. Overexpression of Rat Neurons Nitric Oxide Synthase in Rice Enhances Drought and Salt Tolerance.

    Directory of Open Access Journals (Sweden)

    Wei Cai

    Full Text Available Nitric oxide (NO has been shown to play an important role in the plant response to biotic and abiotic stresses in Arabidopsis mutants with lower or higher levels of endogenous NO. The exogenous application of NO donors or scavengers has also suggested an important role for NO in plant defense against environmental stress. In this study, rice plants under drought and high salinity conditions showed increased nitric oxide synthase (NOS activity and NO levels. Overexpression of rat neuronal NO synthase (nNOS in rice increased both NOS activity and NO accumulation, resulting in improved tolerance of the transgenic plants to both drought and salt stresses. nNOS-overexpressing plants exhibited stronger water-holding capability, higher proline accumulation, less lipid peroxidation and reduced electrolyte leakage under drought and salt conditions than wild rice. Moreover, nNOS-overexpressing plants accumulated less H2O2, due to the observed up-regulation of OsCATA, OsCATB and OsPOX1. In agreement, the activities of CAT and POX were higher in transgenic rice than wild type. Additionally, the expression of six tested stress-responsive genes including OsDREB2A, OsDREB2B, OsSNAC1, OsSNAC2, OsLEA3 and OsRD29A, in nNOS-overexpressing plants was higher than that in the wild type under drought and high salinity conditions. Taken together, our results suggest that nNOS overexpression suppresses the stress-enhanced electrolyte leakage, lipid peroxidation and H2O2 accumulation, and promotes proline accumulation and the expression of stress-responsive genes under stress conditions, thereby promoting increased tolerance to drought and salt stresses.

  18. Tumorigenic potential of pituitary tumor transforming gene (PTTG in vivo investigated using a transgenic mouse model, and effects of cross breeding with p53 (+/− transgenic mice

    Directory of Open Access Journals (Sweden)

    Fong Miranda Y

    2012-11-01

    Full Text Available Abstract Background Pituitary tumor-transforming gene (PTTG is an oncogene that is overexpressed in variety of tumors and exhibits characteristics of a transforming gene. Previous transgenic mouse models to access the tumorigenic potential in the pituitary and ovary have resulted in dysplasia without formation of visible tumors, possibly due to the insufficient expression of PTTG. PTTG expression level is critical for ovarian tumorigenesis in a xenograft model. Therefore, the tumorigenic function of PTTG in vivo remains unclear. We generated a transgenic mouse that overexpresses PTTG driven by the CMV promoter to determine whether PTTG functions as a transforming oncogene that is capable of initiating tumorigenesis. Methods Transgenic animals were generated by microinjection of PTTG transgene into the male pronucleus of FVB 0.5 day old embryos. Expression levels of PTTG in tissues of transgenic animals were analyzed using an immunohistochemical analysis. H&E staining and immunohistostaining were performed to examine the type of tumor in transgenic and PTTG transgenic/p53+/- animals. Results PTTG transgenic offspring (TgPTTG were monitored for tumor development at various ages. H&E analysis was performed to identify the presence of cancer and hyperplastic conditions verified with the proliferation marker PCNA and the microvessel marker CD31. Immunohistochemistry was performed to determine transgene expression, revealing localization to the epithelium of the fallopian tube, with more generalized expression in the liver, lung, kidney, and spleen. At eight months of age, 2 out of 15 TgPTTG developed ovarian cancer, 2 out of 15 developed benign tumors, 2 out of 15 developed cervical dysplasia, and 3 out of 15 developed adenomyosis of the uterus. At ten months of age, 2 out of 10 TgPTTG developed adenocarcinoma of the ovary, 1 out of 10 developed a papillary serous adenocarcinoma, and 2 out of 10 presented with atypia of ovarian epithelial cells

  19. Overexpression of Soybean Isoflavone Reductase (GmIFR) Enhances Resistance to Phytophthora sojae in Soybean.

    Science.gov (United States)

    Cheng, Qun; Li, Ninghui; Dong, Lidong; Zhang, Dayong; Fan, Sujie; Jiang, Liangyu; Wang, Xin; Xu, Pengfei; Zhang, Shuzhen

    2015-01-01

    Isoflavone reductase (IFR) is an enzyme involved in the biosynthetic pathway of isoflavonoid phytoalexin in plants. IFRs are unique to the plant kingdom and are considered to have crucial roles in plant response to various biotic and abiotic environmental stresses. Here, we report the characterization of a novel member of the soybean isoflavone reductase gene family GmIFR. Overexpression of GmIFR transgenic soybean exhibited enhanced resistance to Phytophthora sojae. Following stress treatments, GmIFR was significantly induced by P. sojae, ethephon (ET), abscisic acid (placeCityABA), salicylic acid (SA). It is located in the cytoplasm when transiently expressed in soybean protoplasts. The daidzein levels reduced greatly for the seeds of transgenic plants, while the relative content of glyceollins in transgenic plants was significantly higher than that of non-transgenic plants. Furthermore, we found that the relative expression levels of reactive oxygen species (ROS) of transgenic soybean plants were significantly lower than those of non-transgenic plants after incubation with P. sojae, suggesting an important role of GmIFR might function as an antioxidant to reduce ROS in soybean. The enzyme activity assay suggested that GmIFR has isoflavone reductase activity.

  20. Overexpression of Three Glucosinolate Biosynthesis Genes in Brassica napus Identifies Enhanced Resistance to Sclerotinia sclerotiorum and Botrytis cinerea.

    Directory of Open Access Journals (Sweden)

    Yuanyuan Zhang

    Full Text Available Sclerotinia sclerotiorum and Botrytis cinerea are notorious plant pathogenic fungi with an extensive host range including Brassica crops. Glucosinolates (GSLs are an important group of secondary metabolites characteristic of the Brassicales order, whose degradation products are proving to be increasingly important in plant protection. Enhancing the defense effect of GSL and their associated degradation products is an attractive strategy to strengthen the resistance of plants by transgenic approaches. We generated the lines of Brassica napus with three biosynthesis genes involved in GSL metabolic pathway (BnMAM1, BnCYP83A1 and BnUGT74B1, respectively. We then measured the foliar GSLs of each transgenic lines and inoculated them with S. sclerotiorum and B. cinerea. Compared with the wild type control, over-expressing BnUGT74B1 in B. napus increased the aliphatic and indolic GSL levels by 1.7 and 1.5 folds in leaves respectively; while over-expressing BnMAM1 or BnCYP83A1 resulted in an approximate 1.5-fold higher only in the aliphatic GSL level in leaves. The results of plant inoculation demonstrated that BnUGT74B1-overexpressing lines showed less severe disease symptoms and tissue damage compared with the wild type control, but BnMAM1 or BnCYP83A1-overexpressing lines showed no significant difference in comparison to the controls. These results suggest that the resistance to S. sclerotiorum and B. cinerea in B. napus could be enhanced through tailoring the GSL profiles by transgenic approaches or molecular breeding, which provides useful information to assist plant breeders to design improved breeding strategies.

  1. Lysyl oxidase overexpression accelerates cardiac remodeling and aggravates angiotensin II-induced hypertrophy.

    Science.gov (United States)

    Galán, María; Varona, Saray; Guadall, Anna; Orriols, Mar; Navas, Miquel; Aguiló, Silvia; de Diego, Alicia; Navarro, María A; García-Dorado, David; Rodríguez-Sinovas, Antonio; Martínez-González, José; Rodriguez, Cristina

    2017-09-01

    Lysyl oxidase (LOX) controls matrix remodeling, a key process that underlies cardiovascular diseases and heart failure; however, a lack of suitable animal models has limited our knowledge with regard to the contribution of LOX to cardiac dysfunction. Here, we assessed the impact of LOX overexpression on ventricular function and cardiac hypertrophy in a transgenic LOX (TgLOX) mouse model with a strong cardiac expression of human LOX. TgLOX mice exhibited high expression of the transgene in cardiomyocytes and cardiofibroblasts, which are associated with enhanced LOX activity and H 2 O 2 production and with cardiofibroblast reprogramming. LOX overexpression promoted an age-associated concentric remodeling of the left ventricle and impaired diastolic function. Furthermore, LOX transgenesis aggravated angiotensin II (Ang II)-induced cardiac hypertrophy and dysfunction, which triggered a greater fibrotic response that was characterized by stronger collagen deposition and cross-linking and high expression of fibrotic markers. In addition, LOX transgenesis increased the Ang II-induced myocardial inflammatory infiltrate, exacerbated expression of proinflammatory markers, and decreased that of cardioprotective factors. Mechanistically, LOX overexpression enhanced oxidative stress and potentiated the Ang II-mediated cardiac activation of p38 MAPK while reducing AMPK activation. Our findings suggest that LOX induces an age-dependent disturbance of diastolic function and aggravates Ang II-induced hypertrophy, which provides novel insights into the role of LOX in cardiac performance.-Galán, M., Varona, S., Guadall, A., Orriols, M., Navas, M., Aguiló, S., de Diego, A., Navarro, M. A., García-Dorado, D., Rodríguez-Sinovas, A., Martínez-González, J., Rodriguez, C. Lysyl oxidase overexpression accelerates cardiac remodeling and aggravates angiotensin II-induced hypertrophy. © FASEB.

  2. Transgenic tobacco plants expressing BoRS1 gene from Brassica ...

    Indian Academy of Sciences (India)

    Water stress is by far the leading environmental stress limiting crop yields worldwide. Genetic engineering techniques hold great promise for developing crop cultivars with high tolerance to water stress. In this study, the Brassica oleracea var. acephala BoRS1 gene was transferred into tobacco through ...

  3. Transgenic Mice Over-Expressing RBP4 Have RBP4-Dependent and Light-Independent Retinal Degeneration.

    Science.gov (United States)

    Du, Mei; Phelps, Eric; Balangue, Michael J; Dockins, Aaron; Moiseyev, Gennadiy; Shin, Younghwa; Kane, Shelley; Otalora, Laura; Ma, Jian-Xing; Farjo, Rafal; Farjo, Krysten M

    2017-08-01

    Transgenic mice overexpressing serum retinol-binding protein (RBP4-Tg) develop progressive retinal degeneration, characterized by microglia activation, yet the precise mechanisms underlying retinal degeneration are unclear. Previous studies showed RBP4-Tg mice have normal ocular retinoid levels, suggesting that degeneration is independent of the retinoid visual cycle or light exposure. The present study addresses whether retinal degeneration is light-dependent and RBP4-dependent by testing the effects of dark-rearing and pharmacological lowering of serum RBP4 levels, respectively. RBP4-Tg mice reared on normal mouse chow in normal cyclic light conditions were directly compared to RBP4-Tg mice exposed to chow supplemented with the RBP4-lowering compound A1120 or dark-rearing conditions. Quantitative retinal histological analysis was conducted to assess retinal degeneration, and electroretinography (ERG) and optokinetic tracking (OKT) tests were performed to assess retinal and visual function. Ocular retinoids and bis-retinoid A2E were quantified. Dark-rearing RBP4-Tg mice effectively reduced ocular bis-retinoid A2E levels, but had no significant effect on retinal degeneration or dysfunction in RBP4-Tg mice, demonstrating that retinal degeneration is light-independent. A1120 treatment lowered serum RBP4 levels similar to wild-type mice, and prevented structural retinal degeneration. However, A1120 treatment did not prevent retinal dysfunction in RBP4-Tg mice. Moreover, RBP4-Tg mice on A1120 diet had significant worsening of OKT response and loss of cone photoreceptors compared to RBP4-Tg mice on normal chow. This may be related to the very significant reduction in retinyl ester levels in the retina of mice on A1120-supplemented diet. Retinal degeneration in RBP4-Tg mice is RBP4-dependent and light-independent.

  4. Assessment of cultivation factors that affect biomass and geraniol production in transgenic tobacco cell suspension cultures.

    Directory of Open Access Journals (Sweden)

    Nikolay Vasilev

    Full Text Available A large-scale statistical experimental design was used to determine essential cultivation parameters that affect biomass accumulation and geraniol production in transgenic tobacco (Nicotiana tabacum cv. Samsun NN cell suspension cultures. The carbohydrate source played a major role in determining the geraniol yield and factors such as filling volume, inoculum size and light were less important. Sucrose, filling volume and inoculum size had a positive effect on geraniol yield by boosting growth of plant cell cultures whereas illumination of the cultures stimulated the geraniol biosynthesis. We also found that the carbohydrates sucrose and mannitol showed polarizing effects on biomass and geraniol accumulation. Factors such as shaking frequency, the presence of conditioned medium and solubilizers had minor influence on both plant cell growth and geraniol content. When cells were cultivated under the screened conditions for all the investigated factors, the cultures produced ∼ 5.2 mg/l geraniol after 12 days of cultivation in shaking flasks which is comparable to the yield obtained in microbial expression systems. Our data suggest that industrial experimental designs based on orthogonal arrays are suitable for the selection of initial cultivation parameters prior to the essential medium optimization steps. Such designs are particularly beneficial in the early optimization steps when many factors must be screened, increasing the statistical power of the experiments without increasing the demand on time and resources.

  5. An Intergenic Region Shared by At4g35985 and At4g35987 in Arabidopsis thaliana Is a Tissue Specific and Stress Inducible Bidirectional Promoter Analyzed in Transgenic Arabidopsis and Tobacco Plants

    Science.gov (United States)

    Banerjee, Joydeep; Sahoo, Dipak Kumar; Dey, Nrisingha; Houtz, Robert L.; Maiti, Indu Bhushan

    2013-01-01

    On chromosome 4 in the Arabidopsis genome, two neighboring genes (calmodulin methyl transferase At4g35987 and senescence associated gene At4g35985) are located in a head-to-head divergent orientation sharing a putative bidirectional promoter. This 1258 bp intergenic region contains a number of environmental stress responsive and tissue specific cis-regulatory elements. Transcript analysis of At4g35985 and At4g35987 genes by quantitative real time PCR showed tissue specific and stress inducible expression profiles. We tested the bidirectional promoter-function of the intergenic region shared by the divergent genes At4g35985 and At4g35987 using two reporter genes (GFP and GUS) in both orientations in transient tobacco protoplast and Agro-infiltration assays, as well as in stably transformed transgenic Arabidopsis and tobacco plants. In transient assays with GFP and GUS reporter genes the At4g35985 promoter (P85) showed stronger expression (about 3.5 fold) compared to the At4g35987 promoter (P87). The tissue specific as well as stress responsive functional nature of the bidirectional promoter was evaluated in independent transgenic Arabidopsis and tobacco lines. Expression of P85 activity was detected in the midrib of leaves, leaf trichomes, apical meristemic regions, throughout the root, lateral roots and flowers. The expression of P87 was observed in leaf-tip, hydathodes, apical meristem, root tips, emerging lateral root tips, root stele region and in floral tissues. The bidirectional promoter in both orientations shows differential up-regulation (2.5 to 3 fold) under salt stress. Use of such regulatory elements of bidirectional promoters showing spatial and stress inducible promoter-functions in heterologous system might be an important tool for plant biotechnology and gene stacking applications. PMID:24260266

  6. An intergenic region shared by At4g35985 and At4g35987 in Arabidopsis thaliana is a tissue specific and stress inducible bidirectional promoter analyzed in transgenic arabidopsis and tobacco plants.

    Directory of Open Access Journals (Sweden)

    Joydeep Banerjee

    Full Text Available On chromosome 4 in the Arabidopsis genome, two neighboring genes (calmodulin methyl transferase At4g35987 and senescence associated gene At4g35985 are located in a head-to-head divergent orientation sharing a putative bidirectional promoter. This 1258 bp intergenic region contains a number of environmental stress responsive and tissue specific cis-regulatory elements. Transcript analysis of At4g35985 and At4g35987 genes by quantitative real time PCR showed tissue specific and stress inducible expression profiles. We tested the bidirectional promoter-function of the intergenic region shared by the divergent genes At4g35985 and At4g35987 using two reporter genes (GFP and GUS in both orientations in transient tobacco protoplast and Agro-infiltration assays, as well as in stably transformed transgenic Arabidopsis and tobacco plants. In transient assays with GFP and GUS reporter genes the At4g35985 promoter (P85 showed stronger expression (about 3.5 fold compared to the At4g35987 promoter (P87. The tissue specific as well as stress responsive functional nature of the bidirectional promoter was evaluated in independent transgenic Arabidopsis and tobacco lines. Expression of P85 activity was detected in the midrib of leaves, leaf trichomes, apical meristemic regions, throughout the root, lateral roots and flowers. The expression of P87 was observed in leaf-tip, hydathodes, apical meristem, root tips, emerging lateral root tips, root stele region and in floral tissues. The bidirectional promoter in both orientations shows differential up-regulation (2.5 to 3 fold under salt stress. Use of such regulatory elements of bidirectional promoters showing spatial and stress inducible promoter-functions in heterologous system might be an important tool for plant biotechnology and gene stacking applications.

  7. Clinical significance of Phosphatidyl Inositol Synthase overexpression in oral cancer

    International Nuclear Information System (INIS)

    Kaur, Jatinder; Sawhney, Meenakshi; DattaGupta, Siddartha; Shukla, Nootan K; Srivastava, Anurag; Ralhan, Ranju

    2010-01-01

    We reported increased levels of Phosphatidyl Inositol synthase (PI synthase), (enzyme that catalyses phosphatidyl inositol (PI) synthesis-implicated in intracellular signaling and regulation of cell growth) in smokeless tobacco (ST) exposed oral cell cultures by differential display. This study determined the clinical significance of PI synthase overexpression in oral squamous cell carcinoma (OSCC) and premalignant lesions (leukoplakia), and identified the downstream signaling proteins in PI synthase pathway that are perturbed by smokeless tobacco (ST) exposure. Tissue microarray (TMA) Immunohistochemistry, Western blotting, Confocal laser scan microscopy, RT-PCR were performed to define the expression of PI synthase in clinical samples and in oral cell culture systems. Significant increase in PI synthase immunoreactivity was observed in premalignant lesions and OSCCs as compared to oral normal tissues (p = 0.000). Further, PI synthase expression was significantly associated with de-differentiation of OSCCs, (p = 0.005) and tobacco consumption (p = 0.03, OR = 9.0). Exposure of oral cell systems to smokeless tobacco (ST) in vitro confirmed increase in PI synthase, Phosphatidylinositol 3-kinase (PI3K) and cyclin D1 levels. Collectively, increased PI synthase expression was found to be an early event in oral cancer and a target for smokeless tobacco

  8. Neuronal erythropoietin overexpression is protective against kanamycin-induced hearing loss in mice.

    Science.gov (United States)

    Bächinger, David; Horvath, Lukas; Eckhard, Andreas; Goosmann, Madeline M; Honegger, Tim; Gassmann, Max; Vogel, Johannes; Naldi, Arianne Monge

    2018-07-01

    Aminoglycosides have detrimental effects on the hair cells of the inner ear, yet these agents indisputably are one of the cornerstones in antibiotic therapy. Hence, there is a demand for strategies to prevent aminoglycoside-induced ototoxicity, which are not available today. In vitro data suggests that the pleiotropic growth factor erythropoietin (EPO) is neuroprotective against aminoglycoside-induced hair cell loss. Here, we use a mouse model with EPO-overexpression in neuronal tissue to evaluate whether EPO could also in vivo protect from aminoglycoside-induced hearing loss. Auditory brainstem response (ABR) thresholds were measured in 12-weeks-old mice before and after treatment with kanamycin for 15 days, which resulted in both C57BL/6 and EPO-transgenic animals in a high-frequency hearing loss. However, ABR threshold shifts in EPO-transgenic mice were significantly lower than in C57BL/6 mice (mean difference in ABR threshold shift 13.6 dB at 32 kHz, 95% CI 3.8-23.4 dB, p = 0.003). Correspondingly, quantification of hair cells and spiral ganglion neurons by immunofluorescence revealed that EPO-transgenic mice had a significantly lower hair cell and spiral ganglion neuron loss than C57BL/6 mice. In conclusion, neuronal overexpression of EPO is protective against aminoglycoside-induce hearing loss, which is in accordance with its known neuroprotective effects in other organs, such as the eye or the brain. Copyright © 2018 Elsevier B.V. All rights reserved.

  9. Dose-Dependent Rescue of KO Amelogenin Enamel by Transgenes in Vivo.

    Science.gov (United States)

    Bidlack, Felicitas B; Xia, Yan; Pugach, Megan K

    2017-01-01

    Mice lacking amelogenin (KO) have hypoplastic enamel. Overexpression of the most abundant amelogenin splice variant M180 and LRAP transgenes can substantially improve KO enamel, but only ~40% of the incisor thickness is recovered and the prisms are not as tightly woven as in WT enamel. This implies that the compositional complexity of the enamel matrix is required for different aspects of enamel formation, such as organizational structure and thickness. The question arises, therefore, how important the ratio of different matrix components, and in particular amelogenin splice products, is in enamel formation. Can optimal expression levels of amelogenin transgenes representing both the most abundant splice variants and cleavage product at protein levels similar to that of WT improve the enamel phenotype of KO mice? Addressing this question, our objective was here to understand dosage effects of amelogenin transgenes ( Tg ) representing the major splice variants M180 and LRAP and cleavage product CTRNC on enamel properties. Amelogenin KO mice were mated with M180 Tg , CTRNC Tg and LRAP Tg mice to generate M180 Tg and CTRNC Tg double transgene and M180 Tg , CTRNC Tg , LRAP Tg triple transgene mice with transgene hemizygosity (on one allelle) or homozygosity (on both alleles). Transgene homo- vs. hemizygosity was determined by qPCR and relative transgene expression confirmed by Western blot. Enamel volume and mineral density were analyzed by microCT, thickness and structure by SEM, and mechanical properties by Vickers microhardness testing. There were no differences in incisor enamel thickness between amelogenin KO mice with three or two different transgenes, but mice homozygous for a given transgene had significantly thinner enamel than mice hemizygous for the transgene ( p structure, but only up to a maximum of ~80% that of molar and ~40% that of incisor wild-type enamel.

  10. Transgenic plants producing the bacterial pheromone N-acyl-homoserine lactone exhibit enhanced resistance to the bacterial phytopathogen Erwinia carotovora.

    Science.gov (United States)

    Mäe, A; Montesano, M; Koiv, V; Palva, E T

    2001-09-01

    Bacterial pheromones, mainly different homoserine lactones, are central to a number of bacterial signaling processes, including those involved in plant pathogenicity. We previously demonstrated that N-oxoacyl-homoserine lactone (OHL) is essential for quorum sensing in the soft-rot phytopathogen Erwinia carotovora. In this pathogen, OHL controls the coordinate activation of genes encoding the main virulence determinants, extracellular plant cell wall degrading enzymes (PCWDEs), in a cell density-dependent manner. We suggest that E. carotovora employ quorum sensing to avoid the premature production of PCWDEs and subsequent activation of plant defense responses. To test whether modulating this sensory system would affect the outcome of a plant-pathogen interaction, we generated transgenic tobacco, producing OHL. This was accomplished by ectopic expression in tobacco of the E. carotovora gene expI, which is responsible for OHL biosynthesis. We show that expI-positive transgenic tobacco lines produced the active pheromone and partially complemented the avirulent phenotype of expI mutants. The OHL-producing tobacco lines exhibited enhanced resistance to infection by wild-type E. carotovora. The results were confirmed by exogenous addition of OHL to wild-type plants, which also resulted in increased resistance to E. carotovora.

  11. High level expression of Acidothermus cellulolyticus β-1, 4-endoglucanase in transgenic rice enhances the hydrolysis of its straw by cultured cow gastric fluid

    Directory of Open Access Journals (Sweden)

    Chou Hong

    2011-12-01

    Full Text Available Abstract Background Large-scale production of effective cellulose hydrolytic enzymes is the key to the bioconversion of agricultural residues to ethanol. The goal of this study was to develop a rice plant as a bioreactor for the large-scale production of cellulose hydrolytic enzymes via genetic transformation, and to simultaneously improve rice straw as an efficient biomass feedstock for conversion of cellulose to glucose. Results In this study, the cellulose hydrolytic enzyme β-1, 4-endoglucanase (E1 gene, from the thermophilic bacterium Acidothermus cellulolyticus, was overexpressed in rice through Agrobacterium-mediated transformation. The expression of the bacterial E1 gene in rice was driven by the constitutive Mac promoter, a hybrid promoter of Ti plasmid mannopine synthetase promoter and cauliflower mosaic virus 35S promoter enhancer, with the signal peptide of tobacco pathogenesis-related protein for targeting the E1 protein to the apoplastic compartment for storage. A total of 52 transgenic rice plants from six independent lines expressing the bacterial E1 enzyme were obtained that expressed the gene at high levels without severely impairing plant growth and development. However, some transgenic plants exhibited a shorter stature and flowered earlier than the wild type plants. The E1 specific activities in the leaves of the highest expressing transgenic rice lines were about 20-fold higher than those of various transgenic plants obtained in previous studies and the protein amounts accounted for up to 6.1% of the total leaf soluble protein. A zymogram and temperature-dependent activity analyses demonstrated the thermostability of the E1 enzyme and its substrate specificity against cellulose, and a simple heat treatment can be used to purify the protein. In addition, hydrolysis of transgenic rice straw with cultured cow gastric fluid for one hour at 39°C and another hour at 81°C yielded 43% more reducing sugars than wild type rice

  12. Regulation of Expression of the prb-1b / ACC Deaminase gene by UV-B in Transgenic tomatoes

    International Nuclear Information System (INIS)

    Tamot, B.K.; Pauls, K.P.; Glick, R.

    2003-01-01

    Transgenic tomato plants with 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase gene from Enterobacter cloacae UWA4 under the control of a pathogenesis-related promoter (prb-1b) from tobacco were challenged by abiotic stresses to determine the expression patterns of the transgene. No ACC deaminase RNA or protein was detected bu RT-PCR and in western blots prepared from leaf proteins of transgenic plants after wounding or treatment with alpha-amino butyric acid, xylanase, ethephon, salicylic acid, jasmonic acid , ethylene, or ethylene plus jasmonic acid. However, expression of the ACC deaminase transgene was observed in leaves and roots of transformed tomato lines exposed to UV light. The UV response required a minimum of 48 h of exposure and was specific to UV-B light

  13. Cell cycle stage-specific differential expression of topoisomerase I in tobacco BY-2 cells and its ectopic overexpression and knockdown unravels its crucial role in plant morphogenesis and development.

    Science.gov (United States)

    Singh, Badri Nath; Mudgil, Yashwanti; John, Riffat; Achary, V Mohan Murali; Tripathy, Manas Kumar; Sopory, Sudhir K; Reddy, Malireddy K; Kaul, Tanushri

    2015-11-01

    DNA topoisomerases catalyze the inter-conversion of different topological forms of DNA. Cell cycle coupled differential accumulation of topoisomerase I (Topo I) revealed biphasic expression maximum at S-phase and M/G1-phase of cultured synchronized tobacco BY-2 cells. This suggested its active role in resolving topological constrains during DNA replication (S-phase) and chromosome decondensation (M/G1 phase). Immuno-localization revealed high concentrations of Topo I in nucleolus. Propidium iodide staining and Br-UTP incorporation patterns revealed direct correlation between immunofluorescence intensity and rRNA transcription activity within nucleolus. Immuno-stained chromosomes during metaphase and anaphase suggested possible role of Topo I in resolving topological constrains during mitotic chromosome condensation. Inhibitor studies showed that in comparison to Topo I, Topo II was essential in resolving topological constrains during chromosome condensation. Probably, Topo II substituted Topo I functioning to certain extent during chromosome condensation, but not vice-versa. Transgenic Topo I tobacco lines revealed morphological abnormalities and highlighted its crucial role in plant morphogenesis and development. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  14. Promotion of seminomatous tumors by targeted overexpression of glial cell line-derived neurotrophic factor in mouse testis

    NARCIS (Netherlands)

    Meng, X.; de rooij, D. G.; Westerdahl, K.; Saarma, M.; Sariola, H.

    2001-01-01

    We show with transgenic mice that targeted overexpression of glial cell line-derived neurotrophic factor (GDNF) in undifferentiated spermatogonia promotes malignant testicular tumors, which express germ-cell markers. The tumors are invasive and contain aneuploid cells, but no distant metastases have

  15. Overexpression of Human-Derived DNMT3A Induced Intergenerational Inheritance of Active DNA Methylation Changes in Rat Sperm

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    Xiaoguo Zheng

    2017-12-01

    Full Text Available DNA methylation is the major focus of studies on paternal epigenetic inheritance in mammals, but most previous studies about inheritable DNA methylation changes are passively induced by environmental factors. However, it is unclear whether the active changes mediated by variations in DNA methyltransferase activity are heritable. Here, we established human-derived DNMT3A (hDNMT3A transgenic rats to study the effect of hDNMT3A overexpression on the DNA methylation pattern of rat sperm and to investigate whether this actively altered DNA methylation status is inheritable. Our results revealed that hDNMT3A was overexpressed in the testis of transgenic rats and induced genome-wide alterations in the DNA methylation pattern of rat sperm. Among 5438 reliable loci identified with 64 primer-pair combinations using a methylation-sensitive amplification polymorphism method, 28.01% showed altered amplified band types. Among these amplicons altered loci, 68.42% showed an altered DNA methylation status in the offspring of transgenic rats compared with wild-type rats. Further analysis based on loci which had identical DNA methylation status in all three biological replicates revealed that overexpression of hDNMT3A in paternal testis induced hypermethylation in sperm of both genotype-negative and genotype-positive offspring. Among the differentially methylated loci, 34.26% occurred in both positive and negative offspring of transgenic rats, indicating intergenerational inheritance of active DNA methylation changes in the absence of hDNM3A transmission. Furthermore, 75.07% of the inheritable loci were hyper-methylated while the remaining were hypomethylated. Distribution analysis revealed that the DNA methylation variations mainly occurred in introns and intergenic regions. Functional analysis revealed that genes related to differentially methylated loci were involved in a wide range of functions. Finally, this study demonstrated that active DNA methylation

  16. Over-expression of mango (Mangifera indica L.) MiARF2 inhibits root and hypocotyl growth of Arabidopsis.

    Science.gov (United States)

    Wu, Bei; Li, Yun-He; Wu, Jian-Yong; Chen, Qi-Zhu; Huang, Xia; Chen, Yun-Feng; Huang, Xue-Lin

    2011-06-01

    An auxin response factor 2 gene, MiARF2, was cloned in our previous study [1] from the cotyledon section of mango (Mangifera indica L. cv. Zihua) during adventitious root formation, which shares an 84% amino acid sequence similarity to Arabidopsis ARF2. This study was to examine the effects of over-expression of the full-length MiARF2 open reading frame on the root and hypocotyl growth in Arabidopsis. Phenotype analysis showed that the T(3) transgenic lines had about 20-30% reduction in the length of hypocotyls and roots of the seedlings in comparison with the wild-type. The transcription levels of ANT and ARGOS genes which play a role in controlling organ size and cell proliferation in the transgenic seedlings also decreased. Therefore, the inhibited root and hypocotyl growth in the transgenic seedlings may be associated with the down-regulated transcription of ANT and ARGOS by the over-expression of MiARF2. This study also suggests that although MiARF2 only has a single DNA-binding domain (DBD), it can function as other ARF-like proteins containing complete DBD, middle region (MR) and carboxy-terminal dimerization domain (CTD).

  17. Neuronal erythropoietin overexpression protects mice against age-related hearing loss (presbycusis).

    Science.gov (United States)

    Monge Naldi, Arianne; Belfrage, Celina; Jain, Neha; Wei, Eric T; Canto Martorell, Belén; Gassmann, Max; Vogel, Johannes

    2015-12-01

    So far, typical causes of presbycusis such as degeneration of hair cells and/or primary auditory (spiral ganglion) neurons cannot be treated. Because erythropoietin's (Epo) neuroprotective potential has been shown previously, we determined hearing thresholds of juvenile and aged mice overexpressing Epo in neuronal tissues. Behavioral audiometry revealed in contrast to 5 months of age, that 11-month-old Epo-transgenic mice had up to 35 dB lower hearing thresholds between 1.4 and 32 kHz, and at the highest frequencies (50-80 kHz), thresholds could be obtained in aged Epo-transgenic only but not anymore in old C57BL6 control mice. Click-evoked auditory brainstem response showed similar results. Numbers of spiral ganglion neurons in aged C57BL6 but not Epo-transgenic mice were dramatically reduced mainly in the basal turn, the location of high frequencies. In addition, there was a tendency to better preservation of inner and outer hair cells in Epo-transgenic mice. Hence, Epo's known neuroprotective action effectively suppresses the loss of spiral ganglion cells and probably also hair cells and, thus, development of presbycusis in mice. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Bmi1 overexpression in the cerebellar granule cell lineage of mice affects cell proliferation and survival without initiating medulloblastoma formation

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    Hourinaz Behesti

    2013-01-01

    BMI1 is a potent inducer of neural stem cell self-renewal and neural progenitor cell proliferation during development and in adult tissue homeostasis. It is overexpressed in numerous human cancers – including medulloblastomas, in which its functional role is unclear. We generated transgenic mouse lines with targeted overexpression of Bmi1 in the cerebellar granule cell lineage, a cell type that has been shown to act as a cell of origin for medulloblastomas. Overexpression of Bmi1 in granule cell progenitors (GCPs led to a decrease in cerebellar size due to decreased GCP proliferation and repression of the expression of cyclin genes, whereas Bmi1 overexpression in postmitotic granule cells improved cell survival in response to stress by altering the expression of genes in the mitochondrial cell death pathway and of Myc and Lef-1. Although no medulloblastomas developed in ageing cohorts of transgenic mice, crosses with Trp53−/− mice resulted in a low incidence of medulloblastoma formation. Furthermore, analysis of a large collection of primary human medulloblastomas revealed that tumours with a BMI1high TP53low molecular profile are significantly enriched in Group 4 human medulloblastomas. Our data suggest that different levels and timing of Bmi1 overexpression yield distinct cellular outcomes within the same cellular lineage. Importantly, Bmi1 overexpression at the GCP stage does not induce tumour formation, suggesting that BMI1 overexpression in GCP-derived human medulloblastomas probably occurs during later stages of oncogenesis and might serve to enhance tumour cell survival.

  19. Expression of Acidothermus cellulolyticus thermostable cellulases in tobacco and rice plants

    Directory of Open Access Journals (Sweden)

    Xiran Jiang

    2017-01-01

    Full Text Available The production of cellulases in plants is an economical method for the conversion of lignocellulosic biomass into fuels. Herein we report the expressions of two thermostable Acidothermus cellulolyticus cellulases, endo-1,4-β-D-glucanase (E1 and exoglucanase (Gux1, in tobacco and rice. To evaluate the expression of these recombinant cellulases, we expressed the full-length E1, the catalytic domains of E1 (E1cd and Gux1 (Gux1cd, as well as an E1–Gux1cd fusion enzyme in various subcellular compartments. In the case of tobacco, transgenic plants that expressed apoplast-localized E1 showed the highest level of activity, about three times higher than those that expressed the cytosolic E1. In the case of rice, the level of cellulase-specific activity in the transgenic plants ranged from 11 to 20 nmol 4-methylumbelliferone min−1 mg−1 total soluble protein. The recombinant cellulases exhibited good thermostability below 70 °C. Furthermore, transgenic rice leaves that were stored at room temperature for a month lost about 20% of the initial cellulase activity. Taken together, the results suggested that heterologous expression of thermostable cellulases in plants may be a viable option for biomass conversion.

  20. Non-motor and motor features in LRRK2 transgenic mice.

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    Zoë Bichler

    Full Text Available Non-motor symptoms are increasingly recognized as important features of Parkinson's disease (PD. LRRK2 mutations are common causes of familial and sporadic PD. Non-motor features have not been yet comprehensively evaluated in LRRK2 transgenic mouse models.Using a transgenic mouse model overexpressing the R1441G mutation of the human LRRK2 gene, we have investigated the longitudinal correlation between motor and non-motor symptoms and determined if specific non-motor phenotypes precede motor symptoms.We investigated the onset of motor and non-motor phenotypes on the LRRK2(R1441G BAC transgenic mice and their littermate controls from 4 to 21 month-old using a battery of behavioral tests. The transgenic mutant mice displayed mild hypokinesia in the open field from 16 months old, with gastrointestinal dysfunctions beginning at 6 months old. Non-motor features such as depression and anxiety-like behaviors, sensorial functions (pain sensitivity and olfaction, and learning and memory abilities in the passive avoidance test were similar in the transgenic animals compared to littermate controls.LRRK2(R1441G BAC transgenic mice displayed gastrointestinal dysfunction at an early stage but did not have abnormalities in fine behaviors, olfaction, pain sensitivity, mood disorders and learning and memory compared to non-transgenic littermate controls. The observations on olfaction and gastrointestinal dysfunction in this model validate findings in human carriers. These mice did recapitulate mild Parkinsonian motor features at late stages but compensatory mechanisms modulating the progression of PD in these models should be further evaluated.

  1. Salicylic acid-mediated and RNA-silencing defense mechanisms cooperate in the restriction of systemic spread of plum pox virus in tobacco.

    Science.gov (United States)

    Alamillo, Josefa M; Saénz, Pilar; García, Juan Antonio

    2006-10-01

    Plum pox virus (PPV) is able to replicate in inoculated leaves of Nicotiana tabacum, but is defective in systemic movement in this host. However, PPV produces a systemic infection in transgenic tobacco expressing the silencing suppressor P1/HC-Pro from tobacco etch virus (TEV). In this work we show that PPV is able to move to upper non-inoculated leaves of tobacco plants expressing bacterial salicylate hydroxylase (NahG) that degrades salicylic acid (SA). Replication and accumulation of PPV is higher in the locally infected leaves of plants deficient in SA or expressing TEV P1/HC-Pro silencing suppressor. Accumulation of viral derived small RNAs was reduced in the NahG transgenic plants, suggesting that SA might act as an enhancer of the RNA-silencing antiviral defense in tobacco. Besides, expression of SA-mediated defense transcripts, such as those of pathogenesis-related (PR) proteins PR-1 and PR-2 or alternative oxidase-1, as well as that of the putative RNA-dependent RNA polymerase NtRDR1, is induced in response to PPV infection, and the expression patterns of these defense transcripts are altered in the TEV P1/HC-Pro transgenic plants. Long-distance movement of PPV is highly enhanced in NahG x P1/HC-Pro double-transgenic plants and systemic symptoms in these plants reveal that the expression of an RNA-silencing suppressor and the lack of SA produce additive but distinct effects. Our results suggest that SA might act as an enhancer of the RNA-silencing antiviral defense in tobacco, and that silencing suppressors, such as P1/HC-Pro, also alter the SA-mediated defense. Both an RNA-silencing and an SA-mediated defense mechanism could act together to limit PPV infection.

  2. Overexpression of the PAP1 transcription factor reveals a complex regulation of flavonoid and phenylpropanoid metabolism in Nicotiana tabacum plants attacked by Spodoptera litura.

    Science.gov (United States)

    Mitsunami, Tomoko; Nishihara, Masahiro; Galis, Ivan; Alamgir, Kabir Md; Hojo, Yuko; Fujita, Kohei; Sasaki, Nobuhiro; Nemoto, Keichiro; Sawasaki, Tatsuya; Arimura, Gen-ichiro

    2014-01-01

    Anthocyanin pigments and associated flavonoids have demonstrated antioxidant properties and benefits for human health. Consequently, current plant bioengineers have focused on how to modify flavonoid metabolism in plants. Most of that research, however, does not consider the role of natural biotic stresses (e.g., herbivore attack). To understand the influence of herbivore attack on the metabolic engineering of flavonoids, we examined tobacco plants overexpressing the Arabidopsis PAP1 gene (encoding an MYB transcription factor), which accumulated anthocyanin pigments and other flavonoids/phenylpropanoids. In comparison to wild-type and control plants, transgenic plants exhibited greater resistance to Spodoptera litura. Moreover, herbivory suppressed the PAP1-induced increase of transcripts of flavonoid/phenylpropanoid biosynthetic genes (e.g., F3H) and the subsequent accumulation of these genes' metabolites, despite the unaltered PAP1 mRNA levels after herbivory. The instances of down-regulation were independent of the signaling pathways mediated by defense-related jasmonates but were relevant to the levels of PAP1-induced and herbivory-suppressed transcription factors, An1a and An1b. Although initially F3H transcripts were suppressed by herbivory, after the S. litura feeding was interrupted, F3H transcripts increased. We hypothesize that in transgenic plants responding to herbivory, there is a complex mechanism regulating enriched flavonoid/phenylpropanoid compounds, via biotic stress signals.

  3. Overexpression of the PAP1 transcription factor reveals a complex regulation of flavonoid and phenylpropanoid metabolism in Nicotiana tabacum plants attacked by Spodoptera litura.

    Directory of Open Access Journals (Sweden)

    Tomoko Mitsunami

    Full Text Available Anthocyanin pigments and associated flavonoids have demonstrated antioxidant properties and benefits for human health. Consequently, current plant bioengineers have focused on how to modify flavonoid metabolism in plants. Most of that research, however, does not consider the role of natural biotic stresses (e.g., herbivore attack. To understand the influence of herbivore attack on the metabolic engineering of flavonoids, we examined tobacco plants overexpressing the Arabidopsis PAP1 gene (encoding an MYB transcription factor, which accumulated anthocyanin pigments and other flavonoids/phenylpropanoids. In comparison to wild-type and control plants, transgenic plants exhibited greater resistance to Spodoptera litura. Moreover, herbivory suppressed the PAP1-induced increase of transcripts of flavonoid/phenylpropanoid biosynthetic genes (e.g., F3H and the subsequent accumulation of these genes' metabolites, despite the unaltered PAP1 mRNA levels after herbivory. The instances of down-regulation were independent of the signaling pathways mediated by defense-related jasmonates but were relevant to the levels of PAP1-induced and herbivory-suppressed transcription factors, An1a and An1b. Although initially F3H transcripts were suppressed by herbivory, after the S. litura feeding was interrupted, F3H transcripts increased. We hypothesize that in transgenic plants responding to herbivory, there is a complex mechanism regulating enriched flavonoid/phenylpropanoid compounds, via biotic stress signals.

  4. Overexpression of the PAP1 Transcription Factor Reveals a Complex Regulation of Flavonoid and Phenylpropanoid Metabolism in Nicotiana tabacum Plants Attacked by Spodoptera litura

    Science.gov (United States)

    Mitsunami, Tomoko; Nishihara, Masahiro; Galis, Ivan; Alamgir, Kabir Md; Hojo, Yuko; Fujita, Kohei; Sasaki, Nobuhiro; Nemoto, Keichiro; Sawasaki, Tatsuya; Arimura, Gen-ichiro

    2014-01-01

    Anthocyanin pigments and associated flavonoids have demonstrated antioxidant properties and benefits for human health. Consequently, current plant bioengineers have focused on how to modify flavonoid metabolism in plants. Most of that research, however, does not consider the role of natural biotic stresses (e.g., herbivore attack). To understand the influence of herbivore attack on the metabolic engineering of flavonoids, we examined tobacco plants overexpressing the Arabidopsis PAP1 gene (encoding an MYB transcription factor), which accumulated anthocyanin pigments and other flavonoids/phenylpropanoids. In comparison to wild-type and control plants, transgenic plants exhibited greater resistance to Spodoptera litura. Moreover, herbivory suppressed the PAP1-induced increase of transcripts of flavonoid/phenylpropanoid biosynthetic genes (e.g., F3H) and the subsequent accumulation of these genes' metabolites, despite the unaltered PAP1 mRNA levels after herbivory. The instances of down-regulation were independent of the signaling pathways mediated by defense-related jasmonates but were relevant to the levels of PAP1-induced and herbivory-suppressed transcription factors, An1a and An1b. Although initially F3H transcripts were suppressed by herbivory, after the S. litura feeding was interrupted, F3H transcripts increased. We hypothesize that in transgenic plants responding to herbivory, there is a complex mechanism regulating enriched flavonoid/phenylpropanoid compounds, via biotic stress signals. PMID:25268129

  5. High blood pressure in transgenic mice carrying the rat angiotensinogen gene.

    Science.gov (United States)

    Kimura, S; Mullins, J J; Bunnemann, B; Metzger, R; Hilgenfeldt, U; Zimmermann, F; Jacob, H; Fuxe, K; Ganten, D; Kaling, M

    1992-01-01

    Transgenic mice were generated by injecting the entire rat angiotensinogen gene into the germline of NMRI mice. The resulting transgenic animals were characterized with respect to hemodynamics, parameters of the renin angiotension system, and expression of the transgene. The transgenic line TGM(rAOGEN)123 developed hypertension with a mean arterial blood pressure of 158 mmHg in males and 132 mmHg in females. In contrast, the transgenic line TGM(rAOGEN)92 was not hypertensive. Rat angiotensinogen was detectable only in plasma of animals of line 123. Total plasma angiotensinogen and plasma angiotensin II concentrations were about three times as high as those of negative control mice. In TGM(rAOGEN)123 the transgene was highly expressed in liver and brain. Transcripts were also detected in heart, kidney and testis. In TGM(rAOGEN)92 the brain was the main expressing organ. In situ hybridization revealed an mRNA distribution in the brain of TGM(rAOGEN)123 similar to the one in rat. In TGM(rAOGEN)92 the expression pattern in the brain was aberrant. These data indicate that overexpression of the angiotensinogen gene in liver and brain leads to the development of hypertension in transgenic mice. The TGM(rAOGEN)123 constitutes a high angiotensin II type of hypertension and may provide a new experimental animal model to study the kinetics and function of the renin angiotensin system. Images PMID:1547785

  6. Genetically transformed tobacco plants expressing synthetic EPSPS gene confer tolerance against glyphosate herbicide.

    Science.gov (United States)

    Imran, Muhammad; Asad, Shaheen; Barboza, Andre Luiz; Galeano, Esteban; Carrer, Helaine; Mukhtar, Zahid

    2017-04-01

    Glyphosate quashes the synthesis of 5-enolpyruvylshikimate-3- phosphate synthase (EPSPS) enzyme which intercedes the functioning of shikimate pathway for the production of aromatic amino acids. Herbicide resistant crops are developed using glyphosate insensitive EPSPS gene isolated from Agrobacterium sp. strain CP4, which give farmers a sustainable weed control option. Intentions behind this study were to design and characterize the synthetic herbicide resistant CP4 - EPSPS gene in a model plant system and check the effectiveness of transformed tobacco against application of glyphosate. Putative transgenic plants were obtained from independent transformation events, and stable plant transformation, transgene expression and integration were demonstrated respectively by PCR, qRT-PCR and Southern hybridization. Gene transcript level and gene copy number (1-4) varied among the tested transgenic tobacco lines. Herbicide assays showed that transgenic plants were resistant to glyphosate after 12 days of spraying with glyphosate, and EPSPS activity remained at sufficient level to withstand the spray at 1000 ppm of the chemical. T 1 plants analyzed through immunoblot strips and PCR showed that the gene was being translated into protein and transmitted to the next generation successfully. This codon optimized synthetic CP4 - EPSPS gene is functionally equivalent to the gene for glyphosate resistance available in the commercial crops and hence we recommend this gene for transformation into commercial crops.

  7. α-Lipoic acid prevents lipotoxic cardiomyopathy in acyl CoA-synthase transgenic mice

    International Nuclear Information System (INIS)

    Lee, Young; Naseem, R. Haris; Park, Byung-Hyun; Garry, Daniel J.; Richardson, James A.; Schaffer, Jean E.; Unger, Roger H.

    2006-01-01

    α-Lipoic acid (α-LA) mimics the hypothalamic actions of leptin on food intake, energy expenditure, and activation of AMP-activated protein kinase (AMPK). To determine if, like leptin, α-LA protects against cardiac lipotoxicity, α-LA was fed to transgenic mice with cardiomyocyte-specific overexpression of the acyl CoA synthase (ACS) gene. Untreated ACS-transgenic mice died prematurely with increased triacylglycerol content and dilated cardiomyopathy, impaired systolic function and myofiber disorganization, apoptosis, and interstitial fibrosis on microscopy. In α-LA-treated ACS-transgenic mice heart size, echocardiogram and TG content were normal. Plasma TG fell 50%, hepatic-activated phospho-AMPK rose 6-fold, sterol regulatory element-binding protein-1c declined 50%, and peroxisome proliferator-activated receptor-γ cofactor-1α mRNA rose 4-fold. Since food restriction did not prevent lipotoxicity, we conclude that α-LA treatment, like hyperleptinemia, protects the heart of ACS-transgenic mice from lipotoxicity

  8. Cyclin D1 and mammary carcinoma: new insights from transgenic mouse models

    International Nuclear Information System (INIS)

    Sutherland, Robert L; Musgrove, Elizabeth A

    2002-01-01

    Cyclin D1 is one of the most commonly overexpressed oncogenes in breast cancer, with 45–50% of primary ductal carcinomas overexpressing this oncoprotein. Targeted deletion of the gene encoding cyclin D1 demonstrates an essential role in normal mammary gland development while transgenic studies provide evidence that cyclin D1 is a weak oncogene in mammary epithelium. In a recent exciting development, Yu et al. demonstrate that cyclin D1-deficient mice are resistant to mammary carcinomas induced by c-neu and v-Ha-ras, but not those induced by c-myc or Wnt-1. These findings define a pivotal role for cyclin D1 in a subset of mammary cancers in mice and imply a functional role for cyclin D1 overexpression in human breast cancer

  9. Highly phosphorylated functionalized rice starch produced by transgenic rice expressing the potato GWD1 gene

    DEFF Research Database (Denmark)

    Chen, Yaling; Sun, Xiao-Feng; Zhou, Xin

    2017-01-01

    Starch phosphorylation occurs naturally during starch metabolism in the plant and is catalysed by glucan water dikinases (GWD1) and phosphoglucan water dikinase/glucan water dikinase 3 (PWD/GWD3). We generated six stable individual transgenic lines by over-expressing the potato GWD1 in rice....... Transgenic rice grain starch had 9-fold higher 6-phospho (6-P) monoesters and double amounts of 3-phospho (3-P) monoesters, respectively, compared to control grain. The shape and topography of the transgenic starch granules were moderately altered including surface pores and less well defined edges...... content was positively correlated with short chains of DP6-12. The starch pasting temperature, peak viscosity and the breakdown were lower but the setback was higher for transgenic rice flour. The 6-P content was negatively correlated with texture adhesiveness but positively correlated...

  10. The effect of constitutive over-expression of insulin-like growth factor 1 on the cognitive function in aged mice.

    Science.gov (United States)

    Hu, Ankang; Yuan, Honghua; Wu, Lianlian; Chen, Renjin; Chen, Quangang; Zhang, Tengye; Wang, Zhenzhen; Liu, Peng; Zhu, Xiaorong

    2016-01-15

    The neurotrophic factor insulin-like growth factor (IGF)-1 promotes neurogenesis in the mammalian brain and provides protection against brain injury. However, studies regarding the effects of IGF-1 on cognitive function in aged mice remain limited. We investigated the effects of overexpression of IGF-1 specifically in neural stem cells of the hippocampal dentate gyrus on the recognitive function in 18-month-old transgenic mice. Immunohistocytochemistry and Nissl staining revealed the increased population of BrdU-positive cells as well as the upregulated expression of Nestin and neuronal nuclei (NeuN), respective markers for neural progenitors and neurons, in the hippocampus of the aged IGF-1 transgenic mice versus the wild-type, suggesting that IGF-1 overexpression promotes neurogenesis. In addition, the IGF-1 receptor (IGF-1R), the phosphorylation of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) and extracellular signal-regulated kinase (ERK) were enhanced in the transgenic mice than in the wild-type. Transgenic mice also showed superior performance in the Morris water maze and step-down memory tests to their wild-type counterparts. Moreover, the learning and memory abilities of transgenic mice were significantly undermined with the blockage of CaMKII and ERK signaling pathway. Accordingly, our findings indicated that IGF-1 may mitigate the aged-associated cognitive decline via promoting neurogenesis in the hippocampus and activating CaMKII and ERK signaling by binding with IGF-1R. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Arabidopsis genes, AtNPR1, AtTGA2 and AtPR-5, confer partial resistance to soybean cyst nematode (Heterodera glycines) when overexpressed in transgenic soybean roots

    Science.gov (United States)

    2014-01-01

    Background Extensive studies using the model system Arabidopsis thaliana to elucidate plant defense signaling and pathway networks indicate that salicylic acid (SA) is the key hormone triggering the plant defense response against biotrophic and hemi-biotrophic pathogens, while jasmonic acid (JA) and derivatives are critical to the defense response against necrotrophic pathogens. Several reports demonstrate that SA limits nematode reproduction. Results Here we translate knowledge gained from studies using Arabidopsis to soybean. The ability of thirty-one Arabidopsis genes encoding important components of SA and JA synthesis and signaling in conferring resistance to soybean cyst nematode (SCN: Heterodera glycines) are investigated. We demonstrate that overexpression of three of thirty-one Arabidoposis genes in transgenic soybean roots of composite plants decreased the number of cysts formed by SCN to less than 50% of those found on control roots, namely AtNPR1(33%), AtTGA2 (38%), and AtPR-5 (38%). Three additional Arabidopsis genes decreased the number of SCN cysts by 40% or more: AtACBP3 (53% of the control value), AtACD2 (55%), and AtCM-3 (57%). Other genes having less or no effect included AtEDS5 (77%), AtNDR1 (82%), AtEDS1 (107%), and AtPR-1 (80%), as compared to control. Overexpression of AtDND1 greatly increased susceptibility as indicated by a large increase in the number of SCN cysts (175% of control). Conclusions Knowledge of the pathogen defense system gained from studies of the model system, Arabidopsis, can be directly translated to soybean through direct overexpression of Arabidopsis genes. When the genes, AtNPR1, AtGA2, and AtPR-5, encoding specific components involved in SA regulation, synthesis, and signaling, are overexpressed in soybean roots, resistance to SCN is enhanced. This demonstrates functional compatibility of some Arabidopsis genes with soybean and identifies genes that may be used to engineer resistance to nematodes. PMID:24739302

  12. Arabidopsis genes, AtNPR1, AtTGA2 and AtPR-5, confer partial resistance to soybean cyst nematode (Heterodera glycines) when overexpressed in transgenic soybean roots.

    Science.gov (United States)

    Matthews, Benjamin F; Beard, Hunter; Brewer, Eric; Kabir, Sara; MacDonald, Margaret H; Youssef, Reham M

    2014-04-16

    Extensive studies using the model system Arabidopsis thaliana to elucidate plant defense signaling and pathway networks indicate that salicylic acid (SA) is the key hormone triggering the plant defense response against biotrophic and hemi-biotrophic pathogens, while jasmonic acid (JA) and derivatives are critical to the defense response against necrotrophic pathogens. Several reports demonstrate that SA limits nematode reproduction. Here we translate knowledge gained from studies using Arabidopsis to soybean. The ability of thirty-one Arabidopsis genes encoding important components of SA and JA synthesis and signaling in conferring resistance to soybean cyst nematode (SCN: Heterodera glycines) are investigated. We demonstrate that overexpression of three of thirty-one Arabidoposis genes in transgenic soybean roots of composite plants decreased the number of cysts formed by SCN to less than 50% of those found on control roots, namely AtNPR1(33%), AtTGA2 (38%), and AtPR-5 (38%). Three additional Arabidopsis genes decreased the number of SCN cysts by 40% or more: AtACBP3 (53% of the control value), AtACD2 (55%), and AtCM-3 (57%). Other genes having less or no effect included AtEDS5 (77%), AtNDR1 (82%), AtEDS1 (107%), and AtPR-1 (80%), as compared to control. Overexpression of AtDND1 greatly increased susceptibility as indicated by a large increase in the number of SCN cysts (175% of control). Knowledge of the pathogen defense system gained from studies of the model system, Arabidopsis, can be directly translated to soybean through direct overexpression of Arabidopsis genes. When the genes, AtNPR1, AtGA2, and AtPR-5, encoding specific components involved in SA regulation, synthesis, and signaling, are overexpressed in soybean roots, resistance to SCN is enhanced. This demonstrates functional compatibility of some Arabidopsis genes with soybean and identifies genes that may be used to engineer resistance to nematodes.

  13. FT overexpression induces precocious flowering and normal reproductive development in Eucalyptus.

    Science.gov (United States)

    Klocko, Amy L; Ma, Cathleen; Robertson, Sarah; Esfandiari, Elahe; Nilsson, Ove; Strauss, Steven H

    2016-02-01

    Eucalyptus trees are among the most important species for industrial forestry worldwide. However, as with most forest trees, flowering does not begin for one to several years after planting which can limit the rate of conventional and molecular breeding. To speed flowering, we transformed a Eucalyptus grandis × urophylla hybrid (SP7) with a variety of constructs that enable overexpression of FLOWERING LOCUS T (FT). We found that FT expression led to very early flowering, with events showing floral buds within 1-5 months of transplanting to the glasshouse. The most rapid flowering was observed when the cauliflower mosaic virus 35S promoter was used to drive the Arabidopsis thaliana FT gene (AtFT). Early flowering was also observed with AtFT overexpression from a 409S ubiquitin promoter and under heat induction conditions with Populus trichocarpa FT1 (PtFT1) under control of a heat-shock promoter. Early flowering trees grew robustly, but exhibited a highly branched phenotype compared to the strong apical dominance of nonflowering transgenic and control trees. AtFT-induced flowers were morphologically normal and produced viable pollen grains and viable self- and cross-pollinated seeds. Many self-seedlings inherited AtFT and flowered early. FT overexpression-induced flowering in Eucalyptus may be a valuable means for accelerating breeding and genetic studies as the transgene can be easily segregated away in progeny, restoring normal growth and form. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  14. Antimicrobial activity of γ-thionin-like soybean SE60 in E. coli and tobacco plants

    International Nuclear Information System (INIS)

    Choi, Yeonhee; Choi, Yang Do; Lee, Jong Seob

    2008-01-01

    The SE60, a low molecular weight, sulfur-rich protein in soybean, is known to be homologous to wheat γ-purothionin. To elucidate the functional role of SE60, we expressed SE60 cDNA in Escherichia coli and in tobacco plants. A single protein band was detected by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) after anti-FLAG affinity purification of the protein from transformed E. coli. While the control E. coli cells harboring pFLAG-1 showed standard growth with Isopropyl β-D-1-thiogalactopyranoside (IPTG) induction, E. coli cells expressing the SE60 fusion protein did not grow at all, suggesting that SE60 has toxic effects on E. coli growth. Genomic integration and the expression of transgene in the transgenic tobacco plants were confirmed by Southern and Northern blot analysis, respectively. The transgenic plants demonstrated enhanced resistance against the pathogen Pseudomonas syringae. Taken together, these results strongly suggest that SE60 has antimicrobial activity and play a role in the defense mechanism in soybean plants

  15. BDNF Overexpression Exhibited Bilateral Effect on Neural Behavior in SCT Mice Associated with AKT Signal Pathway.

    Science.gov (United States)

    Chen, Mei-Rong; Dai, Ping; Wang, Shu-Fen; Song, Shu-Hua; Wang, Hang-Ping; Zhao, Ya; Wang, Ting-Hua; Liu, Jia

    2016-10-01

    Spinal cord injury (SCI), a severe health problem in worldwide, was commonly associated with functional disability and reduced quality of life. As the expression of brain-derived neurotrophic factor (BDNF) was substantial event in injured spinal cord, we hypothesized whether BDNF-overexpression could be in favor of the recovery of both sensory function and hindlimb function after SCI. By using BDNF-overexpression transgene mice [CMV-BDNF 26 (CB26) mice] we assessed the role of BDNF on the recovery of neurological behavior in spinal cord transection (SCT) model. BMS score and tail-flick test was performed to evaluate locomotor function and sensory function, respectively. Immunohistochemistry was employed to detect the location and the expression of BDNF, NeuN, 5-HT, GAP-43, GFAP as well as CGRP, and the level of p-AKT and AKT were examined through western blot analysis. BDNF overexpressing resulted in significant locomotor functional recovery from 21 to 28 days after SCT, compared with wild type (WT)+SCT group. Meanwhile, the NeuN, 5-HT and GAP-43 positive cells were markedly increased in ventral horn in BDNF overexpression animals, compared with WT mice with SCT. Moreover, the crucial molecular signal, p-AKT/AKT has been largely up-regulated, which is consistent with the improvement of locomotor function. However, in this study, thermal hyperpathia encountered in sham (CB26) group and WT+SCT mice and further aggravated in CB26 mice after SCT. Also, following SCT, the significant augment of positive-GFAP astrocytes and CGRP fibers were found in WT+SCT mice, and further increase was seen in BDNF over-expression transgene mice. BDNF-overexpression may not only facilitate the recovery of locomotor function via AKT pathway, but also contributed simultaneously to thermal hyperalgesia after SCT.

  16. Ntdin, a tobacco senescence-associated gene, is involved in molybdenum cofactor biosynthesis.

    Science.gov (United States)

    Yang, Seung Hwan; Berberich, Thomas; Miyazaki, Atsushi; Sano, Hiroshi; Kusano, Tomonobu

    2003-10-01

    To date, dozens of genes have been reported to be up-regulated with senescence in higher plants. Radish din1 and its ortholog sen1 of Arabidopsis are known as such, but their function is not clear yet. Here we have isolated their counterpart cDNA from tobacco and designated it as NTDIN: Its product, Ntdin, a 185 amino acid polypeptide with 56.8% and 54.2% identity to Atsen1 and Rsdin1, respectively, is localized in chloroplasts. Transcripts of Ntdin are induced by sulfate or nitrate but not by phosphate, suggesting its involvement in sulfur and nitrogen metabolism. A database search revealed that Ntdin shows similarity with the C-terminal region of Nicotiana plumbaginifolia Cnx5, which functions in molybdenum cofactor (Moco) biosynthesis. Transgenic tobacco plants with suppressed Ntdin are more tolerant to chlorate, a substrate analog of nitrate reductase, than controls, implying low nitrate reductase activity in the transgenic plants due to a deficiency of Moco. Indeed, enzymatic activities of two molybdoenzymes, nitrate reductase and xanthine dehydrogenase, in transgenic plants are found to be significantly lower than in control plants. Direct measurement of Moco contents reveals that those transgenic plants contain about 5% Moco of those of the control plants. Abscisic acid and indole-3-acidic acid, whose biosynthetic pathways require Moco, up-regulated Ntdin expression. Taken together, it is concluded that Ntdin functions in a certain step in Moco biosynthesis.

  17. Characterization of a wheat (Triticum aestivum L.) expansin gene, TaEXPB23, involved in the abiotic stress response and phytohormone regulation.

    Science.gov (United States)

    Han, Yang yang; Li, Ai xiu; Li, Feng; Zhao, Mei rong; Wang, Wei

    2012-05-01

    Expansins are proteins that are generally accepted to be key regulators of cell wall extension and plant growth. We examined the expression pattern of TaEXPB23, a wheat (Triticum aestivum L.) expansin gene, under exogenous phytohormone and abiotic stress treatments. In addition, we evaluated its function in the tolerance to salt stress and high temperature (HT) by overexpressing it in transgenic tobacco plants. In subcellular localization assays, TaEXPB23 localized to the cell wall. Expression analysis demonstrated that the transcription pattern of TaEXPB23 corresponded to wheat coleoptile growth. Real-time RT-PCR analysis revealed that TaEXPB23 transcript expression was upregulated by exogenous methyl jasmonate (MeJA) and salt stress, but downregulated by exogenous gibberellins (GA₃), ethylene (ET), indole-3-acetic acid (IAA) and α-naphthlcetic acid (NAA). Overexpression of TaEXPB23 in tobacco (tabacum) conferred tolerance to salt stress by enhancing water retention ability (WRA) and decreasing osmotic potential (OP). However, transgenic plants overexpressing TaEXPB23 did not show any improvement in the tolerance to HT stress. These results suggested that TaEXPB23 is regulated by phytohormones and is involved in the regulation of salt stress tolerance. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  18. In vivo overexpression of Emi1 promotes chromosome instability and tumorigenesis.

    Science.gov (United States)

    Vaidyanathan, S; Cato, K; Tang, L; Pavey, S; Haass, N K; Gabrielli, B G; Duijf, P H G

    2016-10-13

    Cell cycle genes are often aberrantly expressed in cancer, but how their misexpression drives tumorigenesis mostly remains unclear. From S phase to early mitosis, EMI1 (also known as FBXO5) inhibits the anaphase-promoting complex/cyclosome, which controls cell cycle progression through the sequential degradation of various substrates. By analyzing 7403 human tumor samples, we find that EMI1 overexpression is widespread in solid tumors but not in blood cancers. In solid cancers, EMI1 overexpression is a strong prognostic marker for poor patient outcome. To investigate causality, we generated a transgenic mouse model in which we overexpressed Emi1. Emi1-overexpressing animals develop a wide variety of solid tumors, in particular adenomas and carcinomas with inflammation and lymphocyte infiltration, but not blood cancers. These tumors are significantly larger and more penetrant, abundant, proliferative and metastatic than control tumors. In addition, they are highly aneuploid with tumor cells frequently being in early mitosis and showing mitotic abnormalities, including lagging and incorrectly segregating chromosomes. We further demonstrate in vitro that even though EMI1 overexpression may cause mitotic arrest and cell death, it also promotes chromosome instability (CIN) following delayed chromosome alignment and anaphase onset. In human solid tumors, EMI1 is co-expressed with many markers for CIN and EMI1 overexpression is a stronger marker for CIN than most well-established ones. The fact that Emi1 overexpression promotes CIN and the formation of solid cancers in vivo indicates that Emi1 overexpression actively drives solid tumorigenesis. These novel mechanistic insights have important clinical implications.

  19. Immunogenicity of nuclear-encoded LTB:ST fusion protein from Escherichia coli expressed in tobacco plants.

    Science.gov (United States)

    Rosales-Mendoza, Sergio; Soria-Guerra, Ruth E; Moreno-Fierros, Leticia; Govea-Alonso, Dania O; Herrera-Díaz, Areli; Korban, Schuyler S; Alpuche-Solís, Ángel G

    2011-06-01

    Enterotoxigenic Escherichia coli (ETEC) is one of the main causative agents of diarrhea in infants and for travelers. Inclusion of a heat-stable (ST) toxin into vaccine formulations is mandatory as most ETEC strains can produce both heat-labile (LT) and ST enterotoxins. In this study, a genetic fusion gene encoding for an LTB:ST protein has been constructed and transferred into tobacco via Agrobacterium tumefaciens-mediated transformation. Transgenic tobacco plants carrying the LTB:ST gene are then subjected to GM1-ELISA revealing that the LTB:ST has assembled into pentamers and displays antigenic determinants from both LTB and ST. Protein accumulation of up to 0.05% total soluble protein is detected. Subsequently, mucosal and systemic humoral responses are elicited in mice orally dosed with transgenic tobacco leaves. This has suggested that the plant-derived LTB:ST is immunogenic via the oral route. These findings are critical for the development of a plant-based vaccine capable of eliciting broader protection against ETEC and targeting both LTB and ST. Features of this platform in comparison to transplastomic approaches are discussed.

  20. Overexpression of E3 Ubiquitin Ligase Gene AdBiL Contributes to Resistance against Chilling Stress and Leaf Mold Disease in Tomato

    Directory of Open Access Journals (Sweden)

    Shuangchen Chen

    2017-06-01

    Full Text Available Ubiquitination is a common regulatory mechanism, playing a critical role in diverse cellular and developmental processes in eukaryotes. However, a few reports on the functional correlation between E3 ubiquitin ligases and reactive oxygen species (ROS or reactive nitrogen species (RNS metabolism in response to stress are currently available in plants. In the present study, the E3 ubiquitin ligase gene AdBiL (Adi3 Binding E3 Ligase was introduced into tomato line Ailsa Craig via Agrobacterium-mediated method. Transgenic lines were confirmed for integration into the tomato genome using PCR. Transcription of AdBiL in various transgenic lines was determined using real-time PCR. Evaluation of stress tolerance showed that T1 generation of transgenic tomato lines showed only mild symptoms of chilling injury as evident by higher biomass accumulation and chlorophyll content than those of non-transformed plants. Compared with wild-type plants, the contents of AsA, AsA/DHA, GSH and the activity of GaILDH, γ-GCS and GSNOR were increased, while H2O2, O2.−, MDA, NO, SNOs, and GSNO accumulations were significantly decreased in AdBiL overexpressing plants in response to chilling stress. Furthermore, transgenic tomato plants overexpressing AdBiL showed higher activities of enzymes such as G6PDH, 6PGDH, NADP-ICDH, and NADP-ME involved in pentose phosphate pathway (PPP. The transgenic tomato plants also exhibited an enhanced tolerance against the necrotrophic fungus Cladosporium fulvum. Tyrosine nitration protein was activated in the plants infected with leaf mold disease, while the inhibition could be recovered in AdBiL gene overexpressing lines. Taken together, our results revealed a possible physiological role of AdBiL in the activation of the key enzymes of AsA–GSH cycle, PPP and down-regulation of GSNO reductase, thereby reducing oxidative and nitrosative stress in plants. This study demonstrates an optimized transgenic strategy using AdBiL gene for crop

  1. High level of expression of recombinant human granulocyte-macrophage colony stimulating factor in transgenic rice cell suspension culture

    DEFF Research Database (Denmark)

    Shin, Yun-Ji; Hong, Shin-Young; Kwon, Tae-Ho

    2003-01-01

    Recombinant human granulocyte-macrophage colony stimulating factor (hGM-CSF) has been previously produced in tobacco cell suspension cultures. However, the amount of hGM-CSF accumulated in the culture medium dropped quickly from its maximum of 150 microg/L at 5 d after incubation. To overcome...... of recombinant hGM-CSF in transgenic rice cell suspension culture and protease activity of this culture medium was low compared to that of tobacco culture system....

  2. Overexpression of NtPR-Q Up-Regulates Multiple Defense-Related Genes in Nicotiana tabacum and Enhances Plant Resistance to Ralstonia solanacearum

    Directory of Open Access Journals (Sweden)

    Yuanman Tang

    2017-11-01

    Full Text Available Various classes of plant pathogenesis-related proteins have been identified in the past several decades. PR-Q, a member of the PR3 family encoding chitinases, has played an important role in regulating plant resistance and preventing pathogen infection. In this paper, we functionally characterized NtPR-Q in tobacco plants and found that the overexpression of NtPR-Q in tobacco Yunyan87 resulted in higher resistance to Ralstonia solanacearum inoculation. Surprisingly, overexpression of NtPR-Q led to the activation of many defense-related genes, such as salicylic acid (SA-responsive genes NtPR1a/c, NtPR2 and NtCHN50, JA-responsive gene NtPR1b and ET production-associated genes NtACC Oxidase and NtEFE26. Consistent with the role of NtPR-Q in multiple stress responses, NtPR-Q transcripts were induced by the exogenous hormones SA, ethylene and methyl jasmonate, which could enhance the resistance of tobacco to R. solanacearum. Collectively, our results suggested that NtPR-Q overexpression led to the up-regulation of defense-related genes and enhanced plant resistance to R. solanacearum infection.

  3. Dose-Dependent Rescue of KO Amelogenin Enamel by Transgenes in Vivo

    Directory of Open Access Journals (Sweden)

    Felicitas B. Bidlack

    2017-11-01

    Full Text Available Mice lacking amelogenin (KO have hypoplastic enamel. Overexpression of the most abundant amelogenin splice variant M180 and LRAP transgenes can substantially improve KO enamel, but only ~40% of the incisor thickness is recovered and the prisms are not as tightly woven as in WT enamel. This implies that the compositional complexity of the enamel matrix is required for different aspects of enamel formation, such as organizational structure and thickness. The question arises, therefore, how important the ratio of different matrix components, and in particular amelogenin splice products, is in enamel formation. Can optimal expression levels of amelogenin transgenes representing both the most abundant splice variants and cleavage product at protein levels similar to that of WT improve the enamel phenotype of KO mice? Addressing this question, our objective was here to understand dosage effects of amelogenin transgenes (Tg representing the major splice variants M180 and LRAP and cleavage product CTRNC on enamel properties. Amelogenin KO mice were mated with M180Tg, CTRNCTg and LRAPTg mice to generate M180Tg and CTRNCTg double transgene and M180Tg, CTRNCTg, LRAPTg triple transgene mice with transgene hemizygosity (on one allelle or homozygosity (on both alleles. Transgene homo- vs. hemizygosity was determined by qPCR and relative transgene expression confirmed by Western blot. Enamel volume and mineral density were analyzed by microCT, thickness and structure by SEM, and mechanical properties by Vickers microhardness testing. There were no differences in incisor enamel thickness between amelogenin KO mice with three or two different transgenes, but mice homozygous for a given transgene had significantly thinner enamel than mice hemizygous for the transgene (p < 0.05. The presence of the LRAPTg did not improve the phenotype of M180Tg/CTRNCTg/KO enamel. In the absence of endogenous amelogenin, the addition of amelogenin transgenes representing the most

  4. Mice orally immunized with a transgenic plant expressing the glycoprotein of Crimean-Congo hemorrhagic fever virus

    DEFF Research Database (Denmark)

    Ghiasi, Seyed Mojtaba; Salmanian, A H; Chinikar, S

    2011-01-01

    in their serum and feces, respectively. The mice in the fed/boosted group showed a significant rise in specific IgG antibodies after a single boost. Our results imply that oral immunization of animals with edible materials from transgenic plants is feasible, and further assessments are under way. In addition......While Crimean-Congo hemorrhagic fever (CCHF) has a high mortality rate in humans, the associated virus (CCHFV) does not induce clinical symptoms in animals, but animals play an important role in disease transmission to humans. Our aim in this study was to examine the immunogenicity of the CCHFV...... glycoprotein when expressed in the root and leaf of transgenic plants via hairy roots and stable transformation of tobacco plants, respectively. After confirmatory analyses of transgenic plant lines and quantification of the expressed glycoprotein, mice were either fed with the transgenic leaves or roots, fed...

  5. Comparative study of transgenic Brachypodium distachyon expressing sucrose:fructan 6-fructosyltransferases from wheat and timothy grass with different enzymatic properties.

    Science.gov (United States)

    Tamura, Ken-Ichi; Sanada, Yasuharu; Tase, Kazuhiro; Kawakami, Akira; Yoshida, Midori; Yamada, Toshihiko

    2014-04-01

    Fructans can act as cryoprotectants and contribute to freezing tolerance in plant species, such as in members of the grass subfamily Pooideae that includes Triticeae species and forage grasses. To elucidate the relationship of freezing tolerance, carbohydrate composition and degree of polymerization (DP) of fructans, we generated transgenic plants in the model grass species Brachypodium distachyon that expressed cDNAs for sucrose:fructan 6-fructosyltransferases (6-SFTs) with different enzymatic properties: one cDNA encoded PpFT1 from timothy grass (Phleum pratense), an enzyme that produces high-DP levans; a second cDNA encoded wft1 from wheat (Triticum aestivum), an enzyme that produces low-DP levans. Transgenic lines expressing PpFT1 and wft1 showed retarded growth; this effect was particularly notable in the PpFT1 transgenic lines. When grown at 22 °C, both types of transgenic line showed little or no accumulation of fructans. However, after a cold treatment, wft1 transgenic plants accumulated fructans with DP = 3-40, whereas PpFT1 transgenic plants accumulated fructans with higher DPs (20 to the separation limit). The different compositions of the accumulated fructans in the two types of transgenic line were correlated with the differences in the enzymatic properties of the overexpressed 6-SFTs. Transgenic lines expressing PpFT1 accumulated greater amounts of mono- and disaccharides than wild type and wft1 expressing lines. Examination of leaf blades showed that after cold acclimation, PpFT1 overexpression increased tolerance to freezing; by contrast, the freezing tolerance of the wft1 expressing lines was the same as that of wild type plants. These results provide new insights into the relationship of the composition of water-soluble carbohydrates and the DP of fructans to freezing tolerance in plants.

  6. Expression of cartilage developmental genes in Hoxc8- and Hoxd4-transgenic mice.

    Directory of Open Access Journals (Sweden)

    Claudia Kruger

    2010-02-01

    Full Text Available Hox genes encode transcription factors, which regulate skeletal patterning and chondrocyte differentiation during the development of cartilage, the precursor to mature bone. Overexpression of the homeobox transcription factors Hoxc8 and Hoxd4 causes severe cartilage defects due to delay in cartilage maturation. Matrix metalloproteinases (MMPs, bone morphogenetic proteins (BMPs and fibroblastic growth factors (FGFs are known to play important roles in skeletal development and endochondral bone formation and remodeling. In order to investigate whether these molecules are aberrantly expressed in Hoxc8- and/or Hoxd4-transgenic cartilage, we performed quantitative RT-PCR on chondrocytes from Hox-transgenic mice. Gene expression levels of Bmp4, Fgf8, Fgf10, Mmp9, Mmp13, Nos3, Timp3, Wnt3a and Wnt5a were altered in Hoxc8-transgenic chondrocytes, and Fgfr3, Ihh, Mmp8, and Wnt3a expression levels were altered in Hoxd4-transgenic chondrocytes, respectively. Notably, Wnt3a expression was elevated in Hoxc8- and reduced in Hoxd4-transgenic cartilage. These results suggest that both transcription factors affect cartilage maturation through different molecular mechanisms, and provide the basis for future studies into the role of these genes and possible interactions in pathogenesis of cartilage defects in Hoxc8- and Hoxd4-transgenic mice.

  7. Effect of glycogen synthase overexpression on insulin-stimulated muscle glucose uptake and storage.

    Science.gov (United States)

    Fogt, Donovan L; Pan, Shujia; Lee, Sukho; Ding, Zhenping; Scrimgeour, Angus; Lawrence, John C; Ivy, John L

    2004-03-01

    Insulin-stimulated muscle glucose uptake is inversely associated with the muscle glycogen concentration. To investigate whether this association is a cause and effect relationship, we compared insulin-stimulated muscle glucose uptake in noncontracted and postcontracted muscle of GSL3-transgenic and wild-type mice. GSL3-transgenic mice overexpress a constitutively active form of glycogen synthase, which results in an abundant storage of muscle glycogen. Muscle contraction was elicited by in situ electrical stimulation of the sciatic nerve. Right gastrocnemii from GSL3-transgenic and wild-type mice were subjected to 30 min of electrical stimulation followed by hindlimb perfusion of both hindlimbs. Thirty minutes of contraction significantly reduced muscle glycogen concentration in wild-type (49%) and transgenic (27%) mice, although transgenic mice retained 168.8 +/- 20.5 micromol/g glycogen compared with 17.7 +/- 2.6 micromol/g glycogen for wild-type mice. Muscle of transgenic and wild-type mice demonstrated similar pre- (3.6 +/- 0.3 and 3.9 +/- 0.6 micromol.g(-1).h(-1) for transgenic and wild-type, respectively) and postcontraction (7.9 +/- 0.4 and 7.0 +/- 0.4 micromol.g(-1).h(-1) for transgenic and wild-type, respectively) insulin-stimulated glucose uptakes. However, the [14C]glucose incorporated into glycogen was greater in noncontracted (151%) and postcontracted (157%) transgenic muscle vs. muscle of corresponding wild-type mice. These results indicate that glycogen synthase activity is not rate limiting for insulin-stimulated glucose uptake in skeletal muscle and that the inverse relationship between muscle glycogen and insulin-stimulated glucose uptake is an association, not a cause and effect relationship.

  8. Significance of the lipid phase in the dynamics and functions of the xanthophyll cycle as revealed by PsbS overexpression in tobacco and in-vitro de-epoxidation in monogalactosyldiacylglycerol micelles.

    Science.gov (United States)

    Hieber, A David; Kawabata, Osamu; Yamamoto, Harry Y

    2004-01-01

    The dynamics of the xanthophyll cycle relative to non-photochemical quenching (NPQ) were examined in tobacco plants overexpressing violaxanthin de-epoxidase (VDE), PsbS and PsbS+VDE for effects on NPQ and violaxanthin (V) de-epoxidation over a range of light intensities. Induction of de-epoxidation and NPQ increased in overexpressed VDE and PsbS plants, respectively. Surprisingly, under low light, overexpressing PsbS enhanced de-epoxidation in addition to NPQ. The effect was hypothesized as due to PsbS binding zeaxanthin (Z) or inducing the binding of Z within the quenching complex, thus shifting the equilibrium toward higher de-epoxidation states. Studies in model systems show that Z can stereospecifically inhibit VDE activity against violaxanthin. This effect, observed under conditions of limiting lipid concentration, was interpreted as product feedback inhibition. These results support the hypothesis that the capacity of the thylakoid lipid phase for xanthophylls is limited and modulates xanthophyll-cycle activity, in conjunction with the release of V and binding of Z by pigment-binding proteins. These modulating factors are incorporated into a lipid-matrix model that has elements of a signal transduction system wherein the light-generated protons are the signal, VDE the signal receptor, Z the secondary messenger, the lipid phase the transduction network, and Z-binding proteins the targets.

  9. Salivary Gland Derived BDNF Overexpression in Mice Exerts an Anxiolytic Effect

    OpenAIRE

    Saruta, Juri; To, Masahiro; Sugimoto, Masahiro; Yamamoto, Yuko; Shimizu, Tomoko; Nakagawa, Yusuke; Inoue, Hiroko; Saito, Ichiro; Tsukinoki, Keiichi

    2017-01-01

    Brain-derived neurotrophic factor (BDNF) is abundant in the hippocampus and plays critical roles in memory and synapse formation, as well as exerting antidepressant-like effects in psychiatric disorders. We previously reported that BDNF is expressed in salivary glands and affects blood BDNF content. However, the function of salivary BDNF remains unclear. The aim of this study was to generate transgenic mice overexpressing BDNF in the salivary glands. Hence, we used the Lama construct (hemaggl...

  10. Comparative transcriptomic analyses of differentially expressed genes in transgenic melatonin biosynthesis ovine HIOMT gene in switchgrass

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    Shan Yuan

    2016-11-01

    Full Text Available Melatonin serves pleiotropic functions in prompting plant growth and resistance to various stresses. The accurate biosynthetic pathway of melatonin remains elusive in plant species, while the N-acetyltransferase and O-methyltransferase were considered to be the last two key enzymes during its biosynthesis. To investigate the biosynthesis and metabolic pathway of melatonin in plants, the RNA-seq profile of overexpression of the ovine HIOMT was analyzed and compared with the previous transcriptome of transgenic oAANAT gene in switchgrass, a model plant for cellulosic ethanol production. A total of 946, 405 and 807 differentially expressed unigenes were observed in AANAT vs. control, HIOMT vs. control, and AANAT vs. HIOMT, respectively. The significantly upregulated (F-box/kelch-repeat protein, zinc finger BED domain-containing protein-3 genes were consistent with enhanced phenotypes of shoot, stem and root growth in transgenic oHIOMT switchgrass. Early flowering in overexpression of oHIOMT switchgrass involved in the regulation of flowering-time genes (APETALA2. Several stress resistant related genes (SPX domain-containing membrane protein, copper transporter 1, late blight resistance protein homolog R1A-6 OS etc. were specifically and significantly upregulated in transgenic oHIOMT only, while metabolism-related genes (phenylalanine-4-hydroxylase, tyrosine decarboxylase 1, protein disulfide-isomerase and galactinol synthase 2 etc. were significantly upregulated in transgenic oAANAT only. These results provide new sights into the biosynthetic and physiological functional networks of melatonin in plants.

  11. Establishment of a canine model of human type 2 diabetes mellitus by overexpressing phosphoenolypyruvate carboxykinase.

    Science.gov (United States)

    Jeong, Yeon Woo; Lee, Geun-Shik; Kim, Joung Joo; Park, Sun Woo; Ko, Kyeong Hee; Kang, Mina; Kim, Yu Kyung; Jung, Eui-Man; Hyun, Sang Hwan; Shin, Taeyoung; Jeung, Eui-Bae; Hwang, Woo Suk

    2012-08-01

    Dogs are useful models for studying human metabolic diseases such as type 2 diabetes mellitus due to similarities in physiology, anatomy and life styles with humans. Somatic cell nuclear transfer (SCNT) facilitates the production of transgenic dogs. In this study, we generated transgenic dogs expressing the phosphoenolpyruvate carboxykinase (PEPCK) gene, which is closely involved in the pathogenesis of type 2 diabetes mellitus. In addition, we assessed the cloning efficiency associated with adult or fetal (cloned or natural mating) fibroblasts as a nuclear source. Cloning efficiency was determined by the fusion, pregnancy and cloning rates. The fusion rates were significantly high for fibroblasts from cloned fetuses, but the pregnancy and cloning rates were relatively high for cells from normal fetuses. Based on these data, fetal fibroblasts were selected as the nuclear donor for SCNT and genetically engineered to overexpress the PEPCK gene and dual selection marker genes controlled by the PEPCK promoter. The transgenic cells were introduced into oocytes and transferred into five recipient dogs, resulting in two pregnancies. Finally, three puppies were born and confirmed by microsatellite analysis to be genetically identical to the donor. One puppy successfully overexpressed PEPCK mRNA and protein in the liver. This canine disease model may be useful for studying the pathogenesis and/or therapeutic targets of type 2 diabetes mellitus.

  12. Overexpression of BdMATE Gene Improves Aluminum Tolerance in Setaria viridis

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    Ana P. Ribeiro

    2017-06-01

    Full Text Available Acidic soils are distributed worldwide, predominantly in tropical and subtropical areas, reaching around 50% of the arable soil. This type of soil strongly reduces crop production, mainly because of the presence of aluminum, which has its solubility increased at low pH levels. A well-known physiological mechanism used by plants to cope with Al stress involves activation of membrane transporters responsible for organic acid anions secretion from the root apex to the rhizosphere, which chelate Al, preventing its absorption by roots. In sorghum, a membrane transporter gene belonging to multidrug and toxic compound extrusion (MATE family was identified and characterized as an aluminum-activated citrate transporter gene responsible for Al tolerance in this crop. Setaria viridis is an emerging model for C4 species and it is an important model to validate some genes for further C4 crops transformation, such as sugarcane, maize, and wheat. In the present work, Setaria viridis was used as a model plant to overexpress a newly identified MATE gene from Brachypodium distachyon (BdMATE, closely related to SbMATE, for aluminum tolerance assays. Transgenic S. viridis plants overexpressing a BdMATE presented an improved Al tolerance phenotype, characterized by sustained root growth and exclusion of aluminum from the root apex in transgenic plants, as confirmed by hematoxylin assay. In addition, transgenic plants showed higher root citrate exudation into the rhizosphere, suggesting that Al tolerance improvement in these plants could be related to the chelation of the metal by the organic acid anion. These results suggest that BdMATE gene can be used to transform C4 crops of economic importance with improved aluminum tolerance.

  13. Constitutive expression of DaCBF7, an Antarctic vascular plant Deschampsia antarctica CBF homolog, resulted in improved cold tolerance in transgenic rice plants.

    Science.gov (United States)

    Byun, Mi Young; Lee, Jungeun; Cui, Li Hua; Kang, Yoonjee; Oh, Tae Kyung; Park, Hyun; Lee, Hyoungseok; Kim, Woo Taek

    2015-07-01

    Deschampsia antarctica is an Antarctic hairgrass that grows on the west coast of the Antarctic peninsula. In this report, we have identified and characterized a transcription factor, D. antarctica C-repeat binding factor 7 (DaCBF7), that is a member of the monocot group V CBF homologs. The protein contains a single AP2 domain, a putative nuclear localization signal, and the typical CBF signature. DaCBF7, like other monocot group V homologs, contains a distinct polypeptide stretch composed of 43 amino acids in front of the AP2 motif. DaCBF7 was predominantly localized to nuclei and interacted with the C-repeat/dehydration responsive element (CRT/DRE) core sequence (ACCGAC) in vitro. DaCBF7 was induced by abiotic stresses, including drought, cold, and salinity. To investigate its possible cellular role in cold tolerance, a transgenic rice system was employed. DaCBF7-overexpressing transgenic rice plants (Ubi:DaCBF7) exhibited markedly increased tolerance to cold stress compared to wild-type plants without growth defects; however, overexpression of DaCBF7 exerted little effect on tolerance to drought or salt stress. Transcriptome analysis of a Ubi:DaCBF7 transgenic line revealed 13 genes that were up-regulated in DaCBF7-overexpressing plants compared to wild-type plants in the absence of cold stress and in short- or long-term cold stress. Five of these genes, dehydrin, remorin, Os03g63870, Os11g34790, and Os10g22630, contained putative CRT/DRE or low-temperature responsive elements in their promoter regions. These results suggest that overexpression of DaCBF7 directly and indirectly induces diverse genes in transgenic rice plants and confers enhanced tolerance to cold stress. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  14. A three-component gene expression system and its application for inducible flavonoid overproduction in transgenic Arabidopsis thaliana.

    Science.gov (United States)

    Feng, Yue; Cao, Cong-Mei; Vikram, Meenu; Park, Sunghun; Kim, Hye Jin; Hong, Jong Chan; Cisneros-Zevallos, Luis; Koiwa, Hisashi

    2011-03-08

    Inducible gene expression is a powerful tool to study and engineer genes whose overexpression could be detrimental for the host organisms. However, only limited systems have been adopted in plant biotechnology. We have developed an osmotically inducible system using three components of plant origin, RD29a (Responsive to Dehydration 29A) promoter, CBF3 (C-repeat Binding Factor 3) transcription factor and cpl1-2 (CTD phosphatase-like 1) mutation. The osmotic stress responsible RD29a promoter contains the CBF3 binding sites and thus RD29A-CBF3 feedforward cassette enhances induction of RD29a promoter under stress. The cpl1-2 mutation in a host repressor CPL1 promotes stress responsible RD29a promoter expression. The efficacy of this system was tested using PAP1 (Production of Anthocyanin Pigment 1) transgene, a model transcription factor that regulates the anthocyanin pathway in Arabidopsis. While transgenic plants with only one or two of three components did not reproducibly accumulate anthocyanin pigments above the control level, transgenic cpl1 plants containing homozygous RD29a-PAP1 and RD29a-CBF3 transgenes produced 30-fold higher level of total anthocyanins than control plants upon cold treatment. Growth retardation and phytochemical production of transgenic plants were minimum under normal conditions. The flavonoid profile in cold-induced transgenic plants was determined by LC/MS/MS, which resembled that of previously reported pap1-D plants but enriched for kaempferol derivatives. These results establish the functionality of the inducible three-component gene expression system in plant metabolic engineering. Furthermore, we show that PAP1 and environmental signals synergistically regulate the flavonoid pathway to produce a unique flavonoid blend that has not been produced by PAP1 overexpression or cold treatment alone.

  15. A three-component gene expression system and its application for inducible flavonoid overproduction in transgenic Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Yue Feng

    Full Text Available Inducible gene expression is a powerful tool to study and engineer genes whose overexpression could be detrimental for the host organisms. However, only limited systems have been adopted in plant biotechnology. We have developed an osmotically inducible system using three components of plant origin, RD29a (Responsive to Dehydration 29A promoter, CBF3 (C-repeat Binding Factor 3 transcription factor and cpl1-2 (CTD phosphatase-like 1 mutation. The osmotic stress responsible RD29a promoter contains the CBF3 binding sites and thus RD29A-CBF3 feedforward cassette enhances induction of RD29a promoter under stress. The cpl1-2 mutation in a host repressor CPL1 promotes stress responsible RD29a promoter expression. The efficacy of this system was tested using PAP1 (Production of Anthocyanin Pigment 1 transgene, a model transcription factor that regulates the anthocyanin pathway in Arabidopsis. While transgenic plants with only one or two of three components did not reproducibly accumulate anthocyanin pigments above the control level, transgenic cpl1 plants containing homozygous RD29a-PAP1 and RD29a-CBF3 transgenes produced 30-fold higher level of total anthocyanins than control plants upon cold treatment. Growth retardation and phytochemical production of transgenic plants were minimum under normal conditions. The flavonoid profile in cold-induced transgenic plants was determined by LC/MS/MS, which resembled that of previously reported pap1-D plants but enriched for kaempferol derivatives. These results establish the functionality of the inducible three-component gene expression system in plant metabolic engineering. Furthermore, we show that PAP1 and environmental signals synergistically regulate the flavonoid pathway to produce a unique flavonoid blend that has not been produced by PAP1 overexpression or cold treatment alone.

  16. genetic overexpression of NR2B subunit enhances social recognition memory for different strains and species.

    Science.gov (United States)

    Jacobs, Stephanie A; Tsien, Joe Z

    2012-01-01

    The ability to learn and remember conspecifics is essential for the establishment and maintenance of social groups. Many animals, including humans, primates and rodents, depend on stable social relationships for survival. Social learning and social recognition have become emerging areas of interest for neuroscientists but are still not well understood. It has been established that several hormones play a role in the modulation of social recognition including estrogen, oxytocin and arginine vasopression. Relatively few studies have investigated how social recognition might be improved or enhanced. In this study, we investigate the role of the NMDA receptor in social recognition memory, specifically the consequences of altering the ratio of the NR2B:NR2A subunits in the forebrain regions in social behavior. We produced transgenic mice in which the NR2B subunit of the NMDA receptor was overexpressed postnatally in the excitatory neurons of the forebrain areas including the cortex, amygdala and hippocampus. We investigated the ability of both our transgenic animals and their wild-type littermate to learn and remember juvenile conspecifics using both 1-hr and 24-hr memory tests. Our experiments show that the wild-type animals and NR2B transgenic mice preformed similarly in the 1-hr test. However, transgenic mice showed better performances in 24-hr tests of recognizing animals of a different strain or animals of a different species. We conclude that NR2B overexpression in the forebrain enhances social recognition memory for different strains and animal species.

  17. Amyloid-like protein inclusions in tobacco transgenic plants.

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    Anna Villar-Piqué

    Full Text Available The formation of insoluble protein deposits in human tissues is linked to the onset of more than 40 different disorders, ranging from dementia to diabetes. In these diseases, the proteins usually self-assemble into ordered β-sheet enriched aggregates known as amyloid fibrils. Here we study the structure of the inclusions formed by maize transglutaminase (TGZ in the chloroplasts of tobacco transplastomic plants and demonstrate that they have an amyloid-like nature. Together with the evidence of amyloid structures in bacteria and fungi our data argue that amyloid formation is likely a ubiquitous process occurring across the different kingdoms of life. The discovery of amyloid conformations inside inclusions of genetically modified plants might have implications regarding their use for human applications.

  18. Tapetum development in transgenic tobacco (Nicotiana tabacum L. plants with modlfied level of histone H1 variants

    Directory of Open Access Journals (Sweden)

    Joanna Ślusarczyk

    2011-01-01

    Full Text Available The phenomenon of male sterility has often been observed in investigations on the role of histone H1 in regulation of morphogenetic and cytological processes in transgenic tobacco plants. These changes were accumulated by disturbances in flower development, consisting in lengthening of the pistil style in relation to stamen heads. This prevented pollination and production of seeds. As similar abnormalities occurred also in the present investigations (depending on combination, the sterility% was 84.4 to 19.9, at only 8.1 in the control, the main problem of our investigations was an attempt to explain their reasons. It is commonly known that one of the conditions for formation of fertile pollen is the properly functioning tapetum. Here, we carried out observations of ultrastructure of anther tapetum control cells in respect of abnormalities which occurred during microsporogenesis of transgenic plants with inactivated expression of two major (A, B and two minor (C, D histone H1 variants. The investigations were carried out on the following groups of plants: (1 control group with a full set of histone variants (K, (2 with inactivated A and B variants (-AB; (3 with inactivated A, B, C and D variants (-ABCD, (4 with inactivated C and D variants (-CD. It was found that tapetal development was normal in all the investigated groups of plants, and the sequence of changes was similar as in the control. However, certain ultrastructural differences appeared when tapetum functioned as secretory tissue, and in the degeneration phase. In tapetal cell cytoplasm, with participation of rER, lipid bodies were formed, which, having penetrated to the cell surface and to locules, took part in formation of pollen grain sporoderm. Both in the control and in the remaining combination, excluding -ABCD, these bodies looked similar: they were grey, homogenous and surrounded by black jagged deposits. In -ABCD plants, these bodies were more translucent, slightly rarefied, and

  19. Functional characterization of secondary wall deposition regulating transcription factors MusaVND2 and MusaVND3 in transgenic banana plants.

    Science.gov (United States)

    Negi, Sanjana; Tak, Himanshu; Ganapathi, T R

    2016-03-01

    NAM, ATAF, and CUC (NAC) domain-containing proteins are plant-specific transcription factors involved in stress responses and developmental regulation. MusaVND2 and MusaVND3 are vascular-related NAC domain-containing genes encoding for nuclear-localized proteins. The transcript level of MusaVND2 and MusaVND3 are gradually induced after induction of lignification conditions in banana embryogenic cells. Banana embryogenic cells differentiated to tracheary element-like cells after overexpression of MusaVND2 and MusaVND3 with a differentiation frequency of 63.5 and 23.4 %, respectively, after ninth day. Transgenic banana plants overexpressing either of MusaVND2 or MusaVND3 showed ectopic secondary wall deposition as well as transdifferentiation of cells into tracheary elements. Transdifferentiation to tracheary element-like cells was observed in cortical cells of corm and in epidermal and mesophyll cells of leaves of transgenic plants. Elevated levels of lignin and crystalline cellulose were detected in the transgenic banana lines than control plants. The results obtained are useful for understanding the molecular regulation of secondary wall development in banana.

  20. Overexpression of TaLEA gene from Tamarix androssowii improves salt and drought tolerance in transgenic poplar (Populus simonii × P. nigra.

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    Weidong Gao

    Full Text Available Late embryogenesis abundant (LEA genes were confirmed to confer resistance to drought and water deficiency. An LEA gene from Tamarixandrossowii (named TaLEA was transformed into Xiaohei poplar (Populussimonii × P. nigra via Agrobacterium. Twenty-five independent transgenic lines were obtained that were resistant to kanamycin, and 11 transgenic lines were randomly selected for further analysis. The polymerase chain reaction (PCR and ribonucleic acid (RNA gel blot indicated that the TaLEA gene had been integrated into the poplar genome. The height growth rate, malondialdehyde (MDA content, relative electrolyte leakage and damages due to salt or drought to transgenic and non-transgenic plants were compared under salt and drought stress conditions. The results showed that the constitutive expression of the TaLEA gene in transgenic poplars could induce an increase in height growth rate and a decrease in number and severity of wilted leaves under the salt and drought stresses. The MDA content and relative electrolyte leakage in transgenic lines under salt and drought stresses were significantly lower compared to those in non-transgenic plants, indicating that the TaLEA gene may enhance salt and drought tolerance by protecting cell membranes from damage. Moreover, amongst the lines analyzed for stress tolerance, the transgenic line 11 (T11 showed the highest tolerance levels under both salinity and drought stress conditions. These results indicated that the TaLEA gene could be a salt and drought tolerance candidate gene and could confer a broad spectrum of tolerance under abiotic stresses in poplars.

  1. Overexpression of TaLEA gene from Tamarix androssowii improves salt and drought tolerance in transgenic poplar (Populus simonii × P. nigra).

    Science.gov (United States)

    Gao, Weidong; Bai, Shuang; Li, Qingmei; Gao, Caiqiu; Liu, Guifeng; Li, Guangde; Tan, Feili

    2013-01-01

    Late embryogenesis abundant (LEA) genes were confirmed to confer resistance to drought and water deficiency. An LEA gene from Tamarixandrossowii (named TaLEA) was transformed into Xiaohei poplar (Populussimonii × P. nigra) via Agrobacterium. Twenty-five independent transgenic lines were obtained that were resistant to kanamycin, and 11 transgenic lines were randomly selected for further analysis. The polymerase chain reaction (PCR) and ribonucleic acid (RNA) gel blot indicated that the TaLEA gene had been integrated into the poplar genome. The height growth rate, malondialdehyde (MDA) content, relative electrolyte leakage and damages due to salt or drought to transgenic and non-transgenic plants were compared under salt and drought stress conditions. The results showed that the constitutive expression of the TaLEA gene in transgenic poplars could induce an increase in height growth rate and a decrease in number and severity of wilted leaves under the salt and drought stresses. The MDA content and relative electrolyte leakage in transgenic lines under salt and drought stresses were significantly lower compared to those in non-transgenic plants, indicating that the TaLEA gene may enhance salt and drought tolerance by protecting cell membranes from damage. Moreover, amongst the lines analyzed for stress tolerance, the transgenic line 11 (T11) showed the highest tolerance levels under both salinity and drought stress conditions. These results indicated that the TaLEA gene could be a salt and drought tolerance candidate gene and could confer a broad spectrum of tolerance under abiotic stresses in poplars.

  2. Endothelial Nitric Oxide Synthase Overexpression Restores the Efficiency of Bone Marrow Mononuclear Cell-Based Therapy

    Science.gov (United States)

    Mees, Barend; Récalde, Alice; Loinard, Céline; Tempel, Dennie; Godinho, Marcia; Vilar, José; van Haperen, Rien; Lévy, Bernard; de Crom, Rini; Silvestre, Jean-Sébastien

    2011-01-01

    Bone marrow-derived mononuclear cells (BMMNCs) enhance postischemic neovascularization, and their therapeutic use is currently under clinical investigation. However, cardiovascular risk factors, including diabetes mellitus and hypercholesterolemia, lead to the abrogation of BMMNCs proangiogenic potential. NO has been shown to be critical for the proangiogenic function of BMMNCs, and increased endothelial NO synthase (eNOS) activity promotes vessel growth in ischemic conditions. We therefore hypothesized that eNOS overexpression could restore both the impaired neovascularization response and decreased proangiogenic function of BMMNCs in clinically relevant models of diabetes and hypercholesterolemia. Transgenic eNOS overexpression in diabetic, atherosclerotic, and wild-type mice induced a 1.5- to 2.3-fold increase in postischemic neovascularization compared with control. eNOS overexpression in diabetic or atherosclerotic BMMNCs restored their reduced proangiogenic potential in ischemic hind limb. This effect was associated with an increase in BMMNC ability to differentiate into cells with endothelial phenotype in vitro and in vivo and an increase in BMMNCs paracrine function, including vascular endothelial growth factor A release and NO-dependent vasodilation. Moreover, although wild-type BMMNCs treatment resulted in significant progression of atherosclerotic plaque in ischemic mice, eNOS transgenic atherosclerotic BMMNCs treatment even had antiatherogenic effects. Cell-based eNOS gene therapy has both proangiogenic and antiatherogenic effects and should be further investigated for the development of efficient therapeutic neovascularization designed to treat ischemic cardiovascular disease. PMID:21224043

  3. Transgenic expression of BRCA1 disturbs hematopoietic stem and progenitor cells quiescence and function

    International Nuclear Information System (INIS)

    Bai, Lin; Shi, Guiying; Zhang, Xu; Dong, Wei; Zhang, Lianfeng

    2013-01-01

    The balance between quiescence and proliferation of HSCs is an important regulator of hematopoiesis. Loss of quiescence frequently results in HSCs exhaustion, which underscores the importance of tight regulation of proliferation in these cells. Studies have indicated that cyclin-dependent kinases are involved in the regulation of quiescence in HSCs. BRCA1 plays an important role in the repair of DNA double-stranded breaks, cell cycle, apoptosis and transcription. BRCA1 is expressed in the bone marrow. However, the function of BRCA1 in HSCs is unknown. In our study, we generated BRCA1 transgenic mice to investigate the effects of BRCA1 on the mechanisms of quiescence and differentiation in HSCs. The results demonstrate that over-expression of BRCA1 in the bone marrow impairs the development of B lymphocytes. Furthermore, BRCA1 induced an increase in the number of LSKs, LT-HSCs, ST-HSCs and MPPs. A competitive transplantation assay found that BRCA1 transgenic mice failed to reconstitute hematopoiesis. Moreover, BRCA1 regulates the expression of p21 waf1 /cip1 and p57 kip2 , which results in a loss of quiescence in LSKs. Together, over-expression of BRCA1 in bone marrow disrupted the quiescent of LSKs, induced excessive accumulation of LSKs, and disrupted differentiation of the HSCs, which acts through the down-regulated of p21 waf1 /cip1 and p57 kip2 . - Highlights: • Over-expression of BRCA1 results in impaired B lymphocyte development. • BRCA1 transgenic mice disrupted the quiescent of LSKs, induced excessive accumulation of LSKs. • BRCA1 impairs the function of HSCs through the down-regulated of p21 waf1/cip1 and p57 kip2

  4. Transgenic expression of BRCA1 disturbs hematopoietic stem and progenitor cells quiescence and function

    Energy Technology Data Exchange (ETDEWEB)

    Bai, Lin; Shi, Guiying; Zhang, Xu; Dong, Wei; Zhang, Lianfeng, E-mail: zhanglf@cnilas.org

    2013-10-15

    The balance between quiescence and proliferation of HSCs is an important regulator of hematopoiesis. Loss of quiescence frequently results in HSCs exhaustion, which underscores the importance of tight regulation of proliferation in these cells. Studies have indicated that cyclin-dependent kinases are involved in the regulation of quiescence in HSCs. BRCA1 plays an important role in the repair of DNA double-stranded breaks, cell cycle, apoptosis and transcription. BRCA1 is expressed in the bone marrow. However, the function of BRCA1 in HSCs is unknown. In our study, we generated BRCA1 transgenic mice to investigate the effects of BRCA1 on the mechanisms of quiescence and differentiation in HSCs. The results demonstrate that over-expression of BRCA1 in the bone marrow impairs the development of B lymphocytes. Furthermore, BRCA1 induced an increase in the number of LSKs, LT-HSCs, ST-HSCs and MPPs. A competitive transplantation assay found that BRCA1 transgenic mice failed to reconstitute hematopoiesis. Moreover, BRCA1 regulates the expression of p21{sup waf1}/cip1 and p57{sup kip2}, which results in a loss of quiescence in LSKs. Together, over-expression of BRCA1 in bone marrow disrupted the quiescent of LSKs, induced excessive accumulation of LSKs, and disrupted differentiation of the HSCs, which acts through the down-regulated of p21{sup waf1}/cip1 and p57{sup kip2}. - Highlights: • Over-expression of BRCA1 results in impaired B lymphocyte development. • BRCA1 transgenic mice disrupted the quiescent of LSKs, induced excessive accumulation of LSKs. • BRCA1 impairs the function of HSCs through the down-regulated of p21{sup waf1/cip1} and p57{sup kip2}.

  5. AAV-mediated pancreatic overexpression of Igf1 counteracts progression to autoimmune diabetes in mice.

    Science.gov (United States)

    Mallol, Cristina; Casana, Estefania; Jimenez, Veronica; Casellas, Alba; Haurigot, Virginia; Jambrina, Claudia; Sacristan, Victor; Morró, Meritxell; Agudo, Judith; Vilà, Laia; Bosch, Fatima

    2017-07-01

    Type 1 diabetes is characterized by autoimmune destruction of β-cells leading to severe insulin deficiency. Although many improvements have been made in recent years, exogenous insulin therapy is still imperfect; new therapeutic approaches, focusing on preserving/expanding β-cell mass and/or blocking the autoimmune process that destroys islets, should be developed. The main objective of this work was to test in non-obese diabetic (NOD) mice, which spontaneously develop autoimmune diabetes, the effects of local expression of Insulin-like growth factor 1 (IGF1), a potent mitogenic and pro-survival factor for β-cells with immunomodulatory properties. Transgenic NOD mice overexpressing IGF1 specifically in β-cells (NOD-IGF1) were generated and phenotyped. In addition, miRT-containing, IGF1-encoding adeno-associated viruses (AAV) of serotype 8 (AAV8-IGF1-dmiRT) were produced and administered to 4- or 11-week-old non-transgenic NOD females through intraductal delivery. Several histological, immunological, and metabolic parameters were measured to monitor disease over a period of 28-30 weeks. In transgenic mice, local IGF1 expression led to long-term suppression of diabetes onset and robust protection of β-cell mass from the autoimmune insult. AAV-mediated pancreatic-specific overexpression of IGF1 in adult animals also dramatically reduced diabetes incidence, both when vectors were delivered before pathology onset or once insulitis was established. Transgenic NOD-IGF1 and AAV8-IGF1-dmiRT-treated NOD animals had much less islet infiltration than controls, preserved β-cell mass, and normal insulinemia. Transgenic and AAV-treated islets showed less expression of antigen-presenting molecules, inflammatory cytokines, and chemokines important for tissue-specific homing of effector T cells, suggesting IGF1 modulated islet autoimmunity in NOD mice. Local expression of Igf1 by AAV-mediated gene transfer counteracts progression to diabetes in NOD mice. This study suggests a

  6. The Cotton WRKY Gene GhWRKY41 Positively Regulates Salt and Drought Stress Tolerance in Transgenic Nicotiana benthamiana.

    Directory of Open Access Journals (Sweden)

    Xiaoqian Chu

    Full Text Available WRKY transcription factors constitute a very large family of proteins in plants and participate in modulating plant biological processes, such as growth, development and stress responses. However, the exact roles of WRKY proteins are unclear, particularly in non-model plants. In this study, Gossypium hirsutum WRKY41 (GhWRKY41 was isolated and transformed into Nicotiana benthamiana. Our results showed that overexpression of GhWRKY41 enhanced the drought and salt stress tolerance of transgenic Nicotiana benthamiana. The transgenic plants exhibited lower malondialdehyde content and higher antioxidant enzyme activity, and the expression of antioxidant genes was upregulated in transgenic plants exposed to osmotic stress. A β-glucuronidase (GUS staining assay showed that GhWRKY41 was highly expressed in the stomata when plants were exposed to osmotic stress, and plants overexpressing GhWRKY41 exhibited enhanced stomatal closure when they were exposed to osmotic stress. Taken together, our findings demonstrate that GhWRKY41 may enhance plant tolerance to stress by functioning as a positive regulator of stoma closure and by regulating reactive oxygen species (ROS scavenging and the expression of antioxidant genes.

  7. Potato Annexin STANN1 Promotes Drought Tolerance and Mitigates Light Stress in Transgenic Solanum tuberosum L. Plants

    Science.gov (United States)

    Szalonek, Michal; Sierpien, Barbara; Rymaszewski, Wojciech; Gieczewska, Katarzyna; Garstka, Maciej; Lichocka, Malgorzata; Sass, Laszlo; Paul, Kenny; Vass, Imre; Vankova, Radomira; Dobrev, Peter; Szczesny, Pawel; Marczewski, Waldemar; Krusiewicz, Dominika; Strzelczyk-Zyta, Danuta; Hennig, Jacek; Konopka-Postupolska, Dorota

    2015-01-01

    Annexins are a family of calcium- and membrane-binding proteins that are important for plant tolerance to adverse environmental conditions. Annexins function to counteract oxidative stress, maintain cell redox homeostasis, and enhance drought tolerance. In the present study, an endogenous annexin, STANN1, was overexpressed to determine whether crop yields could be improved in potato (Solanum tuberosum L.) during drought. Nine potential potato annexins were identified and their expression characterized in response to drought treatment. STANN1 mRNA was constitutively expressed at a high level and drought treatment strongly increased transcription levels. Therefore, STANN1 was selected for overexpression analysis. Under drought conditions, transgenic potato plants ectopically expressing STANN1 were more tolerant to water deficit in the root zone, preserved more water in green tissues, maintained chloroplast functions, and had higher accumulation of chlorophyll b and xanthophylls (especially zeaxanthin) than wild type (WT). Drought-induced reductions in the maximum efficiency and the electron transport rate of photosystem II (PSII), as well as the quantum yield of photosynthesis, were less pronounced in transgenic plants overexpressing STANN1 than in the WT. This conferred more efficient non-photochemical energy dissipation in the outer antennae of PSII and probably more efficient protection of reaction centers against photooxidative damage in transgenic plants under drought conditions. Consequently, these plants were able to maintain effective photosynthesis during drought, which resulted in greater productivity than WT plants despite water scarcity. Although the mechanisms underlying this stress protection are not yet clear, annexin-mediated photoprotection is probably linked to protection against light-induced oxidative stress. PMID:26172952

  8. Potato Annexin STANN1 Promotes Drought Tolerance and Mitigates Light Stress in Transgenic Solanum tuberosum L. Plants.

    Science.gov (United States)

    Szalonek, Michal; Sierpien, Barbara; Rymaszewski, Wojciech; Gieczewska, Katarzyna; Garstka, Maciej; Lichocka, Malgorzata; Sass, Laszlo; Paul, Kenny; Vass, Imre; Vankova, Radomira; Dobrev, Peter; Szczesny, Pawel; Marczewski, Waldemar; Krusiewicz, Dominika; Strzelczyk-Zyta, Danuta; Hennig, Jacek; Konopka-Postupolska, Dorota

    2015-01-01

    Annexins are a family of calcium- and membrane-binding proteins that are important for plant tolerance to adverse environmental conditions. Annexins function to counteract oxidative stress, maintain cell redox homeostasis, and enhance drought tolerance. In the present study, an endogenous annexin, STANN1, was overexpressed to determine whether crop yields could be improved in potato (Solanum tuberosum L.) during drought. Nine potential potato annexins were identified and their expression characterized in response to drought treatment. STANN1 mRNA was constitutively expressed at a high level and drought treatment strongly increased transcription levels. Therefore, STANN1 was selected for overexpression analysis. Under drought conditions, transgenic potato plants ectopically expressing STANN1 were more tolerant to water deficit in the root zone, preserved more water in green tissues, maintained chloroplast functions, and had higher accumulation of chlorophyll b and xanthophylls (especially zeaxanthin) than wild type (WT). Drought-induced reductions in the maximum efficiency and the electron transport rate of photosystem II (PSII), as well as the quantum yield of photosynthesis, were less pronounced in transgenic plants overexpressing STANN1 than in the WT. This conferred more efficient non-photochemical energy dissipation in the outer antennae of PSII and probably more efficient protection of reaction centers against photooxidative damage in transgenic plants under drought conditions. Consequently, these plants were able to maintain effective photosynthesis during drought, which resulted in greater productivity than WT plants despite water scarcity. Although the mechanisms underlying this stress protection are not yet clear, annexin-mediated photoprotection is probably linked to protection against light-induced oxidative stress.

  9. Potato Annexin STANN1 Promotes Drought Tolerance and Mitigates Light Stress in Transgenic Solanum tuberosum L. Plants.

    Directory of Open Access Journals (Sweden)

    Michal Szalonek

    Full Text Available Annexins are a family of calcium- and membrane-binding proteins that are important for plant tolerance to adverse environmental conditions. Annexins function to counteract oxidative stress, maintain cell redox homeostasis, and enhance drought tolerance. In the present study, an endogenous annexin, STANN1, was overexpressed to determine whether crop yields could be improved in potato (Solanum tuberosum L. during drought. Nine potential potato annexins were identified and their expression characterized in response to drought treatment. STANN1 mRNA was constitutively expressed at a high level and drought treatment strongly increased transcription levels. Therefore, STANN1 was selected for overexpression analysis. Under drought conditions, transgenic potato plants ectopically expressing STANN1 were more tolerant to water deficit in the root zone, preserved more water in green tissues, maintained chloroplast functions, and had higher accumulation of chlorophyll b and xanthophylls (especially zeaxanthin than wild type (WT. Drought-induced reductions in the maximum efficiency and the electron transport rate of photosystem II (PSII, as well as the quantum yield of photosynthesis, were less pronounced in transgenic plants overexpressing STANN1 than in the WT. This conferred more efficient non-photochemical energy dissipation in the outer antennae of PSII and probably more efficient protection of reaction centers against photooxidative damage in transgenic plants under drought conditions. Consequently, these plants were able to maintain effective photosynthesis during drought, which resulted in greater productivity than WT plants despite water scarcity. Although the mechanisms underlying this stress protection are not yet clear, annexin-mediated photoprotection is probably linked to protection against light-induced oxidative stress.

  10. An engineered pathway for glyoxylate metabolism in tobacco plants aimed to avoid the release of ammonia in photorespiration

    Directory of Open Access Journals (Sweden)

    Carvalho Josirley de FC

    2011-11-01

    Full Text Available Abstract Background The photorespiratory nitrogen cycle in C3 plants involves an extensive diversion of carbon and nitrogen away from the direct pathways of assimilation. The liberated ammonia is re-assimilated, but up to 25% of the carbon may be released into the atmosphere as CO2. Because of the loss of CO2 and high energy costs, there has been considerable interest in attempts to decrease the flux through the cycle in C3 plants. Transgenic tobacco plants were generated that contained the genes gcl and hyi from E. coli encoding glyoxylate carboligase (EC 4.1.1.47 and hydroxypyruvate isomerase (EC 5.3.1.22 respectively, targeted to the peroxisomes. It was presumed that the two enzymes could work together and compete with the aminotransferases that convert glyoxylate to glycine, thus avoiding ammonia production in the photorespiratory nitrogen cycle. Results When grown in ambient air, but not in elevated CO2, the transgenic tobacco lines had a distinctive phenotype of necrotic lesions on the leaves. Three of the six lines chosen for a detailed study contained single copies of the gcl gene, two contained single copies of both the gcl and hyi genes and one line contained multiple copies of both gcl and hyi genes. The gcl protein was detected in the five transgenic lines containing single copies of the gcl gene but hyi protein was not detected in any of the transgenic lines. The content of soluble amino acids including glycine and serine, was generally increased in the transgenic lines growing in air, when compared to the wild type. The content of soluble sugars, glucose, fructose and sucrose in the shoot was decreased in transgenic lines growing in air, consistent with decreased carbon assimilation. Conclusions Tobacco plants have been generated that produce bacterial glyoxylate carboligase but not hydroxypyruvate isomerase. The transgenic plants exhibit a stress response when exposed to air, suggesting that some glyoxylate is diverted away from

  11. Mice overexpressing both non-mutated human SOD1 and mutated SOD1G93A genes: a competent experimental model for studying iron metabolism in amyotrophic lateral sclerosis

    Directory of Open Access Journals (Sweden)

    Anna eGajowiak

    2016-01-01

    Full Text Available Amyotrophic lateral sclerosis (ALS is a progressive neurodegenerative disease characterized by degeneration and loss of motor neurons in the spinal cord, brainstem and motor cortex. Up to 10% of ALS cases are inherited (familial, fALS and associated with mutations, frequently in the superoxide dismutase 1 (SOD1 gene. Rodent transgenic models of ALS are often used to elucidate a complex pathogenesis of this disease. Of importance, both ALS patients and animals carrying mutated human SOD1 gene show symptoms of oxidative stress and iron metabolism misregulation. The aim of our study was to characterize changes in iron metabolism in one of the most commonly used models of ALS – transgenic mice overexpressing human mutated SOD1G93A gene. We analyzed the expression of iron-related genes in asymptomatic, 2-month old and symptomatic, 4-month old SOD1G93A mice. In parallel, respective age-matched mice overexpressing human non-mutated SOD1 transgene and control mice were analyzed. We demonstrate that the overexpression of both SOD1 and SOD1G93A genes account for a substantial increase in SOD1 protein levels and activity in selected tissues and that not all the changes in iron metabolism genes expression are specific for the overexpression of the mutated form of SOD1.

  12. Impaired growth of pancreatic exocrine cells in transgenic mice expressing human activin βE subunit

    International Nuclear Information System (INIS)

    Hashimoto, Osamu; Ushiro, Yuuki; Sekiyama, Kazunari; Yamaguchi, Osamu; Yoshioka, Kazuki; Mutoh, Ken-Ichiro; Hasegawa, Yoshihisa

    2006-01-01

    Activins, TGF-β superfamily members, have multiple functions in a variety of cells and tissues. Recently, additional activin β subunit genes, βC and βE, have been identified. To explore the role of activin E, we created transgenic mice overexpressing human activin βE subunit. There were pronounced differences in the pancreata of the transgenic animals as compared with their wild-type counterparts. Pancreatic weight, expressed relative to total body weight, was significantly reduced. Histologically, adipose replacement of acini in the exocrine pancreas was observed. There was a significant decrease in the number of PCNA-positive cells in the acinar cells, indicating reduced proliferation in the exocrine pancreas of the transgenic mice. However, quantitative pancreatic morphometry showed that the total number and mass of the islets of the transgenic mice were comparable with those of the nontransgenic control mice. Our findings suggest a role for activin E in regulating the proliferation of pancreatic exocrine cells

  13. Triggered Firing and Atrial Fibrillation in Transgenic Mice With Selective Atrial Fibrosis Induced by Overexpression of TGF-β1

    Science.gov (United States)

    Choi, Eue-Keun; Chang, Po-Cheng; Lee, Young-Soo; Lin, Shien-Fong; Zhu, Wuqiang; Maruyama, Mitsunori; Fishbein, Michael C.; Chen, Zhenhui; der Lohe, Michael Rubart-von; Field, Loren J.; Chen, Peng-Sheng

    2013-01-01

    Background Calcium transient triggered firing (CTTF) is induced by large intracellular calcium (Cai) transient and short action potential duration (APD). We hypothesized that CTTF underlies the mechanisms of early afterdepolarization (EAD) and spontaneous recurrent atrial fibrillation (AF) in transgenic (Tx) mice with overexpression of transforming growth factor β1 (TGF-β1). Methods and Results MHC-TGFcys33ser Tx mice develop atrial fibrosis because of elevated levels of TGF-β1. We studied membrane potential and Cai transients of isolated superfused atria from Tx and wild-type (Wt) littermates. Short APD and persistently elevated Cai transients promoted spontaneous repetitive EADs, triggered activity and spontaneous AF after cessation of burst pacing in Tx but not Wt atria (39% vs. 0%, P=0.008). We were able to map optically 4 episodes of spontaneous AF re-initiation. All first and second beats of spontaneous AF originated from the right atrium (4/4, 100%), which is more severely fibrotic than the left atrium. Ryanodine and thapsigargin inhibited spontaneous re-initiation of AF in all 7 Tx atria tested. Western blotting showed no significant changes of calsequestrin or sarco/endoplasmic reticulum Ca2+-ATPase 2a. Conclusions Spontaneous AF may occur in the Tx atrium because of CTTF, characterized by APD shortening, prolonged Cai transient, EAD and triggered activity. Inhibition of Ca2+ release from the sarcoplasmic reticulum suppressed spontaneous AF. Our results indicate that CTTF is an important arrhythmogenic mechanism in TGF-β1 Tx atria. PMID:22447020

  14. Paramutation of tobacco transgenes by small RNA-mediated transcriptional gene silencing

    Czech Academy of Sciences Publication Activity Database

    Crhák Khaitová, Lucie; Fojtová, M.; Křížová, Kateřina; Lunerová Bedřichová, Jana; Fulneček, Jaroslav; Depicker, A.; Kovařík, Aleš

    2011-01-01

    Roč. 6, č. 5 (2011), s. 650-660 ISSN 1559-2294 R&D Projects: GA ČR(CZ) GD204/09/H002; GA MŠk(CZ) LC06004 Grant - others:GA ČR(CZ) GPP501/11/P667 Program:GP Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : transcriptional gene silencing * transgene epialleles * DNA methylation Subject RIV: BO - Biophysics Impact factor: 4.318, year: 2011

  15. High levels of bioplastic are produced in fertile transplastomic tobacco plants engineered with a synthetic operon for the production of polyhydroxybutyrate.

    Science.gov (United States)

    Bohmert-Tatarev, Karen; McAvoy, Susan; Daughtry, Sean; Peoples, Oliver P; Snell, Kristi D

    2011-04-01

    An optimized genetic construct for plastid transformation of tobacco (Nicotiana tabacum) for the production of the renewable, biodegradable plastic polyhydroxybutyrate (PHB) was designed using an operon extension strategy. Bacterial genes encoding the PHB pathway enzymes were selected for use in this construct based on their similarity to the codon usage and GC content of the tobacco plastome. Regulatory elements with limited homology to the host plastome yet known to yield high levels of plastidial recombinant protein production were used to enhance the expression of the transgenes. A partial transcriptional unit, containing genes of the PHB pathway and a selectable marker gene encoding spectinomycin resistance, was flanked at the 5' end by the host plant's psbA coding sequence and at the 3' end by the host plant's 3' psbA untranslated region. This design allowed insertion of the transgenes into the plastome as an extension of the psbA operon, rendering the addition of a promoter to drive the expression of the transgenes unnecessary. Transformation of the optimized construct into tobacco and subsequent spectinomycin selection of transgenic plants yielded T0 plants that were capable of producing up to 18.8% dry weight PHB in samples of leaf tissue. These plants were fertile and produced viable seed. T1 plants producing up to 17.3% dry weight PHB in samples of leaf tissue and 8.8% dry weight PHB in the total biomass of the plant were also isolated.

  16. Development of transgenic crops based on photo-biotechnology.

    Science.gov (United States)

    Ganesan, Markkandan; Lee, Hyo-Yeon; Kim, Jeong-Il; Song, Pill-Soon

    2017-11-01

    The phenotypes associated with plant photomorphogenesis such as the suppressed shade avoidance response and de-etiolation offer the potential for significant enhancement of crop yields. Of many light signal transducers and transcription factors involved in the photomorphogenic responses of plants, this review focuses on the transgenic overexpression of the photoreceptor genes at the uppermost stream of the signalling events, particularly phytochromes, crytochromes and phototropins as the transgenes for the genetic engineering of crops with improved harvest yields. In promoting the harvest yields of crops, the photoreceptors mediate the light regulation of photosynthetically important genes, and the improved yields often come with the tolerance to abiotic stresses such as drought, salinity and heavy metal ions. As a genetic engineering approach, the term photo-biotechnology has been coined to convey the idea that the greater the photosynthetic efficiency that crop plants can be engineered to possess, the stronger the resistance to biotic and abiotic stresses. Development of GM crops based on photoreceptor transgenes (mainly phytochromes, crytochromes and phototropins) is reviewed with the proposal of photo-biotechnology that the photoreceptors mediate the light regulation of photosynthetically important genes, and the improved yields often come with the added benefits of crops' tolerance to environmental stresses. © 2016 John Wiley & Sons Ltd.

  17. Overexpression of avenin-like b proteins in bread wheat (Triticum aestivum L.) improves dough mixing properties by their incorporation into glutenin polymers.

    Science.gov (United States)

    Ma, Fengyun; Li, Miao; Li, Tingting; Liu, Wei; Liu, Yunyi; Li, Yin; Hu, Wei; Zheng, Qian; Wang, Yaqiong; Li, Kexiu; Chang, Junli; Chen, Mingjie; Yang, Guangxiao; Wang, Yuesheng; He, Guangyuan

    2013-01-01

    Avenin-like b proteins are a small family of wheat storage proteins, each containing 18 or 19 cysteine residues. The role of these proteins, with high numbers of cysteine residues, in determining the functional properties of wheat flour is unclear. In the present study, two transgenic lines of the bread wheat overexpressing avenin-like b gene were generated to investigate the effects of Avenin-like b proteins on dough mixing properties. Sodium dodecyl sulfate sedimentation (SDSS) test and Mixograph analysis of these lines demonstrated that overexpression of Avenin-like b proteins in both transgenic wheat lines significantly increased SDSS volume and improved dough elasticity, mixing tolerance and resistance to extension. These changes were associated with the increased proportion of polymeric proteins due to the incorporation of overexpressed Avenin-like b proteins into the glutenin polymers. The results of this study were critical to confirm the hypothesis that Avenin-like b proteins could be integrated into glutenin polymers by inter-chain disulphide bonds, which could help understand the mechanism behind strengthening wheat dough strength.

  18. Overexpression of avenin-like b proteins in bread wheat (Triticum aestivum L. improves dough mixing properties by their incorporation into glutenin polymers.

    Directory of Open Access Journals (Sweden)

    Fengyun Ma

    Full Text Available Avenin-like b proteins are a small family of wheat storage proteins, each containing 18 or 19 cysteine residues. The role of these proteins, with high numbers of cysteine residues, in determining the functional properties of wheat flour is unclear. In the present study, two transgenic lines of the bread wheat overexpressing avenin-like b gene were generated to investigate the effects of Avenin-like b proteins on dough mixing properties. Sodium dodecyl sulfate sedimentation (SDSS test and Mixograph analysis of these lines demonstrated that overexpression of Avenin-like b proteins in both transgenic wheat lines significantly increased SDSS volume and improved dough elasticity, mixing tolerance and resistance to extension. These changes were associated with the increased proportion of polymeric proteins due to the incorporation of overexpressed Avenin-like b proteins into the glutenin polymers. The results of this study were critical to confirm the hypothesis that Avenin-like b proteins could be integrated into glutenin polymers by inter-chain disulphide bonds, which could help understand the mechanism behind strengthening wheat dough strength.

  19. Overexpression of galectin-7 in mouse epidermis leads to loss of cell junctions and defective skin repair.

    Directory of Open Access Journals (Sweden)

    Gaëlle Gendronneau

    Full Text Available The proteins of the galectin family are implicated in many cellular processes, including cell interactions, polarity, intracellular trafficking, and signal transduction. In human and mouse, galectin-7 is almost exclusively expressed in stratified epithelia, notably in the epidermis. Galectin-7 expression is also altered in several human tumors of epithelial origin. This study aimed at dissecting the consequences of galectin-7 overexpression on epidermis structure and functions in vivo.We established transgenic mice specifically overexpressing galectin-7 in the basal epidermal keratinocytes and analyzed the consequences on untreated skin and after UVB irradiation or mechanical injury.The intercellular cohesion of the epidermis is impaired in transgenic animals, with gaps developing between adjacent keratinocytes, associated with loss of adherens junctions. The epidermal architecture is aberrant with perturbations in the multilayered cellular organisation of the tissue, and structural defects in the basement membrane. These transgenic animals displayed a reduced re-epithelialisation potential following superficial wound, due to a defective collective migration of keratinocytes. Finally, a single mild dose of UVB induced an abnormal apoptotic response in the transgenic epidermis.These results indicate that an excess of galectin-7 leads to a destabilisation of adherens junctions associated with defects in epidermal repair. As this phenotype shares similarities with that of galectin-7 null mutant mice, we conclude that a critical level of this protein is required for maintaining proper epidermal homeostasis. This study brings new insight into the mode of action of galectins in normal and pathological situations.

  20. Mammalian cytochrome CYP2E1 triggered differential gene regulation in response to trichloroethylene (TCE) in a transgenic poplar.

    Science.gov (United States)

    Kang, Jun Won; Wilkerson, Hui-Wen; Farin, Federico M; Bammler, Theo K; Beyer, Richard P; Strand, Stuart E; Doty, Sharon L

    2010-08-01

    Trichloroethylene (TCE) is an important environmental contaminant of soil, groundwater, and air. Studies of the metabolism of TCE by poplar trees suggest that cytochrome P450 enzymes are involved. Using poplar genome microarrays, we report a number of putative genes that are differentially expressed in response to TCE. In a previous study, transgenic hybrid poplar plants expressing mammalian cytochrome P450 2E1 (CYP2E1) had increased metabolism of TCE. In the vector control plants for this construct, 24 h following TCE exposure, 517 genes were upregulated and 650 genes were downregulated over 2-fold when compared with the non-exposed vector control plants. However, in the transgenic CYP2E1 plant, line 78, 1,601 genes were upregulated and 1,705 genes were downregulated over 2-fold when compared with the non-exposed transgenic CYP2E1 plant. It appeared that the CYP2E1 transgenic hybrid poplar plants overexpressing mammalian CYP2E1 showed a larger number of differentially expressed transcripts, suggesting a metabolic pathway for TCE to metabolites had been initiated by activity of CYP2E1 on TCE. These results suggest that either the over-expression of the CYP2E1 gene or the abundance of TCE metabolites from CYP450 2E1 activity triggered a strong genetic response to TCE. Particularly, cytochrome p450s, glutathione S-transferases, glucosyltransferases, and ABC transporters in the CYP2E1 transgenic hybrid poplar plants were highly expressed compared with in vector controls.

  1. Expression of the neuronal adaptor protein X11alpha protects against memory dysfunction in a transgenic mouse model of Alzheimer's disease.

    LENUS (Irish Health Repository)

    Mitchell, Jacqueline C

    2010-01-01

    X11alpha is a neuronal-specific adaptor protein that binds to the amyloid-beta protein precursor (AbetaPP). Overexpression of X11alpha reduces Abeta production but whether X11alpha also protects against Abeta-related memory dysfunction is not known. To test this possibility, we crossed X11alpha transgenic mice with AbetaPP-Tg2576 mice. AbetaPP-Tg2576 mice produce high levels of brain Abeta and develop age-related defects in memory function that correlate with increasing Abeta load. Overexpression of X11alpha alone had no detectable adverse effect upon behavior. However, X11alpha reduced brain Abeta levels and corrected spatial reference memory defects in aged X11alpha\\/AbetaPP double transgenics. Thus, X11alpha may be a therapeutic target for Alzheimer\\'s disease.

  2. Evaluation of antigens stability of tobacco seeds as edible vaccine against VTEC strains

    Directory of Open Access Journals (Sweden)

    Luciana Rossi

    2015-11-01

    Full Text Available Plants have represent a promising alternative for biopharmaceutical proteins (Ma et al., 2003; Rossi et al., 2014. Many plant based edible vaccines have been shown to be effective in inducing local immune responses (Rossi et al., 2013. Edible vaccines can activate both mucosal and systemic immunity, as they come in contact with the digestive tract lining. This dual effect would provide first-line defense against pathogens invading through the mucosa. The antigens are released in the intestines are taken up by M cells that are present over the Payer’s patches (in the ileum and the gut associated lymphoid tissue (GALT. Edible vaccines represent an important worldwide goal for the prevention of the enteric diseases, also in livestock. In particular, the enteric infections are a significant clinical problem in pigs. Verocytotoxic Escherichia (E. coli strains are responsible for serious enterotoxaemia that causes important economic losses in the pig industry. The production of a vaccine for oral administration of transgenic seeds could be a practical and efficient system to prevent the infection and to reduce the antibiotic use. This study was focused on tobacco plants, previously transformed by agroinfection for the seed-specific expression of antigenic proteins (F18 adhesive fimbriae and the B subunit of the Vt2e toxin as model of edible vaccines against verocytotoxic E. coli strains. The dietary administration of transgenic tobacco seeds promotes a significant increase in the number of mucosal IgA-producing cells of the tunica propria in both small and large intestine in mice (Rossi et al., 2013. A protective effect of oral administration of transgenic tobacco seeds was also observed against verocytotoxic Escherichia coli infection in piglets (Rossi et al., 2014. The aim of this study was to assess the seed-expression stability, that is a important requirement in the vaccine production, of F 18 and Vt2e-B heterologous genes into the progeny of

  3. Production of Active Bacillus licheniformis Alpha-Amylase in Tobacco and its Application in Starch Liquefaction

    NARCIS (Netherlands)

    Pen, J; MOLENDIJK, L; Quax, Wim J.; SIJMONS, PC; VANOOYEN, AJJ; VANDENELZEN, PJM; RIETVELD, K; HOEKEMA, A

    As a first example of the feasibility of producing industrial bulk enzymes in plants, we have expressed Bacillus licheniformis alpha-amylase in transgenic tobacco, and applied the seeds directly in starch liquification. The enzyme was properly secreted into the intercellular space, and maximum

  4. A primary study of high performance transgenic rice through maize UBI-1 promoter fusing selective maker gene

    International Nuclear Information System (INIS)

    Shen, J.; Cai, P.; Qing, F.

    2012-01-01

    Based on the expression vector pBI121, we successfully constructed a plant over-expression vector of Hspa4 gene fusing with selective maker gene (hygromycin-resistance gene) driven by the Ubi-1 promoter (pBI121-Ubi-Hpt-Hspa4, p121UHH). The plant expression vectors p121UHH and pCAMBIA1301-Ubi-Hspa4 (p1301UH) were transformed into the rice callus, mediated by Agrobacterium tumefaciens. We screened 17 p121UHH-positive transgenic plants and 15 p1301UH-positive transgenic plants by the hygromycin-resistance gene. The pick-up rate of the resistance callus was 51.7% and 42.5%, respectively, and the rate of regeneration for the resistance callus was 51.2% and 49.1%, respectively. The result of polymerase chain reaction (P CR) identification indicated that the pick-up rate of positive transgenic plants was 51.7% and 42.5% and the total transformation efficiency was 16.5% and 6.2%, and the former was 2.66 time s of the later. The results of the experiment indicate that the possibility of the appearance of false positive results in the fusing of a plant over-expression vector with a selective maker gene is much less. (author)

  5. PP005. Vitamin D depletion aggravates hypertension in transgenic rats

    DEFF Research Database (Denmark)

    Bjørkholt Andersen, Louise; Herse, Florian; Christesen, Henrik Thybo

    2013-01-01

    INTRODUCTION: Vitamin D may ameliorate hypertension and kidney disease through genomic and extra-genomic pathways. OBJECTIVE: To investigate the impact of vitamin D in a transgenic rat model of angiotensin II-mediated hypertensive organ failure. METHODS: In 4-week-old age-matched rats overexpress......INTRODUCTION: Vitamin D may ameliorate hypertension and kidney disease through genomic and extra-genomic pathways. OBJECTIVE: To investigate the impact of vitamin D in a transgenic rat model of angiotensin II-mediated hypertensive organ failure. METHODS: In 4-week-old age-matched rats...... determined once weekly. After three weeks, animals were sacrificed. Heart tissue was examined for atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) by RT-PCR. RESULTS: The vitamin D depleted group had higher blood pressure at week 1 (mean difference 23.4mmHg, 95% CI 9.1-37.7) and tended...

  6. Novel Strategy to Control Transgene Expression Mediated by a Sendai Virus-Based Vector Using a Nonstructural C Protein and Endogenous MicroRNAs.

    Directory of Open Access Journals (Sweden)

    Masayuki Sano

    Full Text Available Tissue-specific control of gene expression is an invaluable tool for studying various biological processes and medical applications. Efficient regulatory systems have been utilized to control transgene expression in various types of DNA viral or integrating viral vectors. However, existing regulatory systems are difficult to transfer into negative-strand RNA virus vector platforms because of significant differences in their transcriptional machineries. In this study, we developed a novel strategy for regulating transgene expression mediated by a cytoplasmic RNA vector based on a replication-defective and persistent Sendai virus (SeVdp. Because of the capacity of Sendai virus (SeV nonstructural C proteins to specifically inhibit viral RNA synthesis, overexpression of C protein significantly reduced transgene expression mediated by SeVdp vectors. We found that SeV C overexpression concomitantly reduced SeVdp mRNA levels and genomic RNA synthesis. To control C expression, target sequences for an endogenous microRNA were incorporated into the 3' untranslated region of the C genes. Incorporation of target sequences for miR-21 into the SeVdp vector restored transgene expression in HeLa cells by decreasing C expression. Furthermore, the SeVdp vector containing target sequences for let-7a enabled cell-specific control of transgene expression in human fibroblasts and induced pluripotent stem cells. Our findings demonstrate that SeV C can be used as an effective regulator for controlling transgene expression. This strategy will contribute to efficient and less toxic SeVdp-mediated gene transfer in various biological applications.

  7. Overexpression of CaTLP1, a putative transcription factor in chickpea (Cicer arietinum L.), promotes stress tolerance.

    Science.gov (United States)

    Wardhan, Vijay; Jahan, Kishwer; Gupta, Sonika; Chennareddy, Srinivasarao; Datta, Asis; Chakraborty, Subhra; Chakraborty, Niranjan

    2012-07-01

    Dehydration is the most crucial environmental constraint on plant growth and development, and agricultural productivity. To understand the underlying mechanism of stress tolerance, and to identify proteins for improving such important trait, we screened the dehydration-responsive proteome of chickpea and identified a tubby-like protein, referred to as CaTLP1. The CaTLP1 was found to predominantly bind to double-stranded DNA but incapable of transcriptional activation. We investigated the gene structure and organization and demonstrated, for the first time, that CaTLP1 may be involved in osmotic stress response in plants. The transcripts are strongly expressed in vegetative tissues but weakly in reproductive tissues. CaTLP1 is upregulated by dehydration and high salinity, and by treatment with abscisic acid (ABA), suggesting that its stress-responsive function might be associated with ABA-dependent network. Overexpression of CaTLP1 in transgenic tobacco plants conferred dehydration, salinity and oxidative stress tolerance along with improved shoot and root architecture. Molecular genetic analysis showed differential expression of CaTLP1 under normal and stress condition, and its preferential expression in the nucleus might be associated with enhanced stress tolerance. Our work suggests important roles of CaTLP1 in stress response as well as in the regulation of plant development.

  8. Transgenic plants for enhanced biodegradation and phytoremediation of organic xenobiotics.

    Science.gov (United States)

    Abhilash, P C; Jamil, Sarah; Singh, Nandita

    2009-01-01

    Phytoremediation--the use of plants to clean up polluted soil and water resources--has received much attention in the last few years. Although plants have the inherent ability to detoxify xenobiotics, they generally lack the catabolic pathway for the complete degradation of these compounds compared to microorganisms. There are also concerns over the potential for the introduction of contaminants into the food chain. The question of how to dispose of plants that accumulate xenobiotics is also a serious concern. Hence the feasibility of phytoremediation as an approach to remediate environmental contamination is still somewhat in question. For these reasons, researchers have endeavored to engineer plants with genes that can bestow superior degradation abilities. A direct method for enhancing the efficacy of phytoremediation is to overexpress in plants the genes involved in metabolism, uptake, or transport of specific pollutants. Furthermore, the expression of suitable genes in root system enhances the rhizodegradation of highly recalcitrant compounds like PAHs, PCBs etc. Hence, the idea to amplify plant biodegradation of xenobiotics by genetic manipulation was developed, following a strategy similar to that used to develop transgenic crops. Genes from human, microbes, plants, and animals are being used successfully for this venture. The introduction of these genes can be readily achieved for many plant species using Agrobacterium tumefaciens-mediated plant transformation or direct DNA methods of gene transfer. One of the promising developments in transgenic technology is the insertion of multiple genes (for phase 1 metabolism (cytochrome P450s) and phase 2 metabolism (GSH, GT etc.) for the complete degradation of the xenobiotics within the plant system. In addition to the use of transgenic plants overexpressed with P450 and GST genes, various transgenic plants expressing bacterial genes can be used for the enhanced degradation and remediation of herbicides, explosives

  9. The use of phosphomannose isomerase selection system for Agrobacterium-mediated transformation of tobacco and flax aimed for phytoremediation.

    Science.gov (United States)

    Hilgert, Jitka; Sura-De Jong, Martina; Fišer, Jiří; Tupá, Kateřina; Vrbová, Miroslava; Griga, Miroslav; Macek, Tomáš; Žiarovská, Jana

    2017-05-04

    A plant selection system based on the phosphomannose isomerase gene (pmi) as a selectable marker is often used to avoid selection using antibiotic resistance. Nevertheless, pmi gene is endogenous in several plant species and therefore difficult to use in such cases. Here we evaluated and compared Agrobacterium-mediated transformation of Linum usitatissimum breeding line AGT-952 (without endogenous pmi gene) and Nicotiana tabacum var. WSC-38 (with endogenous pmi gene). Transformation was evaluated for vectors bearing transgenes that have the potential to be involved in improved phytoremediation of contaminated environment. Tobacco regenerants selection resulted in 6.8% transformation efficiency when using a medium supplemented with 30 g/L mannose with stepwise decrease of the sucrose concentration. Similar transformation efficiency (5.3%) was achieved in transformation of flax. Relatively low selection efficiency was achieved (12.5% and 34.8%, respectively). The final detection of efficient pmi selection was conducted using PCR and the non-endogenous genes; pmi transgene for flax and todC2 transgene for tobacco plants.

  10. Truncated presequences of mitochondrial F1-ATPase beta subunit from Nicotiana plumbaginifolia transport CAT and GUS proteins into mitochondria of transgenic tobacco.

    Science.gov (United States)

    Chaumont, F; Silva Filho, M de C; Thomas, D; Leterme, S; Boutry, M

    1994-02-01

    The mitochondrial F1-ATPase beta subunit (ATPase-beta) of Nicotiana plumbaginifolia is nucleus-encoded as a precursor containing an NH2-terminal extension. By sequencing the mature N. tabacum ATPase-beta, we determined the length of the presequence, viz. 54 residues. To define the essential regions of this presequence, we produced a series of 3' deletions in the sequence coding for the 90 NH2-terminal residues of ATPase-beta. The truncated sequences were fused with the chloramphenicol acetyl transferase (cat) and beta-glucuronidase (gus) genes and introduced into tobacco plants. From the observed distribution of CAT and GUS activity in the plant cells, we conclude that the first 23 amino-acid residues of ATPase-beta remain capable of specifically targeting reporter proteins into mitochondria. Immunodetection in transgenic plants and in vitro import experiments with various CAT fusion proteins show that the precursors are processed at the expected cleavage site but also at a cryptic site located in the linker region between the presequence and the first methionine of native CAT.

  11. Functional components of the bacterial CzcCBA efflux system reduce cadmium uptake and accumulation in transgenic tobacco plants.

    Science.gov (United States)

    Nesler, Andrea; DalCorso, Giovanni; Fasani, Elisa; Manara, Anna; Di Sansebastiano, Gian Pietro; Argese, Emanuele; Furini, Antonella

    2017-03-25

    Cadmium (Cd) is a toxic trace element released into the environment by industrial and agricultural practices, threatening the health of plants and contaminating the food/feed chain. Biotechnology can be used to develop plant varieties with a higher capacity for Cd accumulation (for use in phytoremediation programs) or a lower capacity for Cd accumulation (to reduce Cd levels in food and feed). Here we generated transgenic tobacco plants expressing components of the Pseudomonas putida CzcCBA efflux system. Plants were transformed with combinations of the CzcC, CzcB and CzcA genes, and the impact on Cd mobilization was analysed. Plants expressing PpCzcC showed no differences in Cd accumulation, whereas those expressing PpCzcB or PpCzcA accumulated less Cd in the shoots, but more Cd in the roots. Plants expressing both PpCzcB and PpCzcA accumulated less Cd in the shoots and roots compared to controls, whereas plants expressing all three genes showed a significant reduction in Cd levels only in shoots. These results show that components of the CzcCBA system can be expressed in plants and may be useful for developing plants with a reduced capacity to accumulate Cd in the shoots, potentially reducing the toxicity of food/feed crops cultivated in Cd-contaminated soils. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Gene dosage induction of silencing directed against an Arabidopsis Myb transgene in tobacco

    Science.gov (United States)

    An unexpected reduction in petal pigmentation on petunia plants genetically engineered for enhanced flower color was one of the first experimental demonstrations of the natural process of RNA-associated gene silencing. The obvious visual nature of such alterations to pigment patterns of transgenic ...

  13. EMMPRIN overexpression in SVZ neural progenitor cells increases their migration towards ischemic cortex.

    Science.gov (United States)

    Kanemitsu, Michiko; Tsupykov, Oleg; Potter, Gaël; Boitard, Michael; Salmon, Patrick; Zgraggen, Eloisa; Gascon, Eduardo; Skibo, Galina; Dayer, Alexandre G; Kiss, Jozsef Z

    2017-11-01

    Stimulation of endogenous neurogenesis and recruitment of neural progenitors from the subventricular zone (SVZ) neurogenic site may represent a useful strategy to improve regeneration in the ischemic cortex. Here, we tested whether transgenic overexpression of extracellular matrix metalloproteinase inducer (EMMPRIN), the regulator of matrix metalloproteinases (MMPs) expression, in endogenous neural progenitor cells (NPCs) in the subventricular zone (SVZ) could increase migration towards ischemic injury. For this purpose, we applied a lentivector-mediated gene transfer system. We found that EMMPRIN-transduced progenitors exhibited enhanced MMP-2 activity in vitro and showed improved motility in 3D collagen gel as well as in cortical slices. Using a rat model of neonatal ischemia, we showed that EMMPRIN overexpressing SVZ cells invade the injured cortical tissue more efficiently than controls. Our results suggest that EMMPRIN overexpression could be suitable approach to improve capacities of endogenous or transplanted progenitors to invade the injured cortex. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Monitoring changes in anthocyanin and steroid alkaloid glycoside content in lines of transgenic potato plants using liquid chromatography/mass spectrometry.

    Science.gov (United States)

    Stobiecki, Maciej; Matysiak-Kata, Iwona; Frański, Rafał; Skała, Jacek; Szopa, Jan

    2003-03-01

    Transgenic potato plants overexpressing and repressing enzymes of flavonoids biosynthesis were created and analyzed. The selected plants clearly showed the expected changes in anthocyanins synthesis level. Overexpression of a DNA encoding dihydroflavonol 4-reductase (DFR) in sense orientation resulted in an increase in tuber anthocyanins, a 4-fold increase in petunidin and pelargonidin derivatives. A significant decrease in anthocyanin level was observed when the plant was transformed with a corresponding antisense construct. The transformation of potato plants was also accompanied by significant changes in steroid alkaloid glycosides (SAG) level in transgenic potato tuber. The changes in SAGs content was not dependent on flavonoid composition in transgenic potato. However, in an extreme situation where the highest (DFR11) or the lowest (DFRa3) anthocyanin level was detected the positive correlation with steroid alkaloid content was clearly visible. It is suggested that the changes in SAGs content resulted from chromatin stressed upon transformation. A liquid chromatography/mass spectrometry (LC/MS) system with electrospray ionization was applied for profiling qualitative and quantitative changes of steroid alkaloid glycosides in tubers of twelve lines of transgenic potato plants. Except alpha-chaconine and alpha-solanine, in the extracts from dried tuber skin alpha-solamargine and alpha-solasonine, triglycosides of solasonine, were identified in minor amounts, triglycosides of solanidine dehydrodimers were also recognized.

  15. Transcriptome Profiling Reveals the Negative Regulation of Multiple Plant Hormone Signaling Pathways Elicited by Overexpression of C-Repeat Binding Factors

    Directory of Open Access Journals (Sweden)

    Aixin Li

    2017-09-01

    Full Text Available C-repeat binding factors (CBF are a subfamily of AP2 transcription factors that play critical roles in the regulation of plant cold tolerance and growth in low temperature. In the present work, we sought to perform a detailed investigation into global transcriptional regulation of plant hormone signaling associated genes in transgenic plants engineered with CBF genes. RNA samples from Arabidopsis thaliana plants overexpressing two CBF genes, CBF2 and CBF3, were subjected to Illumina HiSeq 2000 RNA sequencing (RNA-Seq. Our results showed that more than half of the hormone associated genes that were differentially expressed in CBF2 or CBF3 transgenic plants were related to auxin signal transduction and metabolism. Most of these alterations in gene expression could lead to repression of auxin signaling. Accordingly, the IAA content was significantly decreased in young tissues of plants overexpressing CBF2 and CBF3 compared with wild type. In addition, genes associated with the biosynthesis of Jasmonate (JA and Salicylic acid (SA, as well as the signal sensing of Brassinolide (BR and SA, were down-regulated, while genes associated with Gibberellin (GA deactivation were up-regulated. In general, overexpression of CBF2 and CBF3 negatively affects multiple plant hormone signaling pathways in Arabidopsis. The transcriptome analysis using CBF2 and CBF3 transgenic plants provides novel and integrated insights into the interaction between CBFs and plant hormones, particularly the modulation of auxin signaling, which may contribute to the improvement of crop yields under abiotic stress via molecular engineering using CBF genes.

  16. Highly phosphorylated functionalized rice starch produced by transgenic rice expressing the potato GWD1 gene

    DEFF Research Database (Denmark)

    Chen, Yaling; Sun, Xiao; Zhou, Xin Mao

    2017-01-01

    Starch phosphorylation occurs naturally during starch metabolism in the plant and is catalysed by glucan water dikinases (GWD1) and phosphoglucan water dikinase/glucan water dikinase 3 (PWD/GWD3). We generated six stable individual transgenic lines by over-expressing the potato GWD1 in rice....... Transgenic rice grain starch had 9-fold higher 6-phospho (6-P) monoesters and double amounts of 3-phospho (3-P) monoesters, respectively, compared to control grain. The shape and topography of the transgenic starch granules were moderately altered including surface pores and less well defined edges....... The gelatinization temperatures of both rice flour and extracted starch were significantly lower than those of the control and hence negatively correlated with the starch phosphate content. The 6-P content was positively correlated with amylose content and relatively long amylopectin chains with DP25-36, and the 3-P...

  17. In planta Transformed Cumin (Cuminum cyminum L.) Plants, Overexpressing the SbNHX1 Gene Showed Enhanced Salt Endurance.

    Science.gov (United States)

    Pandey, Sonika; Patel, Manish Kumar; Mishra, Avinash; Jha, Bhavanath

    2016-01-01

    Cumin is an annual, herbaceous, medicinal, aromatic, spice glycophyte that contains diverse applications as a food and flavoring additive, and therapeutic agents. An efficient, less time consuming, Agrobacterium-mediated, a tissue culture-independent in planta genetic transformation method was established for the first time using cumin seeds. The SbNHX1 gene, cloned from an extreme halophyte Salicornia brachiata was transformed in cumin using optimized in planta transformation method. The SbNHX1 gene encodes a vacuolar Na+/H+ antiporter and is involved in the compartmentalization of excess Na+ ions into the vacuole and maintenance of ion homeostasis Transgenic cumin plants were confirmed by PCR using gene (SbNHX1, uidA and hptII) specific primers. The single gene integration event and overexpression of the gene were confirmed by Southern hybridization and competitive RT-PCR, respectively. Transgenic lines L3 and L13 showed high expression of the SbNHX1 gene compared to L6 whereas moderate expression was detected in L5 and L10 transgenic lines. Transgenic lines (L3, L5, L10 and L13), overexpressing the SbNHX1 gene, showed higher photosynthetic pigments (chlorophyll a, b and carotenoid), and lower electrolytic leakage, lipid peroxidation (MDA content) and proline content as compared to wild type plants under salinity stress. Though transgenic lines were also affected by salinity stress but performed better compared to WT plants. The ectopic expression of the SbNHX1 gene confirmed enhanced salinity stress tolerance in cumin as compared to wild type plants under stress condition. The present study is the first report of engineering salt tolerance in cumin, so far and the plant may be utilized for the cultivation in saline areas.

  18. In planta Transformed Cumin (Cuminum cyminum L. Plants, Overexpressing the SbNHX1 Gene Showed Enhanced Salt Endurance.

    Directory of Open Access Journals (Sweden)

    Sonika Pandey

    Full Text Available Cumin is an annual, herbaceous, medicinal, aromatic, spice glycophyte that contains diverse applications as a food and flavoring additive, and therapeutic agents. An efficient, less time consuming, Agrobacterium-mediated, a tissue culture-independent in planta genetic transformation method was established for the first time using cumin seeds. The SbNHX1 gene, cloned from an extreme halophyte Salicornia brachiata was transformed in cumin using optimized in planta transformation method. The SbNHX1 gene encodes a vacuolar Na+/H+ antiporter and is involved in the compartmentalization of excess Na+ ions into the vacuole and maintenance of ion homeostasis Transgenic cumin plants were confirmed by PCR using gene (SbNHX1, uidA and hptII specific primers. The single gene integration event and overexpression of the gene were confirmed by Southern hybridization and competitive RT-PCR, respectively. Transgenic lines L3 and L13 showed high expression of the SbNHX1 gene compared to L6 whereas moderate expression was detected in L5 and L10 transgenic lines. Transgenic lines (L3, L5, L10 and L13, overexpressing the SbNHX1 gene, showed higher photosynthetic pigments (chlorophyll a, b and carotenoid, and lower electrolytic leakage, lipid peroxidation (MDA content and proline content as compared to wild type plants under salinity stress. Though transgenic lines were also affected by salinity stress but performed better compared to WT plants. The ectopic expression of the SbNHX1 gene confirmed enhanced salinity stress tolerance in cumin as compared to wild type plants under stress condition. The present study is the first report of engineering salt tolerance in cumin, so far and the plant may be utilized for the cultivation in saline areas.

  19. ß-cell specific overexpression of suppressor of cytokine signalling-3 does not protect against multiple low dose streptozotocin induced type 1 diabetes in mice

    DEFF Research Database (Denmark)

    Börjesson, A; Rønn, S G; Karlsen, A E

    2011-01-01

    We investigated the impact of ß-cell specific overexpression of suppressor of cytokine signalling-3 (SOCS-3) on the development of multiple low dose streptozotocin (MLDSTZ) induced Type 1 diabetes and the possible mechanisms involved. MLDSTZ treatment was administered to RIP-SOCS-3 transgenic......RNA in islet cells and secretion of IL-1Ra into culture medium. MLDSTZ treatment caused gradual hyperglycemia both in the wt mice and in the transgenic mice with the latter tending to be more sensitive. In vitro experiments on wt and transgenic islets did not reveal any differences in sensitivity to damaging...

  20. Overexpression of EsMcsu1 from the halophytic plant Eutrema salsugineum promotes abscisic acid biosynthesis and increases drought resistance in alfalfa (Medicago sativa L.).

    Science.gov (United States)

    Zhou, C; Ma, Z Y; Zhu, L; Guo, J S; Zhu, J; Wang, J F

    2015-12-17

    The stress phytohormone abscisic acid (ABA) plays pivotal roles in plants' adaptive responses to adverse environments. Molybdenum cofactor sulfurases influence aldehyde oxidase activity and ABA biosynthesis. In this study, we isolated a novel EsMcsu1 gene encoding a molybdenum cofactor sulfurase from Eutrema salsugineum. EsMcus1 transcriptional patterns varied between organs, and its expression was significantly upregulated by abiotic stress or ABA treatment. Alfalfa plants that overexpressed EsMcsu1 had a higher ABA content than wild-type (WT) plants under drought stress conditions. Furthermore, levels of reactive oxygen species (ROS), ion leakage, and malondialdehyde were lower in the transgenic plants than in the WT plants after drought treatment, suggesting that the transgenic plants experienced less ROS-mediated damage. However, the expression of several stress-responsive genes, antioxidant enzyme activity, and osmolyte (proline and total soluble sugar) levels in the transgenic plants were higher than those in the WT plants after drought treatment. Therefore, EsMcsu1 overexpression improved drought tolerance in alfalfa plants by activating a series of ABA-mediated stress responses.