WorldWideScience

Sample records for acid response mutants

  1. Mutant fatty acid desaturase and methods for directed mutagenesis

    Science.gov (United States)

    Shanklin, John [Shoreham, NY; Whittle, Edward J [Greenport, NY

    2008-01-29

    The present invention relates to methods for producing fatty acid desaturase mutants having a substantially increased activity towards substrates with fewer than 18 carbon atom chains relative to an unmutagenized precursor desaturase having an 18 carbon chain length specificity, the sequences encoding the desaturases and to the desaturases that are produced by the methods. The present invention further relates to a method for altering a function of a protein, including a fatty acid desaturase, through directed mutagenesis involving identifying candidate amino acid residues, producing a library of mutants of the protein by simultaneously randomizing all amino acid candidates, and selecting for mutants which exhibit the desired alteration of function. Candidate amino acids are identified by a combination of methods. Enzymatic, binding, structural and other functions of proteins can be altered by the method.

  2. Induced mutants of Streptococcus lactis by gamma irradiation and its effect on milk acidity

    International Nuclear Information System (INIS)

    Daowd, A.H.; Sabbour, M.M.; Newigy, N.A.; Wahab, G.A.M.

    1988-01-01

    Streptococus lactis was exposed to different doses of gamma-irradiation (10, 50, 100 and 150 Kr). The results of acidity production in sterilized milk inoculated by isolates from radiation treatments and control could be summarized in the following: 1. The mean acidity produced by S. lactis isolates after irradiation at 10 Kr increased to be 0.66% than that of control isolates (0.62%). The acidity produced by the isolates of the 50 Kr treatment showed more increment to reach the peak (0.7%). Thereafter, acidity production decreased by isolates of the 100 Kr (0.53%) and 150 Kr (0.51%) treatments. Heme, the 50 Kr treatment could be considered activation dose to S. lactis starter for acid production. 2. Two mutants were selected. Acidity production by mutant I (from 10 Kr treatment) was 0.95%, and that of mutant II (from 50 Kr treatment) was 1.0%, while acid production by the parent S. lactis culture was 0.62%. Concerning the stability of the isolates, it was found that acid production by mutant I and mutant II slightly decreased by time. The mutants were re-irradiated after 37 and 60 days at doses 10, 25 and 50 Kr. Acid production in milk by isolates of the radiation treatments was determined. The following results were obtained: -The re-irradiation of the mutants decreased the ability of the isolates for acid production than that of parent mutants. -The re-irradiation of the mutants after 60 days yielded isolates which showed more reduction in acid produced than of isolates obtained from re-irridation of the mutants after 37-days. -The higher the dose of the re-irradiation of the mutants, the lower the mean of acid production by isolates of the treatment

  3. Combinations of mutant FAD2 and FAD3 genes to produce high oleic acid and low linolenic acid soybean oil.

    Science.gov (United States)

    Pham, Anh-Tung; Shannon, J Grover; Bilyeu, Kristin D

    2012-08-01

    High oleic acid soybeans were produced by combining mutant FAD2-1A and FAD2-1B genes. Despite having a high oleic acid content, the linolenic acid content of these soybeans was in the range of 4-6 %, which may be high enough to cause oxidative instability of the oil. Therefore, a study was conducted to incorporate one or two mutant FAD3 genes into the high oleic acid background to further reduce the linolenic acid content. As a result, soybean lines with high oleic acid and low linolenic acid (HOLL) content were produced using different sources of mutant FAD2-1A genes. While oleic acid content of these HOLL lines was stable across two testing environments, the reduction of linolenic acid content varied depending on the number of mutant FAD3 genes combined with mutant FAD2-1 genes, on the severity of mutation in the FAD2-1A gene, and on the testing environment. Combination of two mutant FAD2-1 genes and one mutant FAD3 gene resulted in less than 2 % linolenic acid content in Portageville, Missouri (MO) while four mutant genes were needed to achieve the same linolenic acid in Columbia, MO. This study generated non-transgenic soybeans with the highest oleic acid content and lowest linolenic acid content reported to date, offering a unique alternative to produce a fatty acid profile similar to olive oil.

  4. Analysis of nutritional quality of low phytic acid mutants in maize

    International Nuclear Information System (INIS)

    Yuan Ming'an; Luo Hongbing; Wang Zhonghua; Chen Jinhong; Mei Shufang; Shu Xiaoli; Wu Dianxing

    2008-01-01

    Major nutritional quality and components of eight low phytic acid (lpa) mutants and their corresponding wild types were studied. Compared to their corresponding wild types, the content of inorganic P (Pi) was all increased several times, while the content of total P (TP) in mutants was almost the same as their wild types. The contents of crude lipid and amylose were similar, but total starch was significantly different. Crude protein in some of mutants was increased significantly. Most of amino acids were increased, and essential amino acid-Lysine was increased except mutants derived from Q319 and X178. Mineral macronutrients and micronutrients were similar. All results showed that the lpa mutation in maize could enhance the nutritional quality and bioactivities. (authors)

  5. Homologous Recombination Defective Arabidopsis Mutants Exhibit Enhanced Sensitivity to Abscisic Acid.

    Directory of Open Access Journals (Sweden)

    Sujit Roy

    Full Text Available Abscisic acid (ABA acts as an important plant hormone in regulating various aspects of plant growth and developmental processes particularly under abiotic stress conditions. An increased ABA level in plant cells inhibits DNA replication and cell division, causing plant growth retardation. In this study, we have investigated the effects of ABA on the growth responses of some major loss-of-function mutants of DNA double-stand break (DSB repair genes in Arabidopsis during seed germination and early stages of seedling growth for understanding the role of ABA in the induction of genome instability in plants. A comparative analysis of ABA sensitivity of wild-type Arabidopsis and the knockout mutant lines related to DSB sensors, including atatm, atatr, the non-homologous end joining (NHEJ pathway genes, and mutants related to homologous recombination (HR pathway genes showed relatively enhanced sensitivity of atatr and HR-related mutants to ABA treatment. The expression levels of HR-related genes were increased in wild-type Arabidopsis (Col-0 during seed germination and early stages of seedling growth. Immunoblotting experiments detected phosphorylation of histone H2AX in wild-type (Col-0 and DSB repair gene mutants after ABA treatment, indicating the activation of DNA damage response due to ABA treatment. Analyses of DSB repair kinetics using comet assay under neutral condition have revealed comparatively slower DSB repair activity in HR mutants. Overall, our results have provided comprehensive information on the possible effect of ABA on DNA repair machinery in plants and also indicated potential functional involvement of HR pathway in repairing ABA induced DNA damage in Arabidopsis.

  6. Mutant E. coli strain with increased succinic acid production

    Science.gov (United States)

    Donnelly, Mark; Millard, Cynthia S.; Stols, Lucy

    2001-09-25

    A method for isolating succinic acid producing bacteria is provided comprising increasing the biomass of an organism which lacks the ability to catabolize pyruvate, and then subjecting the biomass to glucose-rich medium in an anaerobic environment to enable pyruvate-catabolizing mutants to grow. The invention also provides for a mutant that produces high amounts of succinic acid, which has been derived from a parent which lacked the genes for pyruvate formate lyase and lactate dehydrogenase, and which belongs to the E.coli Group of Bacteria.

  7. Mutant E. coli strain with increased succinic acid production

    Science.gov (United States)

    Donnelly, Mark; Millard, Cynthia S.; Stols, Lucy

    1998-01-01

    A method for isolating succinic acid producing bacteria is provided comprising increasing the biomass of an organism which lacks the ability to catabolize pyruvate, and then subjecting the biomass to glucose-rich medium in an anaerobic environment to enable pyruvate-catabolizing mutants to grow. The invention also provides for a mutant that produces high amounts of succinic acid, which as been derived from a parent which lacked the genes for pyruvate formate lyase and lactate dehydrogenase, and which belongs to the E.coli Group of Bacteria.

  8. Branched-chain fatty acid biosynthesis in a branched-chain amino acid aminotransferase mutant of Staphylococcus carnosus

    DEFF Research Database (Denmark)

    Beck, Hans Christian

    2005-01-01

    Fatty acid biosynthesis by a mutant strain of Staphylococcus carnosus deficient in branched-chain amino acid aminotransferase (IlvE) activity was analysed. This mutant was unable to produce the appropriate branched-chain alpha-ketoacid precursors for branched-chain fatty acid biosynthesis from...... in rich medium and growth in defined medium supplemented with 2-methylpropanoic acid lead to extensive alteration of the fatty acid composition in the cell membrane. In rich medium, a change from 51.7% to 17.1% anteiso-C15:0, and from 3.6% to 33.9% iso-C14:0 fatty acids as compared to the wild-type strain...... for 2-methylpropanoic acid production, revealing that the IlvE protein plays an important, but not essential role in the biosynthesis of branched-chain fatty acids and secondary metabolites in S. carnosus....

  9. Response of a semi-dwarf mutant of rice (Oryza sativa L.) to application of gibberelic acid

    International Nuclear Information System (INIS)

    Orozco, Rafael; Navarro, Willy

    2000-01-01

    The role of gibberellic acid (AG 3 ) on the capacity of elongation of the nodes of a semi-dwarf mutant of rice 2B-95, was studied. This mutant, with a height between 65 to 90 cm, was obtained by Co-60 gamma irradiation from a material called WS tall with an average height of 165 cm. Variety semi-dwarf CR-1113 was used as an additional control. The work was carried out under conditions in vitro applying the AG 3 in concentrations of 0.20, 30 and 40 ppm. The answer of the mutant and of the other two genotypes to exogenous application of the hormone was assessed by measuring the length of the second leaf sheath emerged after germination of the seed according to a special modification of the methodology of Harada and Vergara (1971). The results have indicated that WS genotypes and CR-1113, the length of the sheath of the second leaf at 11 days old, increased by all concentrations of AG 3 evaluated, while in the semi-dwarf mutant 2B-95 the effect was only significant (P 3 . (author) [es

  10. Mutant breeding of Aspergillus niger irradiated by 12C6+ for hyper citric acid

    International Nuclear Information System (INIS)

    Hu Wei; Li Wenjian; Chen Jihong; Liu Jing; Wang Shuyang; Wang Jufang; Lu Dong

    2014-01-01

    In this study, strains of Aspergillus niger No.4 for hyper citric acid were irradiated to different doses by 80 MeV/u 12 C 6+ ion beams. Seven mutant strains showed marked citric acid over-production records and faster productivity than initial Aspergillus niger No.4 by shaking flash fermentation. The maximum product yield was 132.8 gL -1 (the H4002 strain) being a 8.8% increase to the initial strain. The scale-up experiment was carried out in a 100 L bioreactor. The mutant H4002 can accumulate 187gL -1 product yield of citric acid from starch liquefying supernatant. The productivity of citric acid was 2.75 g L -1 h -1 . So, the mutant H4002 possesses rapid sugar katabolism for producing citric acid. Meanwhile, the pellet morphology kept compact and round during the whole submerged fermentation, which was suited to produce citric acid. The results indicate that mutant H4002 has potential ability to produce citric acid rapidly. (authors)

  11. High linolenic acid mutant in soybean induced by X-ray irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Takagi, Y. [Saga Univ. (Japan); Hossain, A. B.M.M.; Yanagita, T.; Kusaba, S.

    1989-12-15

    Soybean [Glycine max (L.) Merr. cv. Bay] seeds were irradiated with X-rays (25kR) and the M{sub 2} progeny was screened for changes in the fatty acid composition of seed oil. X-ray irradiation remarkably increased the variability of the fatty acid composition in the oil of the Bay cultivar. A mutant in which linolenic acid accounted for 18.4 per cent of the total oil cornpared with 9.4 per cent in the Bay cultivar was identified among 2006 M{sub 2} plants. The M{sub 3} generation of the mutant also showed a linolenic acid content approximately two times higher than that of the original variety.

  12. High linolenic acid mutant in soybean induced by X-ray irradiation

    International Nuclear Information System (INIS)

    Takagi, Y.; Hossain, A.B.M.M.; Yanagita, T.; Kusaba, S.

    1989-01-01

    Soybean [Glycine max (L.) Merr. cv. Bay] seeds were irradiated with X-rays (25kR) and the M 2 progeny was screened for changes in the fatty acid composition of seed oil. X-ray irradiation remarkably increased the variability of the fatty acid composition in the oil of the Bay cultivar. A mutant in which linolenic acid accounted for 18.4 per cent of the total oil cornpared with 9.4 per cent in the Bay cultivar was identified among 2006 M 2 plants. The M 3 generation of the mutant also showed a linolenic acid content approximately two times higher than that of the original variety

  13. Breeding L(+)-lactic acid high productive mutant from xylose by nitrogen ions

    International Nuclear Information System (INIS)

    Yang Yingge; Li Wen; Liu Dan; Fan Yonghong; Wang Dongmei; Zheng Zhiming; Yu Zengliang

    2007-01-01

    In order to obtain higher L(+)-lactic acid yield strain fermentating from xylose, the original strain Rhizopus oryzae RLC41-6 was mutated by 10keV N + ion implantation. A mutant strain RQ4012 was obtained. After 72h shake-flask cultivation, the concentration of L(+)-lactic acid reached 74.37g/L, and the productivity was 1.03g/(L.h). Its lactic acid yield was 160% higher than that of the original one, and the mutant strain has high genetic stability. (authors)

  14. Ultraviolet light-induced mutants of Streptococcus lactis subspecies diacetylactis with enhanced acid- or flavor-producing abilities

    International Nuclear Information System (INIS)

    Kuila, R.K.; Ranganathan, B.

    1978-01-01

    A strain of Streptococcus lactis subspecies diacetylactis S 1 isolated from fresh milk was exposed to 7200 ergs/mm 2 of ultraviolet radiation. Over 8100 colonies surviving from 7.4 x 10 6 cells exposed to radiation were screened on citrate agar for detection and isolation of mutants with increased flavor and/or acid production. Of the survivors, 960 were type-I mutants that exhibited clear zone on citrate agar after 18 h (presumed to be high diacetyl producers), and 288 were type-II mutants which did not exhibit clear zones on citrate agar for up to 72 h (high acid producers). Type-II mutants produced an average .93 percent titratable acidity which was 34 percent more than the .69 percent of the parent. Reduction in titratable acidity (56 percent less) was considerable in type-I mutants, compared with the parent culture. Diacetyl + acetoin production by type-I mutants was 137.9 ppM which has 4.5 times more than that of the parental strain. Acetaldehyde production in the mutants varied from 1.5 to 34.5 ppM (parent culture 3.0 ppM). The mutants with increased acid and high acetoin plus diacetyl production were stable after 50 subcultures in milk

  15. Assesment of spineless safflower (Carthamus tinctorius, L.) mutant lines for seed oil content and fatty acid profiles

    International Nuclear Information System (INIS)

    Ragab, A.I.; Kassem, M.; Moustafa, H.A.M.

    2008-01-01

    This study was conducted to assess the new spineless mutants that previously induced through gamma radiation and hybridization techniques in the advanced generation for seed oil content and fatty acid profiles The obtained results cleared that oil percentages of all seven safflower mutants were increased than local variety Giza (1) and the new mutant hybrid 2 line (white petals) had the highest increase value of oil percentage (10%) but the mutant line M14 (dark red petals) had the lowest increase value of oil percentage (3.1 %) The mutant line M7 (yellow petals) had the highest value of total saturated fatty acid (40.38%), because it had the highest value of palmitic fatty acid (25.16%), comparing to 10.01% value for local variety Giza (1), followed by mutant line hybrid 2 (white petals) which had (39.88%) because it had the highest value of caprylic, capric, lauric, myristic and stearic fatty acids. All safflower mutant lines had higher value of oleic fatty acid than that of the local variety Giza (1) the two new safflower mutant lines M7 (yellow petals) and hybrid 2 (white petal) had the highest value of oleic fatty acid 41.22% and 39.88% respectively in comparison with 13.5% for local variety Giza (1), the obtained results are indicating to seed oil content negative correlation between oleic and linoleic acids

  16. Differential disease resistance response in the barley necrotic mutant nec1

    Directory of Open Access Journals (Sweden)

    Kunga Laura

    2011-04-01

    Full Text Available Abstract Background Although ion fluxes are considered to be an integral part of signal transduction during responses to pathogens, only a few ion channels are known to participate in the plant response to infection. CNGC4 is a disease resistance-related cyclic nucleotide-gated ion channel. Arabidopsis thaliana CNGC4 mutants hlm1 and dnd2 display an impaired hypersensitive response (HR, retarded growth, a constitutively active salicylic acid (SA-mediated pathogenesis-related response and elevated resistance against bacterial pathogens. Barley CNGC4 shares 67% aa identity with AtCNGC4. The barley mutant nec1 comprising of a frame-shift mutation of CNGC4 displays a necrotic phenotype and constitutively over-expresses PR-1, yet it is not known what effect the nec1 mutation has on barley resistance against different types of pathogens. Results nec1 mutant accumulated high amount of SA and hydrogen peroxide compared to parental cv. Parkland. Experiments investigating nec1 disease resistance demonstrated positive effect of nec1 mutation on non-host resistance against Pseudomonas syringae pv. tomato (Pst at high inoculum density, whereas at normal Pst inoculum concentration nec1 resistance did not differ from wt. In contrast to augmented P. syringae resistance, penetration resistance against biotrophic fungus Blumeria graminis f. sp. hordei (Bgh, the causal agent of powdery mildew, was not altered in nec1. The nec1 mutant significantly over-expressed race non-specific Bgh resistance-related genes BI-1 and MLO. Induction of BI-1 and MLO suggested putative involvement of nec1 in race non-specific Bgh resistance, therefore the effect of nec1on mlo-5-mediated Bgh resistance was assessed. The nec1/mlo-5 double mutant was as resistant to Bgh as Nec1/mlo-5 plants, suggesting that nec1 did not impair mlo-5 race non-specific Bgh resistance. Conclusions Together, the results suggest that nec1 mutation alters activation of systemic acquired resistance

  17. Enhanced mucosal delivery of antigen with cell wall mutants of lactic acid bacteria.

    Science.gov (United States)

    Grangette, Corinne; Müller-Alouf, Heide; Hols, Pascal; Goudercourt, Denise; Delcour, Jean; Turneer, Mireille; Mercenier, Annick

    2004-05-01

    The potential of recombinant lactic acid bacteria (LAB) to deliver heterologous antigens to the immune system and to induce protective immunity has been best demonstrated by using the C subunit of tetanus toxin (TTFC) as a model antigen. Two types of LAB carriers have mainly been used, Lactobacillus plantarum and Lactococcus lactis, which differ substantially in their abilities to resist passage through the stomach and to persist in the mouse gastrointestinal tract. Here we analyzed the effect of a deficiency in alanine racemase, an enzyme that participates in cell wall synthesis, in each of these bacterial carriers. Recombinant wild-type and mutant strains of L. plantarum NCIMB8826 and L. lactis MG1363 producing TTFC intracellularly were constructed and used in mouse immunization experiments. Remarkably, we observed that the two cell wall mutant strains were far more immunogenic than their wild-type counterparts when the intragastric route was used. However, intestinal TTFC-specific immunoglobulin A was induced only after immunization with the recombinant L. plantarum mutant strain. Moreover, the alanine racemase mutant of either LAB strain allowed induction of a much stronger serum TTFC-specific immune response after immunization via the vagina, which is a quite different ecosystem than the gastrointestinal tract. The design and use of these mutants thus resulted in a major improvement in the mucosal delivery of antigens exhibiting vaccine properties.

  18. A proteome analysis of the response of a Pseudomonas aeruginosa oxyR mutant to iron limitation.

    Science.gov (United States)

    Vinckx, Tiffany; Wei, Qing; Matthijs, Sandra; Noben, Jean-Paul; Daniels, Ruth; Cornelis, Pierre

    2011-06-01

    In Pseudomonas aeruginosa the response to oxidative stress is orchestrated by the LysR regulator OxyR by activation of the transcription of two catalase genes (katA and katB), of the alkyl-hydroxyperoxidases ahpCF and ahpB. Next to the expected high sensitivity to oxidative stress generated by reactive oxygen species (ROS: H(2)O(2), O(2)(-)), the oxyR mutant shows a defective growth under conditions of iron limitation (Vinckx et al. 2008). Although production and uptake of the siderophore pyoverdine is not affected by the absence of oxyR, the mutant is unable to satisfy its need for iron when grown under iron limiting conditions. In order to get a better insight into the effects caused by iron limitation on the physiological response of the oxyR mutant we decided to compare the proteomes of the wild type and the mutant grown in the iron-poor casamino acids medium (CAA), in CAA plus H(2)O(2), and in CAA plus the strong iron chelator ethylenediamine-N,N'-bis(2-hydroxyphenylacetic acid) (EDDHA). Especially in the presence of hydrogen peroxide the oxyR cells increase the production of stress proteins (Dps and IbpA). The superoxide dismutase SodM is produced in higher amounts in the oxyR mutant grown in CAA plus H(2)O(2). The PchB protein, a isochorismate-pyruvate lyase involved in the siderophore pyochelin biosynthesis is not detectable in the extracts from the oxyR mutant grown in the presence of hydrogen peroxide. When cells were grown in the presence of EDDHA, we observed a reduction of the ferric uptake regulator (Fur), and an increase in the two subunits of the succinyl-CoA synthetase and the fumarase FumC1.

  19. Malic acid production by chemically induced Aspergillus niger MTCC 281 mutant from crude glycerol.

    Science.gov (United States)

    Iyyappan, J; Bharathiraja, B; Baskar, G; Jayamuthunagai, J; Barathkumar, S; Anna Shiny, R

    2018-03-01

    In the present investigation, crude glycerol derived from transesterification process was utilized to produce the commercially-valuable malic acid. A combined resistant on methanol and malic acid strain of Aspergillus niger MTCC 281 mutant was generated in solid medium containing methanol (1-5%) and malic acid (40-80 g/L) by the adaptation process for 22 weeks. The ability of induced Aspergillus niger MTCC 281 mutant to utilize crude glycerol and pure glycerol to produce malic acid was studied. The yield of malic acid was increased with 4.45 folds compared with that of parent strain from crude glycerol. The highest concentration of malic acid from crude glycerol by using beneficial mutant was found to be 77.38 ± 0.51 g/L after 192 h at 25 °C. This present study specified that crude glycerol by-product from biodiesel production could be used for producing high amount of malic acid without any pretreatment. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Phenotypic, genetic and molecular characterization of a maize low phytic acid mutant (lpa241)

    DEFF Research Database (Denmark)

    Pilu, R.; Panzeri, D.; Gavazzi, G.

    2003-01-01

    -nutritional factor for animals, and isolation of maize low phytic acid (lpa) mutants provides a novel approach to study its biochemical pathway and to tackle the nutritional problems associated with it. Following chemical mutagenesis of pollen, we have isolated a viable recessive mutant named lpa 241 showing about...... 90% reduction of phytic acid and about a tenfold increase in seed-free phosphate content. Although germination rate was decreased by about 30% compared to wild-type, developement of mutant plants was apparentely unaffected. The results of the genetic, biochemical and molecular characterization...

  1. GID1 modulates stomatal response and submergence tolerance involving abscisic acid and gibberellic acid signaling in rice.

    Science.gov (United States)

    Du, Hao; Chang, Yu; Huang, Fei; Xiong, Lizhong

    2015-11-01

    Plant responses to abiotic stresses are coordinated by arrays of growth and developmental programs. Gibberellic acid (GA) and abscisic acid (ABA) play critical roles in the developmental programs and environmental responses, respectively, through complex signaling and metabolism networks. However, crosstalk between the two phytohormones in stress responses remains largely unknown. In this study, we report that GIBBERELLIN-INSENSITIVE DWARF 1 (GID1), a soluble receptor for GA, regulates stomatal development and patterning in rice (Oryza sativa L.). The gid1 mutant showed impaired biosynthesis of endogenous ABA under drought stress conditions, but it exhibited enhanced sensitivity to exogenous ABA. Scanning electron microscope and infrared thermal image analysis indicated an increase in the stomatal conductance in the gid1 mutant under drought conditions. Interestingly, the gid1 mutant had increased levels of chlorophyll and carbohydrates under submergence conditions, and showed enhanced reactive oxygen species (ROS)-scavenging ability and submergence tolerance compared with the wild-type. Further analyses suggested that the function of GID1 in submergence responses is partially dependent on ABA, and GA signaling by GID1 is involved in submergence tolerance by modulating carbohydrate consumption. Taken together, these findings suggest GID1 plays distinct roles in stomatal response and submergence tolerance through both the ABA and GA signaling pathways in rice. © 2014 Institute of Botany, Chinese Academy of Sciences.

  2. Arabidopsis MAP kinase 4 regulates salicylic acid- and jasmonic acid/ethylene-dependent responses via EDS1 and PAD4

    DEFF Research Database (Denmark)

    Brodersen, P; Petersen, M; Nielsen, Henrik Bjørn

    2006-01-01

    Arabidopsis MPK4 has been implicated in plant defense regulation because mpk4 knockout plants exhibit constitutive activation of salicylic acid (SA)-dependent defenses, but fail to induce jasmonic acid (JA) defense marker genes in response to JA. We show here that mpk4 mutants are also defective...

  3. Chir99021 and Valproic acid reduce the proliferative advantage of Apc mutant cells.

    Science.gov (United States)

    Langlands, Alistair J; Carroll, Thomas D; Chen, Yu; Näthke, Inke

    2018-02-15

    More than 90% of colorectal cancers carry mutations in Apc that drive tumourigenesis. A 'just-right' signalling model proposes that Apc mutations stimulate optimal, but not excessive Wnt signalling, resulting in a growth advantage of Apc mutant over wild-type cells. Reversal of this growth advantage constitutes a potential therapeutic approach. We utilised intestinal organoids to compare the growth of Apc mutant and wild-type cells. Organoids derived from Apc Min/+ mice recapitulate stages of intestinal polyposis in culture. They eventually form spherical cysts that reflect the competitive growth advantage of cells that have undergone loss of heterozygosity (LOH). We discovered that this emergence of cysts was inhibited by Chiron99021 and Valproic acid, which potentiates Wnt signalling. Chiron99021 and Valproic acid restrict the growth advantage of Apc mutant cells while stimulating that of wild-type cells, suggesting that excessive Wnt signalling reduces the relative fitness of Apc mutant cells. As a proof of concept, we demonstrated that Chiron99021-treated Apc mutant organoids were rendered susceptible to TSA-induced apoptosis, while wild-type cells were protected.

  4. Biooxidation of fatty acid distillates to dibasic acids by a mutant of Candida tropicalis.

    Science.gov (United States)

    Gangopadhyay, Sarbani; Nandi, Sumit; Ghosh, Santinath

    2006-01-01

    Fatty acid distillates (FADs) produced during physical refining of vegetable oil contains large amount of free fatty acid. A mutant of Candida tropicalis (M20) obtained after several stages of UV mutation are utilized to produce dicarboxylic acids (DCAs) from the fatty acid distillates of rice bran, soybean, coconut, palm kernel and palm oil. Initially, fermentation study was carried out in shake flasks for 144 h. Products were isolated and identified by GLC analysis. Finally, fermentation was carried out in a 2 L jar fermenter, which yielded 62 g/L and 48 g/L of total dibasic acids from rice bran oil fatty acid distillate and coconut oil fatty acid distillate respectively. FADs can be effectively utilized to produce DCAs of various chain lengths by biooxidation process.

  5. Semi-pilot scale production of citric acid in cane molasses by gamma-ray induced mutants of Aspergillus niger

    Energy Technology Data Exchange (ETDEWEB)

    Islam, M.S.; Begum, R.; Choudhury, N.

    1986-08-01

    Utilizing cane molasses as substrate, semi-pilot scale production of citric acid was investigated in fermentation trays (40 x 35 cm) with several gamma-ray induced mutants of Aspergillus niger. Of the mutants tested, two were found to have high yield efficiency (14/20, 51.06%; 79/20, 50.35%) of sugar to citric acid. The yield of other mutants (HB3, 10/20, 164/20, 277/30 and 112/40) ranged between 30 to 42%. The prospect of utilizing the high yielding mutants for commercial production of citric acid has been discussed.

  6. Semi-pilot scale production of citric acid in cane molasses by gamma-ray induced mutants of Aspergillus niger

    International Nuclear Information System (INIS)

    Islam, M.s.; Begum, R.; Choudhury, N.

    1986-01-01

    Utilizing cane molasses as substrate, semi-pilot scale production of citric acid was investigated in fermentation trays (40 x 35 cm) with several gamma-ray induced mutants of Aspergillus niger. Of the mutants tested, two were found to have high yield efficiency (14/20, 51.06%; 79/20, 50.35%) of sugar to citric acid. The yield of other mutants (HB3, 10/20, 164/20, 277/30 and 112/40) ranged between 30 to 42%. The prospect of utilizing the high yielding mutants for commercial production of citric acid has been discussed. (author)

  7. The glutamate carboxypeptidase AMP1 mediates abscisic acid and abiotic stress responses in Arabidopsis.

    Science.gov (United States)

    Shi, Yiting; Wang, Zheng; Meng, Pei; Tian, Siqi; Zhang, Xiaoyan; Yang, Shuhua

    2013-07-01

    ALTERED MERISTEM PROGRAM1 (AMP1) encodes a glutamate carboxypeptidase that plays an important role in shoot apical meristem development and phytohormone homeostasis. We isolated a new mutant allele of AMP1, amp1-20, from a screen for abscisic acid (ABA) hypersensitive mutants and characterized the function of AMP1 in plant stress responses. amp1 mutants displayed ABA hypersensitivity, while overexpression of AMP1 caused ABA insensitivity. Moreover, endogenous ABA concentration was increased in amp1-20- and decreased in AMP1-overexpressing plants under stress conditions. Application of ABA reduced the AMP1 protein level in plants. Interestingly, amp1 mutants accumulated excess superoxide and displayed hypersensitivity to oxidative stress. The hypersensitivity of amp1 to ABA and oxidative stress was partially rescued by reactive oxygen species (ROS) scavenging agent. Furthermore, amp1 was tolerant to freezing and drought stress. The ABA hypersensitivity and freezing tolerance of amp1 was dependent on ABA signaling. Moreover, amp1 had elevated soluble sugar content and showed hypersensitivity to high concentrations of sugar. By contrast, the contents of amino acids were changed in amp1 mutant compared to the wild-type. This study suggests that AMP1 modulates ABA, oxidative and abotic stress responses, and is involved in carbon and amino acid metabolism in Arabidopsis. © 2013 The Authors. New Phytologist © 2013 New Phytologist Trust.

  8. Responses of Soybean Mutant Lines to Aluminium under In Vitro and In Vivo Condition

    International Nuclear Information System (INIS)

    Yuliasti; Sudarsono

    2011-01-01

    The main limited factors of soybean plants expansion in acid soil are Aluminium (Al) toxicity and low pH. The best approach to solve this problem is by using Al tolerance variety. In vitro or in vivo selections using selective media containing AlCl 3 and induced callus embryonic of mutant lines are reliable methods to develop a new variety. The objectives of this research are to evaluate response of soybean genotypes against AlCl 3 under in vitro and in vivo condition. Addition of 15 part per million (ppm) AlCl 3 into in vitro and in vivo media severely affected plant growth. G3 soybean mutant line was identified as more tolerant than the control soybean cultivar Tanggamus. This mutant line was able to survive under more severe AlCl 3 concentrations (15 ppm) under in vitro conditions. Under in vivo conditions, G1 and G4 mutants were also identified as more tolerant than Tanggamus since they produced more pods and higher dry seed weigh per plant. Moreover, G4 mutant line also produced more dry seed weight per plant than Tanggamus when they were grown on soil containing high Al concentration 8.1 me/100 gr = 81 ppm Al +3 . (author)

  9. Assessing the Role of ETHYLENE RESPONSE FACTOR Transcriptional Repressors in Salicylic Acid-Mediated Suppression of Jasmonic Acid-Responsive Genes.

    Science.gov (United States)

    Caarls, Lotte; Van der Does, Dieuwertje; Hickman, Richard; Jansen, Wouter; Verk, Marcel C Van; Proietti, Silvia; Lorenzo, Oscar; Solano, Roberto; Pieterse, Corné M J; Van Wees, Saskia C M

    2017-02-01

    Salicylic acid (SA) and jasmonic acid (JA) cross-communicate in the plant immune signaling network to finely regulate induced defenses. In Arabidopsis, SA antagonizes many JA-responsive genes, partly by targeting the ETHYLENE RESPONSE FACTOR (ERF)-type transcriptional activator ORA59. Members of the ERF transcription factor family typically bind to GCC-box motifs in the promoters of JA- and ethylene-responsive genes, thereby positively or negatively regulating their expression. The GCC-box motif is sufficient for SA-mediated suppression of JA-responsive gene expression. Here, we investigated whether SA-induced ERF-type transcriptional repressors, which may compete with JA-induced ERF-type activators for binding at the GCC-box, play a role in SA/JA antagonism. We selected ERFs that are transcriptionally induced by SA and/or possess an EAR transcriptional repressor motif. Several of the 16 ERFs tested suppressed JA-dependent gene expression, as revealed by enhanced JA-induced PDF1.2 or VSP2 expression levels in the corresponding erf mutants, while others were involved in activation of these genes. However, SA could antagonize JA-induced PDF1.2 or VSP2 in all erf mutants, suggesting that the tested ERF transcriptional repressors are not required for SA/JA cross-talk. Moreover, a mutant in the co-repressor TOPLESS, that showed reduction in repression of JA signaling, still displayed SA-mediated antagonism of PDF1.2 and VSP2. Collectively, these results suggest that SA-regulated ERF transcriptional repressors are not essential for antagonism of JA-responsive gene expression by SA. We further show that de novo SA-induced protein synthesis is required for suppression of JA-induced PDF1.2, pointing to SA-stimulated production of an as yet unknown protein that suppresses JA-induced transcription. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  10. Cyclopropane fatty acid synthase mutants of probiotic human-derived Lactobacillus reuteri are defective in TNF inhibition.

    Science.gov (United States)

    Jones, Sara E; Whitehead, Kristi; Saulnier, Delphine; Thomas, Carissa M; Versalovic, James; Britton, Robert A

    2011-01-01

    Although commensal microbes have been shown to modulate host immune responses, many of the bacterial factors that mediate immune regulation remain unidentified. Select strains of human-derived Lactobacillus reuteri synthesize immunomodulins that potently inhibit production of the inflammatory cytokine TNF. In this study, genetic and genomic approaches were used to identify and investigate L. reuteri genes required or human TNF immunomodulatory activity. Analysis of membrane fatty acids from multiple L. reuteri strains cultured in MRS medium showed that only TNF inhibitory strains produced the cyclopropane fatty acid (CFA) lactobacillic acid. The enzyme cyclopropane fatty acid synthase is required for synthesis of CFAs such as lactobacillic acid, therefore the cfa gene was inactivated and supernatants from the cfa mutant strain were assayed for TNF inhibitory activity. We found that supernatants from the wild-type strain, but not the cfa mutant, suppressed TNF production by activated THP-1 human monocytoid cells Although this suggested a direct role for lactobacillic acid in immunomodulation, purified lactobacillic acid did not suppress TNF at physiologically relevant concentrations. We further analyzed TNF inhibitory and TNF non-inhibitory strains under different growth conditions and found that lactobacillic acid production did not correlate with TNF inhibition. These results indicate that cfa indirectly contributed to L. reuter immunomodulatory activity and suggest that other mechanisms, such as decreased membrane fluidity or altered expression of immunomodulins, result in the loss of TNF inhibitory activity. By increasing our understanding of immunomodulation by probiotic species, beneficial microbes can be rationally selected to alleviate intestinal inflammation.

  11. Citric acid fermentation by gamma ray induced mutants of Aspergillus niger in different carbohydrate media

    Energy Technology Data Exchange (ETDEWEB)

    Anjuman Ara Begum; Naiyyum Choudhury; Mohammad Serajul Islam (Institute of Food and Radiation Biology, Dacca (Bangladesh))

    1990-01-01

    A natural isolate of Aspergillus niger, CA16, and two of its second step mutants, 136/40 and 277/30, grown on different sugar substrates gave maximum citric acid yields of 34, 70, and 126 mg/ml respectively in sucrose medium. Combination of two sugars in the medium at 50% of each improved the yields of citric acid for the sucrose: glucose, glucose: sorbitol, glucose: xylose, and xylose: sorbitol combinations with the mutant strains. Inclusion of galactose in combinations decreased the citric acid yield. (author).

  12. Citric acid fermentation by gamma ray induced mutants of Aspergillus niger in different carbohydrate media

    International Nuclear Information System (INIS)

    Anjuman Ara Begum; Naiyyum Choudhury; Mohammad Serajul Islam

    1990-01-01

    A natural isolate of Aspergillus niger, CA16, and two of its second step mutants, 136/40 and 277/30, grown on different sugar substrates gave maximum citric acid yields of 34, 70, and 126 mg/ml respectively in sucrose medium. Combination of two sugars in the medium at 50% of each improved the yields of citric acid for the sucrose: glucose, glucose: sorbitol, glucose: xylose, and xylose: sorbitol combinations with the mutant strains. Inclusion of galactose in combinations decreased the citric acid yield. (author)

  13. Arabidopsis AtDjA3 null mutant shows increased sensitivity to abscisic acid, salt, and osmotic stress in germination and postgermination stages

    Directory of Open Access Journals (Sweden)

    Silvia eSalas-Muñoz

    2016-02-01

    Full Text Available DnaJ proteins are essential co-chaperones involved in abiotic and biotic stress responses. Arabidopsis AtDjA3 gene encodes a molecular co-chaperone of 420 amino acids, which belongs to the J-protein family. In this study, we report the functional characterization of the AtDjA3 gene using the Arabidopsis knockout line designated j3 and the 35S::AtDjA3 overexpression lines. Loss of AtDjA3 function was associated with small seed production. In fact, j3 mutant seeds showed a reduction of 24% in seed weight compared to Col-0 seeds. Expression analysis showed that the AtDjA3 gene was modulated in response to NaCl, glucose, and abscisic acid. The j3 line had increased sensitivity to NaCl and glucose treatments in the germination and cotyledon development in comparison to parental Col-0. Furthermore, the j3 mutant line exhibited higher abscisic acid sensitivity in comparison to parental Col-0 and 35S::AtDjA3 overexpression lines. In addition, we examined the expression of ABI3 gene, which is a central regulator in ABA signalling, in j3 mutant and 35S::AtDjA3 overexpression lines. Under 5 μM ABA treatment at 24 h, j3 mutant seedlings displayed higher ABI3 expression, whereas in 35S::AtDjA3 overexpression lines, ABI3 gene expression was repressed. Taken together, these results demonstrate that the AtDjA3 gene is involved in seed development and abiotic stress tolerance.

  14. Increased production of pyruvic acid by Escherichia coli RNase G mutants in combination with cra mutations.

    Science.gov (United States)

    Sakai, Taro; Nakamura, Naoko; Umitsuki, Genryou; Nagai, Kazuo; Wachi, Masaaki

    2007-08-01

    The Escherichia coli RNase G is known as an endoribonuclease responsible for the 5'-end maturation of 16S rRNA and degradation of several specific mRNAs such as adhE and eno mRNAs. In this study, we found that an RNase G mutant derived from the MC1061 strain did not grow on a glucose minimal medium. Genetic analysis revealed that simultaneous defects of cra and ilvIH, encoding a transcriptional regulator of glycolysis/gluconeogenesis and one of isozymes of acetohydroxy acid synthase, respectively, were required for this phenomenon to occur. The results of additional experiments presented here indicate that the RNase G mutation, in combination with cra mutation, caused the increased production of pyruvic acid from glucose, which was then preferentially converted to valine due to the ilvIH mutation, resulting in depletion of isoleucine. In fact, the rng cra double mutant produced increased amount of pyruvate in the medium. These results suggest that the RNase G mutation could be applied in the breeding of producer strains of pyruvate and its derivatives such as valine.

  15. Arabidopsis YAK1 regulates abscisic acid response and drought resistance.

    Science.gov (United States)

    Kim, Dongjin; Ntui, Valentine Otang; Xiong, Liming

    2016-07-01

    Abscisic acid (ABA) is an important phytohormone that controls several plant processes such as seed germination, seedling growth, and abiotic stress response. Here, we report that AtYak1 plays an important role in ABA signaling and postgermination growth in Arabidopsis. AtYak1 knockout mutant plants were hyposensitive to ABA inhibition of seed germination, cotyledon greening, seedling growth, and stomatal movement. atyak1-1 mutant plants display reduced drought stress resistance, as evidenced by water loss rate and survival rate. Molecular genetic analysis revealed that AtYak1 deficiency led to elevated expression of stomatal-related gene, MYB60, and down-regulation of several stress-responsive genes. Altogether, these results indicate that AtYak1 plays a role as a positive regulator in ABA-mediated drought response in Arabidopsis. © 2016 Federation of European Biochemical Societies.

  16. Arabidopsis YAK1 regulates abscisic acid response and drought resistance

    KAUST Repository

    Kim, Dongjin

    2016-06-06

    Abscisic acid (ABA) is an important phytohormone that controls several plant processes such as seed germination, seedling growth, and abiotic stress response. Here, we report that AtYak1 plays an important role in ABA signaling and postgermination growth in Arabidopsis. AtYak1 knockout mutant plants were hyposensitive to ABA inhibition of seed germination, cotyledon greening, seedling growth, and stomatal movement. atyak1-1 mutant plants display reduced drought stress resistance, as evidenced by water loss rate and survival rate. Molecular genetic analysis revealed that AtYak1 deficiency led to elevated expression of stomatal-related gene, MYB60, and down-regulation of several stress-responsive genes. Altogether, these results indicate that AtYak1 plays a role as a positive regulator in ABA-mediated drought response in Arabidopsis. © 2016 Federation of European Biochemical Societies.

  17. Isoenzymes performance of some rice varieties and their mutants

    International Nuclear Information System (INIS)

    Winarno, Ermin; Suliwarno, Ambyah; Ismachin, M.

    1992-01-01

    Isoenzymes performance of some rice varieties and their mutants. Genetics studies on alcohol dehydrogenase, malic enzyme, peroxidase, acid phosphase, and aminopeptidase isoenzymes were carried out on several groups of rice varieties and their mutant lines. The first groups consisted of Atomita I, Pelita I/1, A227/5, Mudgo, TN-1, and IR-26. The second group was Cisadane variety and its five mutants, namely OBS 18, OBS 208, OBS 297, OBS 306, and OBS 330. The third group was mutants line 627-10-3 and its mutants, namely 1063, 1066, 1067, 1076, and 1090. Isoenzymes extracts of the rice leaves were fractionated using polyacrylamide gel disc electrophoresis. The pattern of acid phosphate isoenzyme shows the specific character of rice mutants susceptible to brown plant hopper biotype 1. The gene(s) controlling malic enzyme in Cisadane's mutants is (are) estimated more resistant toward gamma irradiation than gene(s) responsible for controlling the other enzymes. Generally, the isoenzymes zymograms show that gene(s) controlling the mutants enzyme have undergone mutation. This case is shown by the changes of Rm value, as well as the amount and intensity of mutants bands. (authors). 7 refs., 7 figs

  18. Use of X-ray and gamma-induced mutants of lactic acid bacteria in the manufacture of dairy products

    Energy Technology Data Exchange (ETDEWEB)

    Dilanian, Z; Makarian, K; Chuprina, D [Erevan Zootechnical and Veterinary Inst. (USSR). Chair of Dairying

    1976-04-01

    With the aid of X-ray and gamma irradiation were got mutants of lactic acid bacteria, which steadily retain acquired properties. Use of proteolytically active mutant strains in the production of armianski and sovetski cheeses shortened the time of their ripening and increased their quality. Gamma-mutant strain L. lactis 1621/I-M with high phenolstability was received and antibiotic activity with respect to some representatives of pathogenic microflora of the bowels. Use of this mutant in starters for sour milk products will raise their therapeutic effect against intestinal diseases. Deep morphological changes are taking place in lactic acid bacteria under the influence of ionizing radiation.

  19. Use of X-ray and gamma-induced mutants of lactic acid bacteria in the manufacture of dairy products

    International Nuclear Information System (INIS)

    Dilanian, Z.; Makarian, K.; Chuprina, D.

    1976-01-01

    With the aid of X-ray and gamma irradiation were got mutants of lactic acid bacteria, which steadily retain acquired properties. Use of proteolytically active mutant strains in the production of armianski and sovetski cheeses shortened the time of their ripening and increased their quality. Gamma-mutant strain L. lactis 1621/I-M with high phenolstability was received and antibiotic activity with respect to some representatives of pathogenic microflora of the bowels. Use of this mutant in starters for sour milk products will raise their therapeutic effect against intestinal diseases. Deep morphological changes are taking place in lactic acid bacteria under the influence of ionizing radiation. (orig.) [de

  20. Kinetics Studies on citric acid production by gamma ray induced mutant of Aspergillus niger

    International Nuclear Information System (INIS)

    Begum, A.A.; Choudhury, N.; Islam, M.S.

    1991-01-01

    Effect of cultural pH and incubation temperature on citric acid yield and kinetic patterns of citric acid fermentation by a natural isolate of aspergillus niger as CA16 and one of its gamma ray induced mutants were studied using cane molasses as growth and fermentation substrate. Mutant strain, 277/30 gave maximum citric acid yield of 85 g/l at pH 3.5 and 28 degree centigrade in molasses medium adjusted to 16% sugar and 25% prescott salt in the medium. Parent strain, CA16 gave a maximum yield of 34 g/l at pH 4.0 and 26 degree centigrade in molasses medium adjusted to 16% sugar and 100% prescott salt in the medium. In kinetic studies, strains showed combination kinetics of citric acid fermentation where product formation is directly related to growth and cell mass and indirectly related to carbohydrate uptake

  1. Arabidopsis decuple mutant reveals the importance of SnRK2 kinases in osmotic stress responses in vivo

    KAUST Repository

    Fujii, Hiroaki

    2011-01-10

    Osmotic stress associated with drought or salinity is a major factor that limits plant productivity. Protein kinases in the SNF1-related protein kinase 2 (SnRK2) family are activated by osmotic stress, suggesting that the kinases are involved in osmotic stress signaling. However, due to functional redundancy, their contribution to osmotic stress responses remained unclear. In this report, we constructed an Arabidopsis line carrying mutations in all 10 members of the SnRK2 family. The decuple mutant snrk2.1/2/3/4/5/6/7/8/9/10 grew poorly under hyperosmotic stress conditions but was similar to the wild type in culture media in the absence of osmotic stress. The mutant was also defective in gene regulation and the accumulation of abscisic acid (ABA), proline, and inositol 1,4,5-trisphosphate under osmotic stress. In addition, analysis of mutants defective in the ABA-activated SnRK2s (snrk2.2/3/6) and mutants defective in the rest of the SnRK2s (snrk2.1/4/5/7/8/9/10) revealed that SnRK2s are a merging point of ABA-dependent and -independent pathways for osmotic stress responses. These results demonstrate critical functions of the SnRK2s in mediating osmotic stress signaling and tolerance.

  2. Evaluating Genetic Variability of Sorghum Mutant Lines Tolerant to Acid Soil

    International Nuclear Information System (INIS)

    Puspitasari, W.; Human, S.; Wirnas, D.; Trikoesoemaningtyas

    2012-01-01

    High rainfall in some parts in Indonesia causes soil become acidic. The main constraint of acid soil is phosphor (P) deficiency and aluminum (Al) toxicity which decrease plant productivity. To overcome this problem, it is important to develop a crop variety tolerant to such conditions. Sorghum is probably one of the potential crops to meet that objective. Sorghum has been reported to have wide adaptability to various agro-ecology and can be used as food and animal feed. Unfortunately, sorghum is not Indonesian origin so its genetic variability is still low. From previous breeding works with induced mutation, some promising mutant lines have been developed. These mutant lines were included in the experiment carried out in Tenjo with soil condition was classified as acid soil with pH 4.8 and exchangeable-Al content 2.43 me/100 g. The objectives of this experiment were to study the magnitude of genetic variability of agronomy and grain quality characters in sorghum in order to facilitate the breeding improvement of the species. Plant materials used in this study were ten genotypes, including 6 mutant lines and 4 control varieties. The randomized block design with three replications was used in the experiment. The genetic variabilities of agronomic and grain quality characters existed among genotypes, such as plant height, number of leaves, stalk diameter, biomass weight, panicle length, grain yield per plant, 100 seed weight and tannin content in the grain. The broad sense heritabilities of agronomic characters were estimated ranging from medium to high. Grain yield showed significantly positive correlation with agronomic characters observed, but it was negatively correlated with protein content (author)

  3. Unravelling molecular responses to moderate dehydration in harvested fruit of sweet orange (Citrus sinensis L. Osbeck) using a fruit-specific ABA-deficient mutant.

    Science.gov (United States)

    Romero, Paco; Rodrigo, María J; Alférez, Fernando; Ballester, Ana-Rosa; González-Candelas, Luis; Zacarías, Lorenzo; Lafuente, María T

    2012-04-01

    Water stress affects many agronomic traits that may be regulated by the phytohormone abscisic acid (ABA). Within these traits, loss of fruit quality becomes important in many citrus cultivars that develop peel damage in response to dehydration. To study peel dehydration transcriptional responsiveness in harvested citrus fruit and the putative role of ABA in this process, this study performed a comparative large-scale transcriptional analysis of water-stressed fruits of the wild-type Navelate orange (Citrus sinesis L. Osbeck) and its spontaneous ABA-deficient mutant Pinalate, which is more prone to dehydration and to developing peel damage. Major changes in gene expression occurring in the wild-type line were impaired in the mutant fruit. Gene ontology analysis revealed the ability of Navelate fruits to induce the response to water deprivation and di-, tri-valent inorganic cation transport biological processes, as well as repression of the carbohydrate biosynthesis process in the mutant. Exogenous ABA triggered relevant transcriptional changes and repressed the protein ubiquitination process, although it could not fully rescue the physiological behaviour of the mutant. Overall, the results indicated that dehydration responsiveness requires ABA-dependent and -independent signals, and highlight that the ability of citrus fruits to trigger molecular responses against dehydration is an important factor in reducing their susceptibility to developing peel damage.

  4. Arabidopsis thaliana cdd1 mutant uncouples the constitutive activation of salicylic acid signalling from growth defects.

    Science.gov (United States)

    Swain, Swadhin; Roy, Shweta; Shah, Jyoti; Van Wees, Saskia; Pieterse, Corné M; Nandi, Ashis K

    2011-12-01

    Arabidopsis genotypes with a hyperactive salicylic acid-mediated signalling pathway exhibit enhanced disease resistance, which is often coupled with growth and developmental defects, such as dwarfing and spontaneous necrotic lesions on the leaves, resulting in reduced biomass yield. In this article, we report a novel recessive mutant of Arabidopsis, cdd1 (constitutive defence without defect in growth and development1), that exhibits enhanced disease resistance associated with constitutive salicylic acid signalling, but without any observable pleiotropic phenotype. Both NPR1 (NON-EXPRESSOR OF PATHOGENESIS-RELATED GENES1)-dependent and NPR1-independent salicylic acid-regulated defence pathways are hyperactivated in cdd1 mutant plants, conferring enhanced resistance against bacterial pathogens. However, a functional NPR1 allele is required for the cdd1-conferred heightened resistance against the oomycete pathogen Hyaloperonospora arabidopsidis. Salicylic acid accumulates at elevated levels in cdd1 and cdd1 npr1 mutant plants and is necessary for cdd1-mediated PR1 expression and disease resistance phenotypes. In addition, we provide data which indicate that the cdd1 mutation negatively regulates the npr1 mutation-induced hyperactivation of ethylene/jasmonic acid signalling. © 2011 The Authors. Molecular Plant Pathology © 2011 BSPP and Blackwell Publishing Ltd.

  5. Purkinje Cell Compartmentation in the Cerebellum of the Lysosomal Acid Phosphatase 2 Mutant Mouse (Nax - Naked-Ataxia Mutant Mouse)

    Science.gov (United States)

    Bailey, Karen; Rahimi Balaei, Maryam; Mannan, Ashraf; Del Bigio, Marc R.; Marzban, Hassan

    2014-01-01

    The Acp2 gene encodes the beta subunit of lysosomal acid phosphatase, which is an isoenzyme that hydrolyzes orthophosphoric monoesters. In mice, a spontaneous mutation in Acp2 results in severe cerebellar defects. These include a reduced size, abnormal lobulation, and an apparent anterior cerebellar disorder with an absent or hypoplastic vermis. Based on differential gene expression in the cerebellum, the mouse cerebellar cortex can normally be compartmentalized anteroposteriorly into four transverse zones and mediolaterally into parasagittal stripes. In this study, immunohistochemistry was performed using various Purkinje cell compartmentation markers to examine their expression patterns in the Acp2 mutant. Despite the abnormal lobulation and anterior cerebellar defects, zebrin II and PLCβ4 showed similar expression patterns in the nax mutant and wild type cerebellum. However, fewer stripes were found in the anterior zone of the nax mutant, which could be due to a lack of Purkinje cells or altered expression of the stripe markers. HSP25 expression was uniform in the central zone of the nax mutant cerebellum at around postnatal day (P) 18–19, suggesting that HSP25 immunonegative Purkinje cells are absent or delayed in stripe pattern expression compared to the wild type. HSP25 expression became heterogeneous around P22–23, with twice the number of parasagittal stripes in the nax mutant compared to the wild type. Aside from reduced size and cortical disorganization, both the posterior zone and nodular zone in the nax mutant appeared less abnormal than the rest of the cerebellum. From these results, it is evident that the anterior zone of the nax mutant cerebellum is the most severely affected, and this extends beyond the primary fissure into the rostral central zone/vermis. This suggests that ACP2 has critical roles in the development of the anterior cerebellum and it may regulate anterior and central zone compartmentation. PMID:24722417

  6. Masking responses to light in period mutant mice.

    Science.gov (United States)

    Pendergast, Julie S; Yamazaki, Shin

    2011-10-01

    Masking is an acute effect of an external signal on an overt rhythm and is distinct from the process of entrainment. In the current study, we investigated the phase dependence and molecular mechanisms regulating masking effects of light pulses on spontaneous locomotor activity in mice. The circadian genes, Period1 (Per1) and Per2, are necessary components of the timekeeping machinery and entrainment by light appears to involve the induction of the expression of Per1 and Per2 mRNAs in the suprachiasmatic nuclei (SCN). We assessed the roles of the Per genes in regulating masking by assessing the effects of light pulses on nocturnal locomotor activity in C57BL/6J Per mutant mice. We found that Per1(-/-) and Per2(-/-) mice had robust negative masking responses to light. In addition, the locomotor activity of Per1(-/-)/Per2(-/-) mice appeared to be rhythmic in the light-dark (LD) cycle, and the phase of activity onset was advanced (but varied among individual mice) relative to lights off. This rhythm persisted for 1 to 2 days in constant darkness in some Per1(-/-)/Per2(-/-) mice. Furthermore, Per1(-/-)/Per2(-/-) mice exhibited robust negative masking responses to light. Negative masking was phase dependent in wild-type mice such that maximal suppression was induced by light pulses at zeitgeber time 14 (ZT14) and gradually weaker suppression occurred during light pulses at ZT16 and ZT18. By measuring the phase shifts induced by the masking protocol (light pulses were administered to mice maintained in the LD cycle), we found that the phase responsiveness of Per mutant mice was altered compared to wild-types. Together, our data suggest that negative masking responses to light are robust in Per mutant mice and that the Per1(-/-)/Per2(-/-) SCN may be a light-driven, weak/damping oscillator.

  7. An active site mutant of Escherichia coli cyclopropane fatty acid synthase forms new non-natural fatty acids providing insights on the mechanism of the enzymatic reaction.

    Science.gov (United States)

    E, Guangqi; Drujon, Thierry; Correia, Isabelle; Ploux, Olivier; Guianvarc'h, Dominique

    2013-12-01

    We have produced and purified an active site mutant of the Escherichia coli cyclopropane fatty acid synthase (CFAS) by replacing the strictly conserved G236 within cyclopropane synthases, by a glutamate residue, which corresponds to E146 of the homologous mycolic acid methyltransferase, Hma, producing hydroxymethyl mycolic acids. The G236E CFAS mutant had less than 1% of the in vitro activity of the wild type enzyme. We expressed the G236E CFAS mutant in an E. coli (DE3) strain in which the chromosomal cfa gene had been deleted. After extraction of phospholipids and conversion into the corresponding fatty acid methyl esters (FAMEs), we observed the formation of cyclopropanated FAMEs suggesting that the mutant retained some of the normal activity in vivo. However, we also observed the formation of new C17 methyl-branched unsaturated FAMEs whose structures were determined using GC/MS and NMR analyses. The double bond was located at different positions 8, 9 or 10, and the methyl group at position 10 or 9. Thus, this new FAMEs are likely arising from a 16:1 acyl chain of a phospholipid that had been transformed by the G236E CFAS mutant in vivo. The reaction catalyzed by this G236E CFAS mutant thus starts by the methylation of the unsaturated acyl chain at position 10 or 9 yielding a carbocation at position 9 or 10 respectively. It follows then two competing steps, a normal cyclopropanation or hydride shift/elimination events giving different combinations of alkenes. This study not only provides further evidence that cyclopropane synthases (CSs) form a carbocationic intermediate but also opens the way to CSs engineering for the synthesis of non-natural fatty acids. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  8. Effect of abscisic acid on stomatal opening in isolated epidermal strips of abi mutants of Arabidopsis thaliana

    NARCIS (Netherlands)

    Roelfsema, MRG; Prins, HBA

    Abscisic acid-insensitive mutants of Arabidopsis thaliana L. var. Landsberg erecta were selected for their decreased sensitivity to ABA during germination. Two of these mutants, abi-1 and abi-2, display a wilty phenotype as adult plants, indicating disturbed water relations. Experiments were

  9. TolC is important for bacterial survival and oxidative stress response in Salmonella enterica serovar Choleraesuis in an acidic environment.

    Science.gov (United States)

    Lee, Jen-Jie; Wu, Ying-Chen; Kuo, Chih-Jung; Hsuan, Shih-Ling; Chen, Ter-Hsin

    2016-09-25

    The outer membrane protein TolC, which is one of the key components of several multidrug efflux pumps, is thought to be involved in various independent systems in Enterobacteriaceae. Since the acidic environment of the stomach is an important protection barrier against foodborne pathogen infections in hosts, we evaluated whether TolC played a role in the acid tolerance of Salmonella enterica serovar Choleraesuis. Comparison of the acid tolerance of the tolC mutant and the parental wild-type strain showed that the absence of TolC limits the ability of Salmonella to sustain life under extreme acidic conditions. Additionally, the mutant exhibited morphological changes during growth in an acidic medium, leading to the conflicting results of cell viability measured by spectrophotometry and colony-forming unit counting. Reverse-transcriptional-PCR analysis indicated that acid-related molecules, apparatus, or enzymes and oxidation-induced factors were significantly affected by the acidic environment in the null-tolC mutant. The elongated cellular morphology was restored by adding antioxidants to the culture medium. Furthermore, we found that increased cellular antioxidative activity provides an overlapping protection against acid killing, demonstrating the complexity of the bacterial acid stress response. Our findings reinforce the multifunctional characteristics of TolC in acid tolerance or oxidative stress resistance and support the correlative protection mechanism between oxygen- and acid-mediated stress responses in Salmonella enterica serovar Choleraesuis. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Microbial production of squalene by a nicotinic acid-resistant mutant derived from Fusarium sp. No.5-128B

    International Nuclear Information System (INIS)

    Ogawa, T.; Kojima, I.; Takeda, N.; Fukuda, H.

    1994-01-01

    A nicotinic acid-resistant mutant, designated NA201, was obtained from Fusarium sp. no.5-128B by treatment with ultraviolet light. This mutant strain could grow in the presence of up to 500mM nicotinic acid in the culture medium, although the parent strain could not grow at concentrations of nicotinic acid above 200 mM. The Na201 strain exhibited morphological mutations, neither forming aerial hyphae nor secreting a red-brown pigment. However, it retained the resistance to kabicidin at 25 mg-l(-1) of the parent strain. The mutant NA201 cells contained high levels of squalene and low levels of ergosterol, about 53 times higher and five to six times lower, respectively, than those of the parent strain under standard culture conditions. The volumetric oxygen transfer coefficient (Kd) affected the level of squalene in the mutant cells. The Kd for the maximum production of squalene by the mutant was 24 mmol O2 l(-1)h(-1)atm(-1) and the level of squalene in the mutant cells was 26 mg (g cell)(-1) on a dry weight basis. The greatest accumulation of squalene by the Na201 strain, corresponding to 323 mg per liter of culture medium and 35 mg (g cell)(-1) on a dry weight basis, was achieved in a culture in which the Kd was changed from a high to a low value on the third day, with the simultaneous addition of 3% glucose (w/v)

  11. A combined physiological and proteomic approach to reveal lactic-acid-induced alterations in Lactobacillus casei Zhang and its mutant with enhanced lactic acid tolerance.

    Science.gov (United States)

    Wu, Chongde; Zhang, Juan; Chen, Wei; Wang, Miao; Du, Guocheng; Chen, Jian

    2012-01-01

    Lactobacillus casei has traditionally been recognized as a probiotic and frequently used as an adjunct culture in fermented dairy products, where acid stress is an environmental condition commonly encountered. In the present study, we carried out a comparative physiological and proteomic study to investigate lactic-acid-induced alterations in Lactobacillus casei Zhang (WT) and its acid-resistant mutant. Analysis of the physiological data showed that the mutant exhibited 33.8% higher glucose phosphoenolpyruvate:sugar phosphotransferase system activity and lower glycolytic pH compared with the WT under acidic conditions. In addition, significant differences were detected in both cells during acid stress between intracellular physiological state, including intracellular pH, H(+)-ATPase activity, and intracellular ATP pool. Comparison of the proteomic data based on 2D-DIGE and i-TRAQ indicated that acid stress invoked a global change in both strains. The mutant protected the cells against acid damage by regulating the expression of key proteins involved in cellular metabolism, DNA replication, RNA synthesis, translation, and some chaperones. Proteome results were validated by Lactobacillus casei displaying higher intracellular aspartate and arginine levels, and the survival at pH 3.3 was improved 1.36- and 2.10-fold by the addition of 50-mM aspartate and arginine, respectively. To our knowledge, this is the first demonstration that aspartate may be involved in acid tolerance in Lactobacillus casei. Results presented here may help us understand acid resistance mechanisms and help formulate new strategies to enhance the industrial applications of this species.

  12. Masking Responses to Light in Period Mutant Mice

    Science.gov (United States)

    Pendergast, Julie S.; Yamazaki, Shin

    2013-01-01

    Masking is an acute effect of an external signal on an overt rhythm and is distinct from the process of entrainment. In the current study, we investigated the phase dependence and molecular mechanisms regulating masking effects of light pulses on spontaneous locomotor activity in mice. The circadian genes, Period1 (Per1) and Per2, are necessary components of the timekeeping machinery and entrainment by light appears to involve the induction of the expression of Per1 and Per2 mRNAs in the suprachiasmatic nuclei (SCN). We assessed the roles of the Per genes in regulating masking by assessing the effects of light pulses on nocturnal locomotor activity in C57BL/6J Per mutant mice. We found that Per1−/− and Per2−/− mice had robust negative masking responses to light. In addition, the locomotor activity of Per1−/−/Per2−/− mice appeared to be rhythmic in the light-dark (LD) cycle, and the phase of activity onset was advanced (but varied among individual mice) relative to lights off. This rhythm persisted for 1 to 2 days in constant darkness in some Per1−/−/Per2−/− mice. Furthermore, Per1−/−/Per2−/− mice exhibited robust negative masking responses to light. Negative masking was phase dependent in wild-type mice such that maximal suppression was induced by light pulses at zeitgeber time 14 (ZT14) and gradually weaker suppression occurred during light pulses at ZT16 and ZT18. By measuring the phase shifts induced by the masking protocol (light pulses were administered to mice maintained in the LD cycle), we found that the phase responsiveness of Per mutant mice was altered compared to wild-types. Together, our data suggest that negative masking responses to light are robust in Per mutant mice and that the Per1−/−/Per2−/− SCN may be a light-driven, weak/damping oscillator. PMID:21793695

  13. Isolation of a spontaneous CHO amino acid transport mutant by a combination of tritium suicide and replica plating

    International Nuclear Information System (INIS)

    Dantzig, A.H.; Slayman, C.W.; Adelberg, E.A.

    1982-01-01

    A spontaneous transport mutant of Chinese hamster ovary cells, CHY-1, was isolated by a combination of [ 3 H]proline suicide and replica plating. The mutant took up less tritium than the parent, resulting in a lower killing rate during storage. Transport by four separate amino acid transport systems (A, ASC, L, Ly+) was examined. The CHY-1 mutant exhibited normal uptake via the ASC, L, and Ly+ systems. By contrast, uptake of the most specific substrate of the A system, 2-(methylamino)-isobutyric acid, was significantly reduced at low, but not high, concentrations, due to a 3.5-fold increase in Km and a 1.5-fold increase in Vmax. Taken together, these data suggest that the CHY-1 mutation may be in the structural gene coding for the A transport protein. The tritium suicide procedure is discussed, and general equations are derived to predict the maximum storage time for the survival of one mutant cell and the optimum size of the cell population for maximum mutant enrichment

  14. Graviresponsiveness and abscisic-acid content of roots of carotenoid-deficient mutants of Zea mays L

    Science.gov (United States)

    Moore, R.; Smith, J. D.

    1985-01-01

    The abscisic-acid (ABA) content of roots of the carotenoid-deficient w-3, vp-5, and vp-7 mutants of Z. mays was analyzed using gas chromatography-mass spectrometry with an analysis sensitivity of 6 ng ABA g-1 fresh weight (FW). Roots of normal seedlings of the same lines were characterized by the following amounts of ABA (as ng ABA g-1 FW, +/- standard deviation): w-3, 279 +/- 43; vp-5, 237 +/- 26; vp-7, 338 +/- 61. We did not detect any ABA in roots of any of the mutants. Thus, the lack of carotenoids in these mutants correlated positively with the apparent absence of ABA. Primary roots of normal and mutant seedlings were positively gravitropic, with no significant differences in the curvatures of roots of normal as compared with mutant seedlings. These results indicate that ABA 1) is synthesized in maize roots via the carotenoid pathway, and 2) is not necessary for positive gravitropism by primary roots of Z. mays.

  15. Gallic acid induces apoptosis in EGFR-mutant non-small cell lung cancers by accelerating EGFR turnover.

    Science.gov (United States)

    Nam, Boas; Rho, Jin Kyung; Shin, Dong-Myung; Son, Jaekyoung

    2016-10-01

    Gallic acid is a common botanic phenolic compound, which is present in plants and foods worldwide. Gallic acid is implicated in various biological processes such as cell growth and apoptosis. Indeed, gallic acid has been shown to induce apoptosis in many cancer types. However, the molecular mechanisms of gallic acid-induced apoptosis in cancer, particularly lung cancer, are still unclear. Here, we report that gallic acid induces apoptosis in EGFR-mutant non-small cell lung cancer (NSCLC) cells, but not in EGFR-WT NSCLC cells. Treatment with gallic acid resulted in a significant reduction in proliferation and induction of apoptosis, only in EGFR-mutant NSCLC cells. Interestingly, treatment with gallic acid led to a robust decrease in EGFR levels, which is critical for NSCLC survival. Treatment with gallic acid had no significant effect on transcription, but induced EGFR turnover. Indeed, treatment with a proteasome inhibitor dramatically reversed gallic acid-induced EGFR downregulation. Moreover, treatment with gallic acid induced EGFR turnover leading to apoptosis in EGFR-TKI (tyrosine kinase inhibitor)-resistant cell lines, which are dependent on EGFR signaling for survival. Thus, these studies suggest that gallic acid can induce apoptosis in EGFR-dependent lung cancers that are dependent on EGFR for growth and survival via acceleration of EGFR turnover. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Identification of a Classical Mutant in the Industrial Host Aspergillus niger by Systems Genetics: LaeA Is Required for Citric Acid Production and Regulates the Formation of Some Secondary Metabolites

    Directory of Open Access Journals (Sweden)

    Jing Niu

    2016-01-01

    Full Text Available The asexual filamentous fungus Aspergillus niger is an important industrial cell factory for citric acid production. In this study, we genetically characterized a UV-generated A. niger mutant that was originally isolated as a nonacidifying mutant, which is a desirable trait for industrial enzyme production. Physiological analysis showed that this mutant did not secrete large amounts of citric acid and oxalic acid, thus explaining the nonacidifying phenotype. As traditional complementation approaches to characterize the mutant genotype were unsuccessful, we used bulk segregant analysis in combination with high-throughput genome sequencing to identify the mutation responsible for the nonacidifying phenotype. Since A. niger has no sexual cycle, parasexual genetics was used to generate haploid segregants derived from diploids by loss of whole chromosomes. We found that the nonacidifying phenotype was caused by a point mutation in the laeA gene. LaeA encodes a putative methyltransferase-domain protein, which we show here to be required for citric acid production in an A. niger lab strain (N402 and in other citric acid production strains. The unexpected link between LaeA and citric acid production could provide new insights into the transcriptional control mechanisms related to citric acid production in A. niger. Interestingly, the secondary metabolite profile of a ΔlaeA strain differed from the wild-type strain, showing both decreased and increased metabolite levels, indicating that LaeA is also involved in regulating the production of secondary metabolites. Finally, we show that our systems genetics approach is a powerful tool to identify trait mutations.

  17. Identification of a Classical Mutant in the Industrial Host Aspergillus niger by Systems Genetics: LaeA Is Required for Citric Acid Production and Regulates the Formation of Some Secondary Metabolites.

    Science.gov (United States)

    Niu, Jing; Arentshorst, Mark; Nair, P Deepa S; Dai, Ziyu; Baker, Scott E; Frisvad, Jens C; Nielsen, Kristian F; Punt, Peter J; Ram, Arthur F J

    2015-11-13

    The asexual filamentous fungus Aspergillus niger is an important industrial cell factory for citric acid production. In this study, we genetically characterized a UV-generated A. niger mutant that was originally isolated as a nonacidifying mutant, which is a desirable trait for industrial enzyme production. Physiological analysis showed that this mutant did not secrete large amounts of citric acid and oxalic acid, thus explaining the nonacidifying phenotype. As traditional complementation approaches to characterize the mutant genotype were unsuccessful, we used bulk segregant analysis in combination with high-throughput genome sequencing to identify the mutation responsible for the nonacidifying phenotype. Since A. niger has no sexual cycle, parasexual genetics was used to generate haploid segregants derived from diploids by loss of whole chromosomes. We found that the nonacidifying phenotype was caused by a point mutation in the laeA gene. LaeA encodes a putative methyltransferase-domain protein, which we show here to be required for citric acid production in an A. niger lab strain (N402) and in other citric acid production strains. The unexpected link between LaeA and citric acid production could provide new insights into the transcriptional control mechanisms related to citric acid production in A. niger. Interestingly, the secondary metabolite profile of a ΔlaeA strain differed from the wild-type strain, showing both decreased and increased metabolite levels, indicating that LaeA is also involved in regulating the production of secondary metabolites. Finally, we show that our systems genetics approach is a powerful tool to identify trait mutations. Copyright © 2016 Niu et al.

  18. Anti-H-Y responses of H-2b mutant mice.

    Science.gov (United States)

    Simpson, E; Gordon, R D; Chandler, P R; Bailey, D

    1978-10-01

    Two strains of H-2b mutant mice, H-2ba and H-2bf, in which the mutational event took place at H-2K, make anti-H-Y cytotoxic T cell responses which are H-2-restricted, Db-associated and indistinguishable in target cell specificity from those of H-2b mice. Thus, alteration of the H-2K molecule affects neither the Ir gene controlling the response, nor the associative antigen. On the other hand, one H-2Db mutant strain, H-2bo, although it makes a good anti-H-Y cytotoxic response, shows target cell specificity restricted to its own Dbo antigen(s), and neither H-2b, H-2ba or H-2bf anti-H-Y cytotoxic cells kill H-2bo male target cells. Thus, the alteration of the H-2Db molecule does not affect the Ir gene of H-2b mice, but it does alter the H-2Db-associative antigen.

  19. Ethylene signaling renders the jasmonate response of Arabidopsis insensitive to future suppression by salicylic acid

    OpenAIRE

    Leon Reyes, H.A.; Du, Y.; Koornneef, A.; Proietti, S.; Körbes, A.P.; Memelink, J.; Pieterse, C.M.J.; Ritsema, T.

    2010-01-01

    Cross-talk between jasmonate (JA), ethylene (ET), and Salicylic acid (SA) signaling is thought to operate as a mechanism to fine-tune induced defenses that are activated in response to multiple attackers. Here, 43 Arabidopsis genotypes impaired in hormone signaling or defense-related processes were screened for their ability to express SA-mediated suppression of JA-responsive gene expression. Mutant cev1, which displays constitutive expression of JA and ET responses, appeared to be insensitiv...

  20. The Mediator subunit SFR6/MED16 controls defence gene expression mediated by salicylic acid and jasmonate responsive pathways.

    Science.gov (United States)

    Wathugala, Deepthi L; Hemsley, Piers A; Moffat, Caroline S; Cremelie, Pieter; Knight, Marc R; Knight, Heather

    2012-07-01

    • Arabidopsis SENSITIVE TO FREEZING6 (SFR6) controls cold- and drought-inducible gene expression and freezing- and osmotic-stress tolerance. Its identification as a component of the MEDIATOR transcriptional co-activator complex led us to address its involvement in other transcriptional responses. • Gene expression responses to Pseudomonas syringae, ultraviolet-C (UV-C) irradiation, salicylic acid (SA) and jasmonic acid (JA) were investigated in three sfr6 mutant alleles by quantitative real-time PCR and susceptibility to UV-C irradiation and Pseudomonas infection were assessed. • sfr6 mutants were more susceptible to both Pseudomonas syringae infection and UV-C irradiation. They exhibited correspondingly weaker PR (pathogenesis-related) gene expression than wild-type Arabidopsis following these treatments or after direct application of SA, involved in response to both UV-C and Pseudomonas infection. Other genes, however, were induced normally in the mutants by these treatments. sfr6 mutants were severely defective in expression of plant defensin genes in response to JA; ectopic expression of defensin genes was provoked in wild-type but not sfr6 by overexpression of ERF5. • SFR6/MED16 controls both SA- and JA-mediated defence gene expression and is necessary for tolerance of Pseudomonas syringae infection and UV-C irradiation. It is not, however, a universal regulator of stress gene transcription and is likely to mediate transcriptional activation of specific regulons only. © 2012 The Authors. New Phytologist © 2012 New Phytologist Trust.

  1. Bacillus subtilis mutants deficient in the adaptive response to simple alkylating agents

    Energy Technology Data Exchange (ETDEWEB)

    Morohoshi, F.; Munakata, N.

    1985-03-01

    Three mutant strains exhibiting hyper-sensitivity to N-methyl-N'-nitro-N-nitrosoguanidine, but not to methyl methanesulfonate, were selected by a replica method from mutagenized spores of Bacillus subtilis. All three were totally deficient in the adaptive response to N-methyl-N'-nitro-N-nitrosoguanidine with regard to both lethality and mutagenesis. The activity to destroy O/sup 6/-methylguanine residues in the methylated DNA was not elevated in the mutant cells by the pretreatment with sublethal concentrations of N-methyl-N-nitro-N-nitrosoguanidine. This deficiency corresponded to the persistance of O/sup 6/-methylguanine residues in the DNA of both control and pretreated mutant cells challenged with the drug. The lethal and mutagenic sensitivity of the mutant strains were observed only for methyl- or ethyl-nitroso compounds that are thought to be active as inducers and are also active in O-alkylation. Except for the insensitivity to methyl methanesulfonate, the phenotypes of these mutants look very similar to those of ada mutants isolated previously in Escherichia coli.

  2. The Arabidopsis mutant cev1 links cell wall signaling to jasmonate and ethylene responses.

    Science.gov (United States)

    Ellis, Christine; Karafyllidis, Ioannis; Wasternack, Claus; Turner, John G

    2002-07-01

    Biotic and abiotic stresses stimulate the synthesis of jasmonates and ethylene, which, in turn, induce the expression of genes involved in stress response and enhance defense responses. The cev1 mutant has constitutive expression of stress response genes and has enhanced resistance to fungal pathogens. Here, we show that cev1 plants have increased production of jasmonate and ethylene and that its phenotype is suppressed by mutations that interrupt jasmonate and ethylene signaling. Genetic mapping, complementation analysis, and sequence analysis revealed that CEV1 is the cellulose synthase CeSA3. CEV1 was expressed predominantly in root tissues, and cev1 roots contained less cellulose than wild-type roots. Significantly, the cev1 mutant phenotype could be reproduced by treating wild-type plants with cellulose biosynthesis inhibitors, and the cellulose synthase mutant rsw1 also had constitutive expression of VSP. We propose that the cell wall can signal stress responses in plants.

  3. The 7B-1 mutant in tomato shows blue-light-specific resistance to osmotic stress and abscisic acid.

    Science.gov (United States)

    Fellner, Martin; Sawhney, Vipen K

    2002-03-01

    Germination of wild-type (WT) tomato ( Lycopersicon esculentum Mill.) seed is inhibited by mannitol (100-140 mM) in light, but not in darkness, suggesting that light amplifies the responsiveness of the seed to osmotic stress (M. Fellner, V.K. Sawhney (2001) Theor Appl Genet 102:215-221). Here we report that white light (W) and especially blue light (B) strongly enhance the mannitol-induced inhibition of seed germination, and that the effect of red light (R) is weak or nil. The inhibitory effect of mannitol could be completely overcome by fluridone, an inhibitor of abscisic acid (ABA) biosynthesis, indicating that mannitol inhibits seed germination via ABA accumulation in seeds. The inhibition of WT seed germination by exogenous ABA was also amplified by W or B, but not by R. In a recessive, ABA-overproducing, 7B-1 mutant of tomato, seed germination and hypocotyl growth were resistant to inhibition by mannitol or exogenous ABA, both in W or B. Experiments with fluridone suggested that inhibition of hypocotyl growth by W or B is also partially via ABA accumulation. De-etiolation in the mutant was especially less in B compared to the WT, and there was no difference in hypocotyl growth between the two genotypes in R. Our data suggest that B amplifies the responsiveness of tomato seeds and hypocotyls to mannitol and ABA, and that W- or B-specific resistance of the 7B-1 mutant to osmotic stress or ABA is a consequence of a defect in B perception or signal transduction.

  4. A mutation in the Arabidopsis HYL1 gene encoding a dsRNA binding protein affects responses to abscisic acid, auxin, and cytokinin

    Science.gov (United States)

    Lu, C.; Fedoroff, N.

    2000-01-01

    Both physiological and genetic evidence indicate interconnections among plant responses to different hormones. We describe a pleiotropic recessive Arabidopsis transposon insertion mutation, designated hyponastic leaves (hyl1), that alters the plant's responses to several hormones. The mutant is characterized by shorter stature, delayed flowering, leaf hyponasty, reduced fertility, decreased rate of root growth, and an altered root gravitropic response. It also exhibits less sensitivity to auxin and cytokinin and hypersensitivity to abscisic acid (ABA). The auxin transport inhibitor 2,3,5-triiodobenzoic acid normalizes the mutant phenotype somewhat, whereas another auxin transport inhibitor, N-(1-naph-thyl)phthalamic acid, exacerbates the phenotype. The gene, designated HYL1, encodes a 419-amino acid protein that contains two double-stranded RNA (dsRNA) binding motifs, a nuclear localization motif, and a C-terminal repeat structure suggestive of a protein-protein interaction domain. We present evidence that the HYL1 gene is ABA-regulated and encodes a nuclear dsRNA binding protein. We hypothesize that the HYL1 protein is a regulatory protein functioning at the transcriptional or post-transcriptional level.

  5. Identification of a novel ga-related bush mutant in pumpkin (cucurbita moschata duchesne)

    International Nuclear Information System (INIS)

    Wu, T.; Cao, J.

    2015-01-01

    Pumpkin (Cucurbita moschata Duchesne) bush mutant plants were characterized by short stems. The sensitivity of pumpkin bush mutant plants to exogenous hormones was identified in this study. Results revealed that internode elongation of bush mutant plants could respond to gibberellins (GA4+7 and GA3), but not to indole acetic acid (IAA) and brassinosteroids (BR); by contrast, the mutant phenotype of bush mutant plants could not be fully rescued by GA4+7 and GA3. The internode of bush mutant plants yielded a lower KS expression level than that of vine plants. Therefore, pumpkin bush mutant plants were designated as GA-related mutant plants eliciting a partial response to GAs; the action of IAA and BR might not be involved in the internode growth of pumpkin bush mutant plants, specifically Cucurbita moschata Duch. (author)

  6. Comparative metabolic profiling of mce1 operon mutant vs wild-type Mycobacterium tuberculosis strains.

    Science.gov (United States)

    Queiroz, Adriano; Medina-Cleghorn, Daniel; Marjanovic, Olivera; Nomura, Daniel K; Riley, Lee W

    2015-11-01

    Mycobacterium tuberculosis disrupted in a 13-gene operon (mce1) accumulates free mycolic acids (FM) in its cell wall and causes accelerated death in mice. Here, to more comprehensively analyze differences in their cell wall lipid composition, we used an untargeted metabolomics approach to compare the lipid profiles of wild-type and mce1 operon mutant strains. By liquid chromatography-mass spectrometry, we identified >400 distinct lipids significantly altered in the mce1 mutant compared to wild type. These lipids included decreased levels of saccharolipids and glycerophospholipids, and increased levels of alpha-, methoxy- and keto mycolic acids (MA), and hydroxyphthioceranic acid. The mutant showed reduced expression of mmpL8, mmpL10, stf0, pks2 and papA2 genes involved in transport and metabolism of lipids recognized to induce proinflammatory response; these lipids were found to be decreased in the mutant. In contrast, the transcripts of mmpL3, fasI, kasA, kasB, acpM and RV3451 involved in MA transport and metabolism increased; MA inhibits inflammatory response in macrophages. Since the mce1 operon is known to be regulated in intracellular M. tuberculosis, we speculate that the differences we observed in cell wall lipid metabolism and composition may affect host response to M. tuberculosis infection and determine the clinical outcome of such an infection. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  7. Lactobacillus casei combats acid stress by maintaining cell membrane functionality.

    Science.gov (United States)

    Wu, Chongde; Zhang, Juan; Wang, Miao; Du, Guocheng; Chen, Jian

    2012-07-01

    Lactobacillus casei strains have traditionally been recognized as probiotics and frequently used as adjunct culture in fermented dairy products where lactic acid stress is a frequently encountered environmental condition. We have investigated the effect of lactic acid stress on the cell membrane of L. casei Zhang [wild type (WT)] and its acid-resistant mutant Lbz-2. Both strains were grown under glucose-limiting conditions in chemostats; following challenge by low pH, the cell membrane stress responses were investigated. In response to acid stress, cell membrane fluidity decreased and its fatty acid composition changed to reduce the damage caused by lactic acid. Compared with the WT, the acid-resistant mutant exhibited numerous survival advantages, such as higher membrane fluidity, higher proportions of unsaturated fatty acids, and higher mean chain length. In addition, cell integrity analysis showed that the mutant maintained a more intact cellular structure and lower membrane permeability after environmental acidification. These results indicate that alteration in membrane fluidity, fatty acid distribution, and cell integrity are common mechanisms utilized by L. casei to withstand severe acidification and to reduce the deleterious effect of lactic acid on the cell membrane. This detailed comparison of cell membrane responses between the WT and mutant add to our knowledge of the acid stress adaptation and thus enable new strategies to be developed aimed at improving the industrial performance of this species under acid stress.

  8. Enhanced Abscisic Acid-Mediated Responses in nia1nia2noa1-2 Triple Mutant Impaired in NIA/NR- and AtNOA1-Dependent Nitric Oxide Biosynthesis in Arabidopsis1[W

    Science.gov (United States)

    Lozano-Juste, Jorge; León, José

    2010-01-01

    Nitric oxide (NO) regulates a wide range of plant processes from development to environmental adaptation. Despite its reported regulatory functions, it remains unclear how NO is synthesized in plants. We have generated a triple nia1nia2noa1-2 mutant that is impaired in nitrate reductase (NIA/NR)- and Nitric Oxide-Associated1 (AtNOA1)-mediated NO biosynthetic pathways. NO content in roots of nia1nia2 and noa1-2 plants was lower than in wild-type plants and below the detection limit in nia1nia2noa1-2 plants. NIA/NR- and AtNOA1-mediated biosynthesis of NO were thus active and responsible for most of the NO production in Arabidopsis (Arabidopsis thaliana). The nia1nia2noa1-2 plants displayed reduced size, fertility, and seed germination potential but increased dormancy and resistance to water deficit. The increasing deficiency in NO of nia1nia2, noa1-2, and nia1nia2noa1-2 plants correlated with increased seed dormancy, hypersensitivity to abscisic acid (ABA) in seed germination and establishment, as well as dehydration resistance. In nia1nia2noa1-2 plants, enhanced drought tolerance was due to a very efficient stomata closure and inhibition of opening by ABA, thus uncoupling NO from ABA-triggered responses in NO-deficient guard cells. The NO-deficient mutants in NIA/NR- and AtNOA1-mediated pathways in combination with the triple mutant will be useful tools to functionally characterize the role of NO and the contribution of both biosynthetic pathways in regulating plant development and defense. PMID:20007448

  9. Plastid Located WHIRLY1 Enhances the Responsiveness of Arabidopsis Seedlings Toward Abscisic Acid

    Science.gov (United States)

    Isemer, Rena; Krause, Kirsten; Grabe, Nils; Kitahata, Nobutaka; Asami, Tadao; Krupinska, Karin

    2012-01-01

    WHIRLY1 is a protein that can be translocated from the plastids to the nucleus, making it an ideal candidate for communicating information between these two compartments. Mutants of Arabidopsis thaliana lacking WHIRLY1 (why1) were shown to have a reduced sensitivity toward salicylic acid (SA) and abscisic acid (ABA) during germination. Germination assays in the presence of abamine, an inhibitor of ABA biosynthesis, revealed that the effect of SA on germination was in fact caused by a concomitant stimulation of ABA biosynthesis. In order to distinguish whether the plastid or the nuclear isoform of WHIRLY1 is adjusting the responsiveness toward ABA, sequences encoding either the complete WHIRLY1 protein or a truncated form lacking the plastid transit peptide were overexpressed in the why1 mutant background. In plants overexpressing the full-length sequence, WHIRLY1 accumulated in both plastids and the nucleus, whereas in plants overexpressing the truncated sequence, WHIRLY1 accumulated exclusively in the nucleus. Seedlings containing recombinant WHIRLY1 in both compartments were hypersensitive toward ABA. In contrast, seedlings possessing only the nuclear form of WHIRLY1 were as insensitive toward ABA as the why1 mutants. ABA was furthermore shown to lower the rate of germination of wildtype seeds even in the presence of abamine which is known to inhibit the formation of xanthoxin, the plastid located precursor of ABA. From this we conclude that plastid located WHIRLY1 enhances the responsiveness of seeds toward ABA even when ABA is supplied exogenously. PMID:23269926

  10. The transcriptional response of Lactobacillus sanfranciscensis DSM 20451T and its tcyB mutant lacking a functional cystine transporter to diamide stress.

    Science.gov (United States)

    Stetina, Mandy; Behr, Jürgen; Vogel, Rudi F

    2014-07-01

    As a result of its strong adaptation to wheat and rye sourdoughs, Lactobacillus sanfranciscensis has the smallest genome within the genus Lactobacillus. The concomitant absence of some important antioxidative enzymes and the inability to synthesize glutathione suggest a role of cystine transport in maintenance of an intracellular thiol balance. Diamide [synonym 1,1'-azobis(N,N-dimethylformamide)] disturbs intracellular and membrane thiol levels in oxidizing protein thiols depending on its initial concentration. In this study, RNA sequencing was used to reveal the transcriptional response of L. sanfranciscensis DSM 20451(T) (wild type [WT]) and its ΔtcyB mutant with a nonfunctional cystine transporter after thiol stress caused by diamide. Along with the different expression of genes involved in amino acid starvation, pyrimidine synthesis, and energy production, our results show that thiol stress in the wild type can be compensated through activation of diverse chaperones and proteases whereas the ΔtcyB mutant shifts its metabolism in the direction of survival. Only a small set of genes are significantly differentially expressed between the wild type and the mutant. In the WT, mainly genes which are associated with a heat shock response are upregulated whereas glutamine import and synthesis genes are downregulated. In the ΔtcyB mutant, the whole opp operon was more highly expressed, as well as a protein which probably includes enzymes for methionine transport. The two proteins encoded by spxA and nrdH, which are involved in direct or indirect oxidative stress responses, are also upregulated in the mutant. This work emphasizes that even in the absence of definitive antioxidative enzymes, bacteria with a small genome and a high frequency of gene inactivation and elimination use small molecules such as the cysteine/cystine couple to overcome potential cell damage resulting from oxidative stress. Copyright © 2014, American Society for Microbiology. All Rights

  11. Mutant alleles of FAD2-1A and FAD2-1B combine to produce soybeans with the high oleic acid seed oil trait

    Directory of Open Access Journals (Sweden)

    Pham Anh-Tung

    2010-09-01

    Full Text Available Abstract Background The alteration of fatty acid profiles in soybean [Glycine max (L. Merr.] to improve soybean oil quality is an important and evolving theme in soybean research to meet nutritional needs and industrial criteria in the modern market. Soybean oil with elevated oleic acid is desirable because this monounsaturated fatty acid improves the nutrition and oxidative stability of the oil. Commodity soybean oil typically contains 20% oleic acid and the target for high oleic acid soybean oil is approximately 80% of the oil; previous conventional plant breeding research to raise the oleic acid level to just 50-60% of the oil was hindered by the genetic complexity and environmental instability of the trait. The objective of this work was to create the high oleic acid trait in soybeans by identifying and combining mutations in two delta-twelve fatty acid desaturase genes, FAD2-1A and FAD2-1B. Results Three polymorphisms found in the FAD2-1B alleles of two soybean lines resulted in missense mutations. For each of the two soybean lines, there was one unique amino acid change within a highly conserved region of the protein. The mutant FAD2-1B alleles were associated with an increase in oleic acid levels, although the FAD2-1B mutant alleles alone were not capable of producing a high oleic acid phenotype. When existing FAD2-1A mutations were combined with the novel mutant FAD2-1B alleles, a high oleic acid phenotype was recovered only for those lines which were homozygous for both of the mutant alleles. Conclusions We were able to produce conventional soybean lines with 80% oleic acid in the oil in two different ways, each requiring the contribution of only two genes. The high oleic acid soybean germplasm developed contained a desirable fatty acid profile, and it was stable in two production environments. The presumed causative sequence polymorphisms in the FAD2-1B alleles were developed into highly efficient molecular markers for tracking the

  12. Mutant alleles of FAD2-1A and FAD2-1B combine to produce soybeans with the high oleic acid seed oil trait.

    Science.gov (United States)

    Pham, Anh-Tung; Lee, Jeong-Dong; Shannon, J Grover; Bilyeu, Kristin D

    2010-09-09

    The alteration of fatty acid profiles in soybean [Glycine max (L.) Merr.] to improve soybean oil quality is an important and evolving theme in soybean research to meet nutritional needs and industrial criteria in the modern market. Soybean oil with elevated oleic acid is desirable because this monounsaturated fatty acid improves the nutrition and oxidative stability of the oil. Commodity soybean oil typically contains 20% oleic acid and the target for high oleic acid soybean oil is approximately 80% of the oil; previous conventional plant breeding research to raise the oleic acid level to just 50-60% of the oil was hindered by the genetic complexity and environmental instability of the trait. The objective of this work was to create the high oleic acid trait in soybeans by identifying and combining mutations in two delta-twelve fatty acid desaturase genes, FAD2-1A and FAD2-1B. Three polymorphisms found in the FAD2-1B alleles of two soybean lines resulted in missense mutations. For each of the two soybean lines, there was one unique amino acid change within a highly conserved region of the protein. The mutant FAD2-1B alleles were associated with an increase in oleic acid levels, although the FAD2-1B mutant alleles alone were not capable of producing a high oleic acid phenotype. When existing FAD2-1A mutations were combined with the novel mutant FAD2-1B alleles, a high oleic acid phenotype was recovered only for those lines which were homozygous for both of the mutant alleles. We were able to produce conventional soybean lines with 80% oleic acid in the oil in two different ways, each requiring the contribution of only two genes. The high oleic acid soybean germplasm developed contained a desirable fatty acid profile, and it was stable in two production environments. The presumed causative sequence polymorphisms in the FAD2-1B alleles were developed into highly efficient molecular markers for tracking the mutant alleles. The resources described here for the creation

  13. A genetic screen in Myxococcus xanthus identifies mutants that uncouple outer membrane exchange from a downstream cellular response.

    Science.gov (United States)

    Dey, Arup; Wall, Daniel

    2014-12-01

    Upon physical contact with sibling cells, myxobacteria transiently fuse their outer membranes (OMs) and exchange OM proteins and lipids. From previous work, TraA and TraB were identified to be essential factors for OM exchange (OME) in donor and recipient cells. To define the genetic complexity of OME, we carried out a comprehensive forward genetic screen. The screen was based on the observation that Myxococcus xanthus nonmotile cells, by a Tra-dependent mechanism, block swarm expansion of motile cells when mixed. Thus, mutants defective in OME or a downstream responsive pathway were readily identified as escape flares from mixed inocula seeded on agar. This screen was surprisingly powerful, as we found >50 mutants defective in OME. Importantly, all of the mutations mapped to the traAB operon, suggesting that there may be few, if any, proteins besides TraA and TraB directly required for OME. We also found a second and phenotypically different class of mutants that exhibited wild-type OME but were defective in a responsive pathway. This pathway is postulated to control inner membrane homeostasis by covalently attaching amino acids to phospholipids. The identified proteins are homologous to the Staphylococcus aureus MprF protein, which is involved in membrane adaptation and antibiotic resistance. Interestingly, we also found that a small number of nonmotile cells were sufficient to block the swarming behavior of a large gliding-proficient population. This result suggests that an OME-derived signal could be amplified from a few nonmotile producers to act on many responder cells. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  14. Ascorbic acid deficiency activates cell death and disease resistance responses in Arabidopsis.

    Science.gov (United States)

    Pavet, Valeria; Olmos, Enrique; Kiddle, Guy; Mowla, Shaheen; Kumar, Sanjay; Antoniw, John; Alvarez, María E; Foyer, Christine H

    2005-11-01

    Programmed cell death, developmental senescence, and responses to pathogens are linked through complex genetic controls that are influenced by redox regulation. Here we show that the Arabidopsis (Arabidopsis thaliana) low vitamin C mutants, vtc1 and vtc2, which have between 10% and 25% of wild-type ascorbic acid, exhibit microlesions, express pathogenesis-related (PR) proteins, and have enhanced basal resistance against infections caused by Pseudomonas syringae. The mutants have a delayed senescence phenotype with smaller leaf cells than the wild type at maturity. The vtc leaves have more glutathione than the wild type, with higher ratios of reduced glutathione to glutathione disulfide. Expression of green fluorescence protein (GFP) fused to the nonexpressor of PR protein 1 (GFP-NPR1) was used to detect the presence of NPR1 in the nuclei of transformed plants. Fluorescence was observed in the nuclei of 6- to 8-week-old GFP-NPR1 vtc1 plants, but not in the nuclei of transformed GFP-NPR1 wild-type plants at any developmental stage. The absence of senescence-associated gene 12 (SAG12) mRNA at the time when constitutive cell death and basal resistance were detected confirms that elaboration of innate immune responses in vtc plants does not result from activation of early senescence. Moreover, H2O2-sensitive genes are not induced at the time of systemic acquired resistance execution. These results demonstrate that ascorbic acid abundance modifies the threshold for activation of plant innate defense responses via redox mechanisms that are independent of the natural senescence program.

  15. Biochemical characteristics of mutant lines of currant tomato

    International Nuclear Information System (INIS)

    Gorbatenko, I.Yu.; Khrustaleva, V.V.; Shcherbakov, V.K.

    1988-01-01

    The currant tomato is used in breeding for fruit quality. It contains up to 50 mg% ascorbic acid, a large quantity of sugar and 8-10% of dry matter. The weight of the fruit, however, does not exceed 1.2-1.5 g. The plants have long, spreading and very branchy stems. Gamma ray induced mutants of currant tomato were used, as initial material in breeding for fruit quality in varieties suitable for mechanized harvesting. The research was carried out mainly at the Department of Vegetable Growing Ukrainian Scientific Research Institute of Irrigation Farming. The regional variety Lebyazhinskij (suitable for mechanized harvesting) was adopted as the standard. Its fruits contain: 5.6% dry matter, 2.7% sugars, 0.543% titrated acidity, 26.6 mg/100 g ascorbic acid, 0.425 mg% carotene and 0.35% cellulose. The biochemical characteristics of the tomato mutants are shown. In terms of fruit dry matter, all mutants surpassed the standard. The acidity and the ascorbic acid content varied considerably. Most noteworthy in terms of carotene were the lines GP-5, GP-9 and GP-12. An important factor in the production of tomato paste is the fruit cellulose content. The lowest cellulose content is found in mutant GP-3. As shown, all of the mutants were early ripening. The mutants surpassed the standard in simultaneous fruit ripening. Mutant lines GP-3, GP-6, GP-9 and GP-12 will be used in the breeding programme for improving fruit quality of varieties suitable for mechanized harvesting

  16. Insights into regioselective metabolism of mefenamic acid by cytochrome P450 BM3 mutants through crystallography, docking, molecular dynamics, and free energy calculations

    DEFF Research Database (Denmark)

    Capoferri, Luigi; Leth, Rasmus; Ter Haar, Ernst

    2016-01-01

    of the protein mutant M11 was expressed, purified, and crystallized, and its X-ray structure was used as template for modeling. A multistep approach was used that combines molecular docking, molecular dynamics (MD) simulation, and binding free-energy calculations to address protein flexibility. In this way...... active-site mutations such as V87I were reported to invert regioselectivity in NSAID hydroxylation. In this work, we combine crystallography and molecular simulation to study the effect of single mutations on binding and regioselective metabolism of mefenamic acid by M11 mutants. The heme domain...... of mefenamic acid by M11 and its mutants by including protein flexibility and dynamics in free-energy computation. In addition, we could obtain structural insights into the change in regioselectivity of mefenamic acid hydroxylation due to single active-site mutations. Our findings confirm that use of MD...

  17. Sulfhydryl oxidation of mutants with cysteine in place of acidic residues in the lactose permease.

    Science.gov (United States)

    Voss, J; Sun, J; Venkatesan, P; Kaback, H R

    1998-06-02

    To examine further the role of charge-pair interactions in the structure and function of lactose permease, Asp237 (helix VII), Asp240 (helix VII), Glu126 (cytoplasmic loop IV/V), Glu269 (helix VIII), and Glu325 (helix X) were replaced individually with Cys in a functional mutant devoid of Cys residues. Each mutant was then oxidized with H2O2 in order to generate a sulfinic and/or sulfonic acid at these positions. Due to the isosteric relationship between aspartate and sulfinate, in particular, and the lower pKa of the sulfinic and sulfonic acid side chains, oxidized derivatives of Cys are useful probes for examining the role of carboxylates. Asp237-->Cys or Asp240-->Cys permease is inactive, as shown previously, but H2O2 oxidation restores activity to an extent similar to that observed when a negative charge is reintroduced by other means. Glu126-->Cys, Glu269-->Cys, or Glu325-->Cys permease is inactive, but oxidation does not restore active lactose transport. The data are consistent with previous observations indicating that Asp237 and Asp240 are not critical for active lactose transport, while Glu126, Glu269, and Glu325 are irreplaceable. Although Glu269-->Cys permease does not transport lactose, the oxidized mutant exhibits significant transport of beta,D-galactosylpyranosyl 1-thio-beta,D-galactopyranoside, a property observed with Glu269-->Asp permease. The observation supports the idea that an acidic residue at position 269 is important for substrate recognition. Finally, oxidized Glu325-->Cys permease catalyzes equilibrium exchange with an apparent pKa of about 6.5, more than a pH unit lower than that observed with Glu325-->Asp permease, thereby providing strong confirmatory evidence that a negative charge at position 325 determines the rate of translocation of the ternary complex between the permease, substrate, and H+.

  18. The plastidial retrograde signal methyl erythritol cyclopyrophosphate is a regulator of salicylic acid and jasmonic acid crosstalk

    Science.gov (United States)

    Lemos, Mark; Xiao, Yanmei; Bjornson, Marta; Wang, Jin-zheng; Hicks, Derrick; de Souza, Amancio; Wang, Chang-Quan; Yang, Panyu; Ma, Shisong; Dinesh-Kumar, Savithramma; Dehesh, Katayoon

    2016-01-01

    The exquisite harmony between hormones and their corresponding signaling pathways is central to prioritizing plant responses to simultaneous and/or successive environmental trepidations. The crosstalk between jasmonic acid (JA) and salicylic acid (SA) is an established effective mechanism that optimizes and tailors plant adaptive responses. However, the underlying regulatory modules of this crosstalk are largely unknown. Global transcriptomic analyses of mutant plants (ceh1) with elevated levels of the stress-induced plastidial retrograde signaling metabolite 2-C-methyl-D-erythritol cyclopyrophosphate (MEcPP) revealed robustly induced JA marker genes, expected to be suppressed by the presence of constitutively high SA levels in the mutant background. Analyses of a range of genotypes with varying SA and MEcPP levels established the selective role of MEcPP-mediated signal(s) in induction of JA-responsive genes in the presence of elevated SA. Metabolic profiling revealed the presence of high levels of the JA precursor 12-oxo-phytodienoic acid (OPDA), but near wild type levels of JA in the ceh1 mutant plants. Analyses of coronatine-insensitive 1 (coi1)/ceh1 double mutant plants confirmed that the MEcPP-mediated induction is JA receptor COI1 dependent, potentially through elevated OPDA. These findings identify MEcPP as a previously unrecognized central regulatory module that induces JA-responsive genes in the presence of high SA, thereby staging a multifaceted plant response within the environmental context. PMID:26733689

  19. Auxin-dependent microtubule responses and seedling development are affected in a rice mutant resistant to EPC

    International Nuclear Information System (INIS)

    Nick, P.; Yatou, O.; Furuya, M.; Lambert, A.M.

    1994-01-01

    Mutants in rice (Oryza sativa L. cv. japonica) were used to study the role of the cytoskeleton in signal-dependent morphogenesis. Mutants obtained by gamma ray irradiation were selected that failed to show inhibition of coleoptile elongation by the anti microtubular drug ethyl-N-phenylcarbamate (EPC). The mutation EPC-Resistant 31 (ER31), isolated from such a screen, caused lethality in putatively homozygous embryos. Heterozygotes exhibited drug resistance, impaired development of crown roots, and characteristic changes in the pattern of cell elongation: cell elongation was enhanced in mesocotyls and leaf sheaths, but inhibited in coleoptiles. The orientation of cortical microtubules changed correspondingly: for etiolated seedlings, compared with the wild-type, they were more transverse with respect to the long cell axis in mesocotyls and leaf sheaths, but more longitudinal in coleoptiles. In mutant coleoptiles, in contrast to wild-type, microtubules did not reorient in response to auxin, and their response to microtubule-eliminating and microtubule-stabilizing drugs was conspicuously reduced. In contrast, they responded normally to other stimuli such as gibberellins or red light. Auxin sensitivity as assayed by the dose-response for callus induction did not show any significant differences between wild-type and mutant. The mutant phenotype is interpreted in terms of an interrupted link between auxin-triggered signal transduction and microtubule reorientation. (author)

  20. Identification of a Transcription Factor Controlling pH-Dependent Organic Acid Response in Aspergillus niger

    DEFF Research Database (Denmark)

    Poulsen, Lars; Andersen, Mikael Rørdam; Lantz, Anna Eliasson

    2012-01-01

    exhibiting an oxalate overproducing phenotype were identified. The yield of oxalate was increased up to 158% compared to the wild type and the corresponding transcription factor was therefore entitled Oxalic Acid repression Factor, OafA. Detailed physiological characterization of one of the ΔoafA mutants......, compared to the wild type, showed that both strains produced substantial amounts of gluconic acid, but the mutant strain was more efficient in re-uptake of gluconic acid and converting it to oxalic acid, particularly at high pH (pH 5.0). Transcriptional profiles showed that 241 genes were differentially......Acid formation in Aspergillus niger is known to be subjected to tight regulation, and the acid production profiles are fine-tuned to respond to the ambient pH. Based on transcriptome data, putative trans-acting pH responding transcription factors were listed and through knock out studies, mutants...

  1. Stress Marker Signatures in Lesion Mimic Single and Double Mutants Identify a Crucial Leaf Age-Dependent Salicylic Acid Related Defense Signal.

    Science.gov (United States)

    Kaurilind, Eve; Brosché, Mikael

    2017-01-01

    Plants are exposed to abiotic and biotic stress conditions throughout their lifespans that activates various defense programs. Programmed cell death (PCD) is an extreme defense strategy the plant uses to manage unfavorable environments as well as during developmentally induced senescence. Here we investigated the role of leaf age on the regulation of defense gene expression in Arabidopsis thaliana. Two lesion mimic mutants with misregulated cell death, catalase2 (cat2) and defense no death1 (dnd1) were used together with several double mutants to dissect signaling pathways regulating defense gene expression associated with cell death and leaf age. PCD marker genes showed leaf age dependent expression, with the highest expression in old leaves. The salicylic acid (SA) biosynthesis mutant salicylic acid induction deficient2 (sid2) had reduced expression of PCD marker genes in the cat2 sid2 double mutant demonstrating the importance of SA biosynthesis in regulation of defense gene expression. While the auxin- and jasmonic acid (JA)- insensitive auxin resistant1 (axr1) double mutant cat2 axr1 also led to decreased expression of PCD markers; the expression of several marker genes for SA signaling (ISOCHORISMATE SYNTHASE 1, PR1 and PR2) were additionally decreased in cat2 axr1 compared to cat2. The reduced expression of these SA markers genes in cat2 axr1 implicates AXR1 as a regulator of SA signaling in addition to its known role in auxin and JA signaling. Overall, the current study reinforces the important role of SA signaling in regulation of leaf age-related transcript signatures.

  2. Early transcriptional responses of internalization defective Brucella abortus mutants in professional phagocytes, RAW 264.7.

    Science.gov (United States)

    Cha, Seung Bin; Lee, Won Jung; Shin, Min Kyoung; Jung, Myung Hwan; Shin, Seung Won; Yoo, An Na; Kim, Jong Wan; Yoo, Han Sang

    2013-06-27

    Brucella abortus is an intracellular zoonotic pathogen which causes undulant fever, endocarditis, arthritis and osteomyelitis in human and abortion and infertility in cattle. This bacterium is able to invade and replicate in host macrophage instead of getting removed by this defense mechanism. Therefore, understanding the interaction between virulence of the bacteria and the host cell is important to control brucellosis. Previously, we generated internalization defective mutants and analyzed the envelope proteins. The present study was undertaken to evaluate the changes in early transcriptional responses between wild type and internalization defective mutants infected mouse macrophage, RAW 264.7. Both of the wild type and mutant infected macrophages showed increased expression levels in proinflammatory cytokines, chemokines, apoptosis and G-protein coupled receptors (Gpr84, Gpr109a and Adora2b) while the genes related with small GTPase which mediate intracellular trafficking was decreased. Moreover, cytohesin 1 interacting protein (Cytip) and genes related to ubiquitination (Arrdc3 and Fbxo21) were down-regulated, suggesting the survival strategy of this bacterium. However, we could not detect any significant changes in the mutant infected groups compared to the wild type infected group. In summary, it was very difficult to clarify the alterations in host cellular transcription in response to infection with internalization defective mutants. However, we found several novel gene changes related to the GPCR system, ubiquitin-proteosome system, and growth arrest and DNA damages in response to B. abortus infection. These findings may contribute to a better understanding of the molecular mechanisms underlying host-pathogen interactions and need to be studied further.

  3. Novel mutations in β-tubulin gene in Trichoderma harzianum mutants resistant to methyl benzimidazol-2-yl carbamate.

    Science.gov (United States)

    Li, M; Zhang, H Y; Liang, B

    2013-01-01

    Twelve-low resistant (LR) mutants of Trichoderma harzianum with the capability of grow fast at 0.8 μg/mL methyl benzimidazol-2-yl carbamate (MBC) were obtained using UV mutagenesis. MR and HR mutants which could grow fast at 10 and 100 μg/mL MBC, respectively, were isolated by step-up selection protocols in which UV-treated mutants were induced and mycelial sector screening was made in plates with growth medium. Subsequently, β-tubulin genes of 14 mutants were cloned to describe-the molecular lesion likely to be responsible-for MBC resistance. Comparison of the β-tubulin sequences of the mutant and sensitive strains of T. harzianum revealed 2 new MBC-binding sites differed from those in other plant pathogens. A single mutation at-amino acid 168, having Phe (TTC) instead of Ser (TCC)', was demonstrated for the HR mutant; a double mutation in amino acid 13 resulting in the substitution of Gly (GGC) by Val (GTG) was observed in β-tubulin gene of MR mutant. On the other hand, no substitutions were identified in the β-tubulin gene and its 5'-flanking regions in 12 LR mutants of T. harzianum.

  4. Abscisic Acid–Responsive Guard Cell Metabolomes of Arabidopsis Wild-Type and gpa1 G-Protein Mutants[C][W

    Science.gov (United States)

    Jin, Xiaofen; Wang, Rui-Sheng; Zhu, Mengmeng; Jeon, Byeong Wook; Albert, Reka; Chen, Sixue; Assmann, Sarah M.

    2013-01-01

    Individual metabolites have been implicated in abscisic acid (ABA) signaling in guard cells, but a metabolite profile of this specialized cell type is lacking. We used liquid chromatography–multiple reaction monitoring mass spectrometry for targeted analysis of 85 signaling-related metabolites in Arabidopsis thaliana guard cell protoplasts over a time course of ABA treatment. The analysis utilized ∼350 million guard cell protoplasts from ∼30,000 plants of the Arabidopsis Columbia accession (Col) wild type and the heterotrimeric G-protein α subunit mutant, gpa1, which has ABA-hyposensitive stomata. These metabolomes revealed coordinated regulation of signaling metabolites in unrelated biochemical pathways. Metabolites clustered into different temporal modules in Col versus gpa1, with fewer metabolites showing ABA-altered profiles in gpa1. Ca2+-mobilizing agents sphingosine-1-phosphate and cyclic adenosine diphosphate ribose exhibited weaker ABA-stimulated increases in gpa1. Hormone metabolites were responsive to ABA, with generally greater responsiveness in Col than in gpa1. Most hormones also showed different ABA responses in guard cell versus mesophyll cell metabolomes. These findings suggest that ABA functions upstream to regulate other hormones, and are also consistent with G proteins modulating multiple hormonal signaling pathways. In particular, indole-3-acetic acid levels declined after ABA treatment in Col but not gpa1 guard cells. Consistent with this observation, the auxin antagonist α-(phenyl ethyl-2-one)-indole-3-acetic acid enhanced ABA-regulated stomatal movement and restored partial ABA sensitivity to gpa1. PMID:24368793

  5. Conserved role of unc-79 in ethanol responses in lightweight mutant mice.

    Directory of Open Access Journals (Sweden)

    David J Speca

    2010-08-01

    Full Text Available The mechanisms by which ethanol and inhaled anesthetics influence the nervous system are poorly understood. Here we describe the positional cloning and characterization of a new mouse mutation isolated in an N-ethyl-N-nitrosourea (ENU forward mutagenesis screen for animals with enhanced locomotor activity. This allele, Lightweight (Lwt, disrupts the homolog of the Caenorhabditis elegans (C. elegans unc-79 gene. While Lwt/Lwt homozygotes are perinatal lethal, Lightweight heterozygotes are dramatically hypersensitive to acute ethanol exposure. Experiments in C. elegans demonstrate a conserved hypersensitivity to ethanol in unc-79 mutants and extend this observation to the related unc-80 mutant and nca-1;nca-2 double mutants. Lightweight heterozygotes also exhibit an altered response to the anesthetic isoflurane, reminiscent of unc-79 invertebrate mutant phenotypes. Consistent with our initial mapping results, Lightweight heterozygotes are mildly hyperactive when exposed to a novel environment and are smaller than wild-type animals. In addition, Lightweight heterozygotes exhibit increased food consumption yet have a leaner body composition. Interestingly, Lightweight heterozygotes voluntarily consume more ethanol than wild-type littermates. The acute hypersensitivity to and increased voluntary consumption of ethanol observed in Lightweight heterozygous mice in combination with the observed hypersensitivity to ethanol in C. elegans unc-79, unc-80, and nca-1;nca-2 double mutants suggests a novel conserved pathway that might influence alcohol-related behaviors in humans.

  6. Nature of mutants induced by ionizing radiation in cultured hamster cells. II. Antigenic response and reverse mutation of HPRT-deficient mutants induced by. gamma. -rays or ethyl methanesulphonate

    Energy Technology Data Exchange (ETDEWEB)

    Brown, R; Stretch, A; Thacker, J

    1986-04-01

    A large series of independent mutants deficient in HPRT enzyme activity, isolated from V79-4 hamster cells, were assessed for properties which reflect the nature of the genetic changes induced. A total of 88 mutants were screened, 43 isolated from ..gamma..-ray-treated cultures and 45 induced by ethyl methanesulphonate (EMS). Firstly, each mutant was assayed for the presence of protein with the antigenic response of HPRT. In a competitive inhibition assay, 31% of EMS-induced mutants were CRM-positive compared to 7% of the ..gamma..-ray series. Secondly, each mutant was tested for ability to revert to HPRT proficiency. All except 2 of the EMS-induced mutants reverted with ethyl nitrosourea ENU, and many reverted spontaneously, under the given conditions. However reversion was not detected in about 80% of ..gamma..-ray-induced mutants, suggesting that the types of forward mutation caused by ionizing radiation differ qualitatively from those caused by EMS. (Auth.). 30 refs.; 6 figs.; 2 tabs.

  7. Abscisic acid deficiency increases defence responses against Myzus persicae in Arabidopsis.

    Science.gov (United States)

    Hillwig, Melissa S; Chiozza, Mariana; Casteel, Clare L; Lau, Siau Ting; Hohenstein, Jessica; Hernández, Enrique; Jander, Georg; MacIntosh, Gustavo C

    2016-02-01

    Comparison of Arabidopsis thaliana (Arabidopsis) gene expression induced by Myzus persicae (green peach aphid) feeding, aphid saliva infiltration and abscisic acid (ABA) treatment showed a significant positive correlation. In particular, ABA-regulated genes are over-represented among genes that are induced by M. persicae saliva infiltration into Arabidopsis leaves. This suggests that the induction of ABA-related gene expression could be an important component of the Arabidopsis-aphid interaction. Consistent with this hypothesis, M. persicae populations induced ABA production in wild-type plants. Furthermore, aphid populations were smaller on Arabidopsis aba1-1 mutants, which cannot synthesize ABA, and showed a significant preference for wild-type plants compared with the mutant. Total free amino acids, which play an important role in aphid nutrition, were not altered in the aba1-1 mutant line, but the levels of isoleucine (Ile) and tryptophan (Trp) were differentially affected by aphids in wild-type and mutant plants. Recently, indole glucosinolates have been shown to promote aphid resistance in Arabidopsis. In this study, 4-methoxyindol-3-ylmethylglucosinolate was more abundant in the aba1-1 mutant than in wild-type Arabidopsis, suggesting that the induction of ABA signals that decrease the accumulation of defence compounds may be beneficial for aphids. © 2015 BSPP AND JOHN WILEY & SONS LTD.

  8. The plastidial retrograde signal methyl erythritol cyclopyrophosphate is a regulator of salicylic acid and jasmonic acid crosstalk.

    Science.gov (United States)

    Lemos, Mark; Xiao, Yanmei; Bjornson, Marta; Wang, Jin-Zheng; Hicks, Derrick; Souza, Amancio de; Wang, Chang-Quan; Yang, Panyu; Ma, Shisong; Dinesh-Kumar, Savithramma; Dehesh, Katayoon

    2016-03-01

    The exquisite harmony between hormones and their corresponding signaling pathways is central to prioritizing plant responses to simultaneous and/or successive environmental trepidations. The crosstalk between jasmonic acid (JA) and salicylic acid (SA) is an established effective mechanism that optimizes and tailors plant adaptive responses. However, the underlying regulatory modules of this crosstalk are largely unknown. Global transcriptomic analyses of mutant plants (ceh1) with elevated levels of the stress-induced plastidial retrograde signaling metabolite 2-C-methyl-D-erythritol cyclopyrophosphate (MEcPP) revealed robustly induced JA marker genes, expected to be suppressed by the presence of constitutively high SA levels in the mutant background. Analyses of a range of genotypes with varying SA and MEcPP levels established the selective role of MEcPP-mediated signal(s) in induction of JA-responsive genes in the presence of elevated SA. Metabolic profiling revealed the presence of high levels of the JA precursor 12-oxo-phytodienoic acid (OPDA), but near wild type levels of JA in the ceh1 mutant plants. Analyses of coronatine-insensitive 1 (coi1)/ceh1 double mutant plants confirmed that the MEcPP-mediated induction is JA receptor COI1 dependent, potentially through elevated OPDA. These findings identify MEcPP as a previously unrecognized central regulatory module that induces JA-responsive genes in the presence of high SA, thereby staging a multifaceted plant response within the environmental context. © The Author 2016. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  9. Mutants of GABA transaminase (POP2 suppress the severe phenotype of succinic semialdehyde dehydrogenase (ssadh mutants in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Frank Ludewig

    Full Text Available BACKGROUND: The gamma-aminubutyrate (GABA shunt bypasses two steps of the tricarboxylic acid cycle, and is present in both prokaryotes and eukaryotes. In plants, the pathway is composed of the calcium/calmodulin-regulated cytosolic enzyme glutamate decarboxylase (GAD, the mitochondrial enzymes GABA transaminase (GABA-T; POP2 and succinic semialdehyde dehydrogenase (SSADH. We have previously shown that compromising the function of the GABA-shunt, by disrupting the SSADH gene of Arabidopsis, causes enhanced accumulation of reactive oxygen intermediates (ROIs and cell death in response to light and heat stress. However, to date, genetic investigations of the relationships between enzymes of the GABA shunt have not been reported. PRINCIPAL FINDINGS: To elucidate the role of succinic semialdehyde (SSA, gamma-hydroxybutyrate (GHB and GABA in the accumulation of ROIs, we combined two genetic approaches to suppress the severe phenotype of ssadh mutants. Analysis of double pop2 ssadh mutants revealed that pop2 is epistatic to ssadh. Moreover, we isolated EMS-generated mutants suppressing the phenotype of ssadh revealing two new pop2 alleles. By measuring thermoluminescence at high temperature, the peroxide contents of ssadh and pop2 mutants were evaluated, showing that only ssadh plants accumulate peroxides. In addition, pop2 ssadh seedlings are more sensitive to exogenous SSA or GHB relative to wild type, because GHB and/or SSA accumulate in these plants. SIGNIFICANCE: We conclude that the lack of supply of succinate and NADH to the TCA cycle is not responsible for the oxidative stress and growth retardations of ssadh mutants. Rather, we suggest that the accumulation of SSA, GHB, or both, produced downstream of the GABA-T transamination step, is toxic to the plants, resulting in high ROI levels and impaired development.

  10. Semi-dwarf mutants in triticale and wheat breeding

    International Nuclear Information System (INIS)

    Driscoll, C.J.

    1984-01-01

    The triticale lines Beagle and DR-IRA have been subjected to ionizing irradiation and chemical mutagenesis in order to produce semi-dwarf mutants. Beagle is 100 cm tall and DR-IRA 80 cm under average field conditions. A bulk then pedigree method is currently represented by 158 single plots of M 6 (or in some cases M 7 ) mutants that are from 5 to 35 cm shorter than the control variety. The shortest mutants are 65 cm in height. Forty of these mutants are also earlier flowering than the control varieties. Replicated yield testing will be conducted on confirmed mutants in 1983. Response to gibberellic acid of these mutants will also be determined. The Cornerstone male-sterility mutant (ms1c) on chromosome arm 4Aα has been combined with the GA-insensitive/reduced height gene Gai/Rht1 which is also on chromosome arm 4Aα. The ms1c mutant has also been combined with Gai/Rht2 on chromosome 4D and with both Gai/Rht1 and Gai/Rht2. The combination ms1c and Gai/Rht1 has been chosen as the basis of a composite cross. Thirteen varieties were tested with GA 3 and seven (Warigal, Aroona, Oxley, Banks, Avocet, Matipo and Toquifen) which contain Gai/Rht1 were crossed with ms1c Gai/Rht1 and entered into an interpollinating F 2 . The entire composite is homozygous for this semi-dwarf allele and selection will be practiced for increased height on a GA-insensitive background. (author)

  11. Ethylene signaling renders the jasmonate response of Arabidopsis insensitive to future suppression by salicylic Acid.

    Science.gov (United States)

    Leon-Reyes, Antonio; Du, Yujuan; Koornneef, Annemart; Proietti, Silvia; Körbes, Ana P; Memelink, Johan; Pieterse, Corné M J; Ritsema, Tita

    2010-02-01

    Cross-talk between jasmonate (JA), ethylene (ET), and Salicylic acid (SA) signaling is thought to operate as a mechanism to fine-tune induced defenses that are activated in response to multiple attackers. Here, 43 Arabidopsis genotypes impaired in hormone signaling or defense-related processes were screened for their ability to express SA-mediated suppression of JA-responsive gene expression. Mutant cev1, which displays constitutive expression of JA and ET responses, appeared to be insensitive to SA-mediated suppression of the JA-responsive marker genes PDF1.2 and VSP2. Accordingly, strong activation of JA and ET responses by the necrotrophic pathogens Botrytis cinerea and Alternaria brassicicola prior to SA treatment counteracted the ability of SA to suppress the JA response. Pharmacological assays, mutant analysis, and studies with the ET-signaling inhibitor 1-methylcyclopropene revealed that ET signaling renders the JA response insensitive to subsequent suppression by SA. The APETALA2/ETHYLENE RESPONSE FACTOR transcription factor ORA59, which regulates JA/ET-responsive genes such as PDF1.2, emerged as a potential mediator in this process. Collectively, our results point to a model in which simultaneous induction of the JA and ET pathway renders the plant insensitive to future SA-mediated suppression of JA-dependent defenses, which may prioritize the JA/ET pathway over the SA pathway during multi-attacker interactions.

  12. Hormonal regulation of seed development and germination in tomato : studies on abscisic acid- and gibberellin-deficient mutants

    NARCIS (Netherlands)

    Groot, S.P.C.

    1987-01-01

    The role of endogenous gibberellins (GAs) and abscisic acid (ABA) in seed development and germination of tomato, was studied with the use of GA- and/or ABA-deficient mutants.

    GAs are indispensable for the development of fertile flowers. Fertility of GA-deficient flowers is restored

  13. Transcriptome Comparative Profiling of Barley eibi1 Mutant Reveals Pleiotropic Effects of HvABCG31 Gene on Cuticle Biogenesis and Stress Responsive Pathways

    Directory of Open Access Journals (Sweden)

    Eviatar Nevo

    2013-10-01

    Full Text Available Wild barley eibi1 mutant with HvABCG31 gene mutation has low capacity to retain leaf water, a phenotype associated with reduced cutin deposition and a thin cuticle. To better understand how such a mutant plant survives, we performed a genome-wide gene expression analysis. The leaf transcriptomes between the near-isogenic lines eibi1 and the wild type were compared using the 22-k Barley1 Affymetrix microarray. We found that the pleiotropic effect of the single gene HvABCG31 mutation was linked to the co-regulation of metabolic processes and stress-related system. The cuticle development involved cytochrome P450 family members and fatty acid metabolism pathways were significantly up-regulated by the HvABCG31 mutation, which might be anticipated to reduce the levels of cutin monomers or wax and display conspicuous cuticle defects. The candidate genes for responses to stress were induced by eibi1 mutant through activating the jasmonate pathway. The down-regulation of co-expressed enzyme genes responsible for DNA methylation and histone deacetylation also suggested that HvABCG31 mutation may affect the epigenetic regulation for barley development. Comparison of transcriptomic profiling of barley under biotic and abiotic stresses revealed that the functions of HvABCG31 gene to high-water loss rate might be different from other osmotic stresses of gene mutations in barley. The transcriptional profiling of the HvABCG31 mutation provided candidate genes for further investigation of the physiological and developmental changes caused by the mutant.

  14. Rapid identification of lettuce seed germination mutants by bulked segregant analysis and whole genome sequencing.

    Science.gov (United States)

    Huo, Heqiang; Henry, Isabelle M; Coppoolse, Eric R; Verhoef-Post, Miriam; Schut, Johan W; de Rooij, Han; Vogelaar, Aat; Joosen, Ronny V L; Woudenberg, Leo; Comai, Luca; Bradford, Kent J

    2016-11-01

    Lettuce (Lactuca sativa) seeds exhibit thermoinhibition, or failure to complete germination when imbibed at warm temperatures. Chemical mutagenesis was employed to develop lettuce lines that exhibit germination thermotolerance. Two independent thermotolerant lettuce seed mutant lines, TG01 and TG10, were generated through ethyl methanesulfonate mutagenesis. Genetic and physiological analyses indicated that these two mutations were allelic and recessive. To identify the causal gene(s), we applied bulked segregant analysis by whole genome sequencing. For each mutant, bulked DNA samples of segregating thermotolerant (mutant) seeds were sequenced and analyzed for homozygous single-nucleotide polymorphisms. Two independent candidate mutations were identified at different physical positions in the zeaxanthin epoxidase gene (ABSCISIC ACID DEFICIENT 1/ZEAXANTHIN EPOXIDASE, or ABA1/ZEP) in TG01 and TG10. The mutation in TG01 caused an amino acid replacement, whereas the mutation in TG10 resulted in alternative mRNA splicing. Endogenous abscisic acid contents were reduced in both mutants, and expression of the ABA1 gene from wild-type lettuce under its own promoter fully complemented the TG01 mutant. Conventional genetic mapping confirmed that the causal mutations were located near the ZEP/ABA1 gene, but the bulked segregant whole genome sequencing approach more efficiently identified the specific gene responsible for the phenotype. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

  15. Nutrient value protein of mutant J3 of agaricus blazei by irradiation

    International Nuclear Information System (INIS)

    Jiang Zhihe; Lin Yong; Xiao Shuxia

    2003-01-01

    The studies of nutrient value protein of mutant J 3 of Agaricus blazei were carried out. The results showed that mutant J 3 had 9 kinds of essential amino acids, the compositions of essential amino acids is 46.0% of total amino acids. The amino acid score is 94.5, the essential amino acid index is 92.4, biological value is 89.0, the ratio coefficient of amino acids is 72.3, the nutrient value is 32.5. The chemical score is 74.18. These results proved that the protein of mutant J 3 had high nutrient value

  16. Arabidopsis cysteine-rich receptor-like kinase 45 functions in the responses to abscisic acid and abiotic stresses

    KAUST Repository

    Zhang, Xiujuan

    2013-06-01

    The phytohormone abscisic acid (ABA) regulates seed germination, plant growth and development, and response to abiotic stresses such as drought and salt stresses. Receptor-like kinases are well known signaling components that mediate plant responses to developmental and environmental stimuli. Here, we characterized the biological function of an ABA and stress-inducible cysteine-rich receptor-like protein kinase, CRK45, in ABA signaling in Arabidopsis thaliana. The crk45 mutant was less sensitive to ABA than the wild type during seed germination and early seedling development, whereas CRK45 overexpression plants were more sensitive to ABA compared to the wild type. Furthermore, overexpression of CRK45 led to hypersensitivity to salt and glucose inhibition of seed germination, whereas the crk45 mutant showed the opposite phenotypes. In addition, CRK45 overexpression plants had enhanced tolerance to drought. Gene expression analyses revealed that the expression of representative stress-responsive genes was significantly enhanced in CRK45 overexpression plants in response to salt stress. ABA biosynthetic genes such as NCED3,. 22NCED3, 9-Cis-Epoxycarotenoid Dioxygenase 3.NCED5,. 33NCED5, 9-Cis-Epoxycarotenoid Dioxygenase 5.ABA2,. 44ABA2, Abscisic Acid Deficient 2. and AAO355AAO3, Abscisic Aldehyde Oxidase 3. were also constitutively elevated in the CRK45 overexpression plants. We concluded that CRK45 plays an important role in ABA signaling that regulates Arabidopsis seeds germination, early seedling development and abiotic stresses response, by positively regulating ABA responses in these processes. © 2013 Elsevier Masson SAS.

  17. Sinapic acid or its derivatives interfere with abscisic acid homeostasis during Arabidopsis thaliana seed germination.

    Science.gov (United States)

    Bi, Baodi; Tang, Jingliang; Han, Shuang; Guo, Jinggong; Miao, Yuchen

    2017-06-06

    Sinapic acid and its esters have broad functions in different stages of seed germination and plant development and are thought to play a role in protecting against ultraviolet irradiation. To better understand the interactions between sinapic acid esters and seed germination processes in response to various stresses, we analyzed the role of the plant hormone abscisic acid (ABA) in the regulation of sinapic acid esters involved in seed germination and early seedling growth. We found that exogenous sinapic acid promotes seed germination in a dose-dependent manner in Arabidopsis thaliana. High-performance liquid chromatography mass spectrometry analysis showed that exogenous sinapic acid increased the sinapoylcholine content of imbibed seeds. Furthermore, sinapic acid affected ABA catabolism, resulting in reduced ABA levels and increased levels of the ABA-glucose ester. Using mutants deficient in the synthesis of sinapate esters, we showed that the germination of mutant sinapoylglucose accumulator 2 (sng2) and bright trichomes 1 (brt1) seeds was more sensitive to ABA than the wild-type. Moreover, Arabidopsis mutants deficient in either abscisic acid deficient 2 (ABA2) or abscisic acid insensitive 3 (ABI3) displayed increased expression of the sinapoylglucose:choline sinapoyltransferase (SCT) and sinapoylcholine esterase (SCE) genes with sinapic acid treatment. This treatment also affected the accumulation of sinapoylcholine and free choline during seed germination. We demonstrated that sinapoylcholine, which constitutes the major phenolic component in seeds among various minor sinapate esters, affected ABA homeostasis during seed germination and early seedling growth in Arabidopsis. Our findings provide insights into the role of sinapic acid and its esters in regulating ABA-mediated inhibition of Arabidopsis seed germination in response to drought stress.

  18. Flavonoids and Auxin Transport Inhibitors Rescue Symbiotic Nodulation in the Medicago truncatula Cytokinin Perception Mutant cre1

    Science.gov (United States)

    Ng, Jason Liang Pin; Hassan, Samira; Truong, Thy T.; Hocart, Charles H.; Laffont, Carole; Frugier, Florian; Mathesius, Ulrike

    2015-01-01

    Initiation of symbiotic nodules in legumes requires cytokinin signaling, but its mechanism of action is largely unknown. Here, we tested whether the failure to initiate nodules in the Medicago truncatula cytokinin perception mutant cre1 (cytokinin response1) is due to its altered ability to regulate auxin transport, auxin accumulation, and induction of flavonoids. We found that in the cre1 mutant, symbiotic rhizobia cannot locally alter acro- and basipetal auxin transport during nodule initiation and that these mutants show reduced auxin (indole-3-acetic acid) accumulation and auxin responses compared with the wild type. Quantification of flavonoids, which can act as endogenous auxin transport inhibitors, showed a deficiency in the induction of free naringenin, isoliquiritigenin, quercetin, and hesperetin in cre1 roots compared with wild-type roots 24 h after inoculation with rhizobia. Coinoculation of roots with rhizobia and the flavonoids naringenin, isoliquiritigenin, and kaempferol, or with the synthetic auxin transport inhibitor 2,3,5,-triiodobenzoic acid, rescued nodulation efficiency in cre1 mutants and allowed auxin transport control in response to rhizobia. Our results suggest that CRE1-dependent cytokinin signaling leads to nodule initiation through the regulation of flavonoid accumulation required for local alteration of polar auxin transport and subsequent auxin accumulation in cortical cells during the early stages of nodulation. PMID:26253705

  19. Strain improvement and metabolic flux analysis in the wild-type and a mutant Lactobacillus lactis strain for L(+)-lactic acid production.

    Science.gov (United States)

    Bai, Dong-Mei; Zhao, Xue-Ming; Li, Xin-Gang; Xu, Shi-Min

    2004-12-20

    The effects of initial glucose concentration and calcium lactate concentration on the lactic acid production by the parent strain, Lactobacillus lactis BME5-18, were studied. The results of the experiments indicated that glucose and lactate repressed the cell growth and the lactic acid production by Lactobacillus lactis BME5-18. A L(+)-lactic acid overproducing strain, Lactobacillus lactis BME5-18M, was screened by mutagenizing the parent strain with ultraviolet (UV) light irradiation and selecting the high glucose and lactate calcium concentration repression resistant mutant. Starting with a concentration of 100g L(-1) glucose, the mutant produced 98.6 g L(-1) lactic acid after 60 h in flasks, 73.9% higher than that of the parent strain. The L(+)-lactic acid purity was 98.1% by weight based on the amount of total lactic acid. The culture of the parent strain could not be analyzed well by conventional metabolic flux analysis techniques, since some pyruvate were accumulated intracellularly. Therefore, a revised flux analysis method was proposed by introducing intracellular pyruvate pool. Further studies demonstrate that there is a high level of NADH oxidase activity (12.11 mmol mg(-1) min(-1)) in the parent strain. The molecular mechanisms of the strain improvement were proposed, i.e., the high level of NADH oxidase activity was eliminated and the uptake rate of glucose was increased from 82.1 C-mmol (g DW h)(-1) to 98.9 C-mmol (g DW h)(-1) by mutagenizing the parent strain with UV, and therefore the mutant strain converts mostly pyruvate to lactic acid with a higher productivity (1.76 g L(-1) h(-1)) than the parent strain (0.95 g L(-1) h(-1)).

  20. Chinese hamster ovary cell mutants defective in heparan sulfate biosynthesis

    International Nuclear Information System (INIS)

    Bame, K.J.; Kiser, C.S.; Esko, J.D.

    1987-01-01

    The authors have isolated Chinese hamster ovary cell mutants defective in proteoglycan synthesis by radiographic screening for cells unable to incorporate 35 SO 4 into acid-precipitable material. Some mutants did not incorporate 35 SO 4 into acid-precipitable material, whereas others incorporated about 3-fold less radioactivity. HPLC anion exchange chromatographic analysis of radiolabelled glycosaminoglycans isolated from these mutants revealed many are defective in heparan sulfate biosynthesis. Mutants 803 and 677 do not synthesize heparan sulfate, although they produce chondroitin sulfate: strain 803 makes chondroitin sulfate normally, whereas 677 overaccumulates chondroitin sulfate by a factor of three. These mutants fall into the same complementation group, suggesting that the mutations are allelic. A second group of heparan sulfate biosynthetic mutants, consisting of cell lines 625, 668 and 679, produce undersulfated heparan sulfate and normal chondroitin sulfate. Treatment of the chains with nitrous acid should determine the position of the sulfate groups along the chain. These mutants may define a complementation group that is defective in the enzymes which modify the heparan sulfate chain. To increase the authors repertoire of heparan sulfate mutants, they are presently developing an in situ enzyme assay to screen colonies replica plated on filter discs for sulfotransferase defects

  1. Deoxyribonucleic acid repair in Escherichia coli mutants deficient in the 5'----3' exonuclease activity of deoxyribonucleic acid polymerase I and exonuclease VII

    International Nuclear Information System (INIS)

    Chase, J.W.; Masker, W.E.

    1977-01-01

    A series of Escherichia coli strains deficient in the 5'----3' exonuclease activity associated with deoxyribonucleic acid (DNA) polymerase I (exonuclease VI) and exonuclease VII has been constructed. Both of these enzymes are capable of pyrimidine dimer excision in vitro. These strains were examined for conditional lethality, sensitivity to ultraviolet (UV) and X-irradiation, postirradiation DNA degradation, and ability to excise pyrimidine dimers. It was found that strains deficient in both exonuclease VI (polAex-) and exonuclease VII (xseA-) are significantly reduced in their ability to survive incubation at elevated temperature (43 degrees C) beyond the reduction previously observed for the polAex single mutants. The UV and X-ray sensitivity of the exonuclease VI-deficient strains was not increased by the addition of the xseA7 mutation. Mutants deficient in both enzymes are about as efficient as wild-type strains at excising dimers produced by up to 40 J/m2 UV. At higher doses strains containing only polAex- mutations show reduced ability to excise dimers; however, the interpretation of dimer excision data at these doses is complicated by extreme postirradiation DNA degradation in these strains. The additional deficiency in the polAex xseA7 double-mutant strains has no significant effect on either postirradiation DNA degradation or the apparent deficiency in dimer excision at high UV doses observed in polAex single mutants

  2. Deficiency of PdxR in Streptococcus mutans affects vitamin B6 metabolism, acid tolerance response and biofilm formation.

    Science.gov (United States)

    Liao, S; Bitoun, J P; Nguyen, A H; Bozner, D; Yao, X; Wen, Z T

    2015-08-01

    Streptococcus mutans, a key etiological agent of the human dental caries, lives primarily on the tooth surface in tenacious biofilms. The SMU864 locus, designated pdxR, is predicted to encode a member of the novel MocR/GabR family proteins, which are featured with a winged helix DNA-binding N-terminal domain and a C-terminal domain highly homologous to the pyridoxal phosphate-dependent aspartate aminotransferases. A pdxR-deficient mutant, TW296, was constructed using allelic exchange. PdxR deficiency in S. mutans had little effect on cell morphology and growth when grown in brain heart infusion. However, when compared with its parent strain, UA159, the PdxR-deficient mutant displayed major defects in acid tolerance response and formed significantly fewer biofilms (P mutans is known to require vitamin B6 to grow in defined medium, B6 vitamers, especially pyridoxal, were strongly inhibitory at millimolar concentrations, against S. mutans growth and biofilm formation. Our results suggest that PdxR in S. mutans plays an important role in regulation of vitamin B6 metabolism, acid tolerance response and biofilm formation. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  3. Responsiveness to acidity via metal ion regulators mediates virulence in the gastric pathogen Helicobacter pylori.

    Science.gov (United States)

    Bury-Moné, Stéphanie; Thiberge, Jean-Michel; Contreras, Monica; Maitournam, Aboubakar; Labigne, Agnès; De Reuse, Hilde

    2004-07-01

    The virulence of pathogenic bacteria is dependent on their adaptation to and survival in the stressful conditions encountered in their hosts. Helicobacter pylori exclusively colonizes the acid stomach of primates, making it an ideal study model. Little is known about how H. pylori responds to the moderately acidic conditions encountered at its colonization site, the gastric mucus layer. Thus, we compared gene expression profiles of H. pylori 26695 grown at neutral and acidic pH, and validated the data for a selection of genes by real-time polymerase chain reaction, dot-blots or enzymatic assays. During growth in acidic conditions, 56 genes were upregulated and 45 genes downregulated. We found that acidity is a signal modulating the expression of several virulence factors. Regulation of genes related to metal ion homeostasis suggests protective mechanisms involving diminished transport and enhanced storage. Genes encoding subunits of the F0F1 ATPase and of a newly identified Na+/H+ antiporter (NhaC-HP0946) were downregulated, revealing that this bacterium uses original mechanisms to control proton entry. Five of the upregulated genes encoded proteins controlling intracellular ammonia synthesis, including urease, amidase and formamidase, underlining the major role of this buffering compound in the protection against acidity in H. pylori. Regulatory networks and transcriptome analysis as well as enzymatic assays implicated two metal-responsive transcriptional regulators (NikR and Fur) and an essential two-component response regulator (HP0166, OmpR-like) as effectors of the H. pylori acid response. Finally, a nikR-fur mutant is attenuated in the mouse model, emphasizing the link between response to acidity, metal metabolism and virulence in this gastric pathogen.

  4. Chaperone protein HYPK interacts with the first 17 amino acid region of Huntingtin and modulates mutant HTT-mediated aggregation and cytotoxicity

    Energy Technology Data Exchange (ETDEWEB)

    Choudhury, Kamalika Roy [Crystallography and Molecular Biology Division, Saha Institute of Nuclear Physics, 1/AF Bidhannagar, Kolkata 700064 (India); Centre for Neuroscience, Indian Institute of Science, Bangalore 560012 (India); Bhattacharyya, Nitai P., E-mail: nitai_sinp@yahoo.com [Biomedical Genomics Centre, PG Polyclinic Building, 5, Suburbun Hospital Road, Kolkata 700020 (India)

    2015-01-02

    Highlights: • HYPK reduces mutant HTT-mediated aggregate formation and cytotoxicity. • Interaction of HYPK with HTT requires N-terminal 17 amino acid of HTT (HTT-N17). • Deletion of HTT-N17 leads to SDS-soluble, smaller, nuclear aggregates. • These smaller aggregates do not associate with HYPK and are more cytotoxic. • Maybe, interaction of HYPK with amphipathic HTT-N17 block HTT aggregate formation. - Abstract: Huntington’s disease is a polyglutamine expansion disorder, characterized by mutant HTT-mediated aggregate formation and cytotoxicity. Many reports suggests roles of N-terminal 17 amino acid domain of HTT (HTT-N17) towards subcellular localization, aggregate formation and subsequent pathogenicity induced by N-terminal HTT harboring polyQ stretch in pathogenic range. HYPK is a HTT-interacting chaperone which can reduce N-terminal mutant HTT-mediated aggregate formation and cytotoxicity in neuronal cell lines. However, how HYPK interacts with N-terminal fragment of HTT remained unknown. Here we report that specific interaction of HYPK with HTT-N17 is crucial for the chaperone activity of HYPK. Deletion of HTT-N17 leads to formation of tinier, SDS-soluble nuclear aggregates formed by N-terminal mutant HTT. The increased cytotoxicity imparted by these tiny aggregates might be contributed due to loss of interaction with HYPK.

  5. Barley Brassinosteroid Mutants Provide an Insight into Phytohormonal Homeostasis in Plant Reaction to Drought Stress

    Czech Academy of Sciences Publication Activity Database

    Gruszka, D.; Janeczko, A.; Dziurka, M.; Pociecha, E.; Oklešťková, Jana; Szarejko, I.

    2016-01-01

    Roč. 7, DEC 2 (2016), č. článku 1824. ISSN 1664-462X R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : abscisic- acid * arabidopsis-thaliana * jasmonic - acid * gibberellin biosynthesis * constitutive activation * abiotic stresses * brassica-napus * rice * responses * tolerance * barley * brassinosteroids * drought * homeostasis * mutants * phytohormones Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.298, year: 2016

  6. The role of the atypical kinases ABC1K7 and ABC1K8 in abscisic acid responses

    Directory of Open Access Journals (Sweden)

    Anna eManara

    2016-03-01

    Full Text Available The ABC1K family of atypical kinases (activity of bc1 complex kinase is represented in bacteria, archaea and eukaryotes. In plants they regulate diverse physiological processes in the chloroplasts and mitochondria, but their precise functions are poorly defined. ABC1K7 and ABC1K8 are probably involved in oxidative stress responses, isoprenyl lipid synthesis and distribution of iron within chloroplasts. Because reactive oxygen species take part in abscisic acid (ABA-mediated processes, we investigated the functions of ABC1K7 and ABC1K8 during germination, stomatal movement and leaf senescence. Both genes were upregulated by ABA treatment and some ABA-responsive physiological processes were affected in abc1k7 and abc1k8 mutants. Germination was more severely affected by ABA, osmotic stress and salt stress in the single and double mutants; the stomatal aperture was smaller in the mutants under standard growth conditions and was not further reduced by exogenous ABA application; ABA-induced senescence symptoms were more severe in the leaves of the single and double mutants compared to wild type leaves. Taken together, our results suggest that ABC1K7 and ABC1K8 might be involved in the cross-talk between ABA and ROS signaling.

  7. CKB1 is involved in abscisic acid and gibberellic acid signaling to regulate stress responses in Arabidopsis thaliana.

    Science.gov (United States)

    Yuan, Congying; Ai, Jianping; Chang, Hongping; Xiao, Wenjun; Liu, Lu; Zhang, Cheng; He, Zhuang; Huang, Ji; Li, Jinyan; Guo, Xinhong

    2017-05-01

    Casein kinase II (CK2), an evolutionarily well-conserved Ser/Thr kinase, plays critical roles in all higher organisms including plants. CKB1 is a regulatory subunit beta of CK2. In this study, homozygous T-DNA mutants (ckb1-1 and ckb1-2) and over-expression plants (35S:CKB1-1, 35S:CKB1-2) of Arabidopsis thaliana were studied to understand the role of CKB1 in abiotic stress and gibberellic acid (GA) signaling. Histochemical staining showed that although CKB1 was expressed in all organs, it had a relatively higher expression in conducting tissues. The ckb1 mutants showed reduced sensitivity to abscisic acid (ABA) during seed germination and seedling growth. The increased stomatal aperture, leaf water loss and proline accumulation were observed in ckb1 mutants. In contrast, the ckb1 mutant had increased sensitivity to polyaluminum chloride during seed germination and hypocotyl elongation. We obtained opposite results in over-expression plants. The expression levels of a number of genes in the ABA and GA regulatory network had changed. This study demonstrates that CKB1 is an ABA signaling-related gene, which subsequently influences GA metabolism, and may play a positive role in ABA signaling.

  8. Construction and functional analysis of Trichoderma harzianum mutants that modulate maize resistance to the pathogen Curvularia lunata.

    Science.gov (United States)

    Fan, Lili; Fu, Kehe; Yu, Chuanjin; Ma, Jia; Li, Yaqian; Chen, Jie

    2014-01-01

    Agrobacterium tumefaciens-mediated transformation (ATMT) was used to generate an insertional mutant library of the mycelial fungus Trichoderma harzianum. From a total of 450 mutants, six mutants that showed significant influence on maize resistance to C. lunata were analyzed in detail. Maize coated with these mutants was more susceptible to C. lunata compared with those coated with a wild-type (WT) strain. Similar to other fungal ATMT libraries, all six mutants were single copy integrations, which occurred preferentially in noncoding regions (except two mutants) and were frequently accompanied by the loss of border sequences. Two mutants (T66 and T312) that were linked to resistance were characterized further. Maize seeds coated with T66 and T312 were more susceptible to C. lunata than those treated with WT. Moreover, the mutants affected the resistance of maize to C. lunata by enhancing jasmonate-responsive gene expression. T66 and T312 induced maize resistance to C. lunata infection through a jasmonic acid-dependent pathway.

  9. Heterotrimeric G proteins-mediated resistance to necrotrophic pathogens includes mechanisms independent of salicylic acid-, jasmonic acid/ethylene- and abscisic acid-mediated defense signaling.

    Science.gov (United States)

    Trusov, Yuri; Sewelam, Nasser; Rookes, James Edward; Kunkel, Matt; Nowak, Ekaterina; Schenk, Peer Martin; Botella, José Ramón

    2009-04-01

    Heterotrimeric G proteins are involved in the defense response against necrotrophic fungi in Arabidopsis. In order to elucidate the resistance mechanisms involving heterotrimeric G proteins, we analyzed the effects of the Gβ (subunit deficiency in the mutant agb1-2 on pathogenesis-related gene expression, as well as the genetic interaction between agb1-2 and a number of mutants of established defense pathways. Gβ-mediated signaling suppresses the induction of salicylic acid (SA)-, jasmonic acid (JA)-, ethylene (ET)- and abscisic acid (ABA)-dependent genes during the initial phase of the infection with Fusarium oxysporum (up to 48 h after inoculation). However, at a later phase it enhances JA/ET-dependent genes such as PDF1.2 and PR4. Quantification of the Fusarium wilt symptoms revealed that Gβ- and SA-deficient mutants were more susceptible than wild-type plants, whereas JA- and ET-insensitive and ABA-deficient mutants demonstrated various levels of resistance. Analysis of the double mutants showed that the Gβ-mediated resistance to F. oxysporum and Alternaria brassicicola was mostly independent of all of the previously mentioned pathways. However, the progressive decay of agb1-2 mutants was compensated by coi1-21 and jin1-9 mutations, suggesting that at this stage of F. oxysporum infection Gβ acts upstream of COI1 and ATMYC2 in JA signaling. © 2008 The Authors. Journal compilation © 2008 Blackwell Publishing Ltd.

  10. Biosynthesis of Citric Acid from Glycerol by Acetate Mutants of Yarrowia lipolytica in Fed-Batch Fermentation

    Directory of Open Access Journals (Sweden)

    Anita Rywińska

    2009-01-01

    Full Text Available Pure and crude glycerol from biodiesel production have been used as substrates for citric acid production by acetate-negative mutants of Yarrowia lipolytica in fed-batch fermentation. Both the final concentration and the yield of the product were the highest when Y. lipolytica Wratislavia AWG7 strain was used in the culture with pure or crude glycerol. With a medium containing 200 g/L of glycerol, production reached a maximum of citric acid of 139 g/L after 120 h. This high yield of the product (up to 0.69 g of citric acid per gram of glycerol consumed was achieved with both pure and crude glycerol. Lower yield of citric acid in the culture with Y. lipolytica Wratislavia K1 strain (about 0.45 g/g resulted from increased erythritol concentrations (up to 40 g/L, accumulated simultaneously with the citric acid. The concentration of isocitric acid, a by-product in this fermentation, was very low, in the range from 2.6 to 4.6 g/L.

  11. Identification of a transcription factor controlling pH-dependent organic acid response in Aspergillus niger.

    Directory of Open Access Journals (Sweden)

    Lars Poulsen

    Full Text Available Acid formation in Aspergillus niger is known to be subjected to tight regulation, and the acid production profiles are fine-tuned to respond to the ambient pH. Based on transcriptome data, putative trans-acting pH responding transcription factors were listed and through knock out studies, mutants exhibiting an oxalate overproducing phenotype were identified. The yield of oxalate was increased up to 158% compared to the wild type and the corresponding transcription factor was therefore entitled Oxalic Acid repression Factor, OafA. Detailed physiological characterization of one of the ΔoafA mutants, compared to the wild type, showed that both strains produced substantial amounts of gluconic acid, but the mutant strain was more efficient in re-uptake of gluconic acid and converting it to oxalic acid, particularly at high pH (pH 5.0. Transcriptional profiles showed that 241 genes were differentially expressed due to the deletion of oafA and this supported the argument of OafA being a trans-acting transcription factor. Furthermore, expression of two phosphoketolases was down-regulated in the ΔoafA mutant, one of which has not previously been described in fungi. It was argued that the observed oxalate overproducing phenotype was a consequence of the efficient re-uptake of gluconic acid and thereby a higher flux through glycolysis. This results in a lower flux through the pentose phosphate pathway, demonstrated by the down-regulation of the phosphoketolases. Finally, the physiological data, in terms of the specific oxygen consumption, indicated a connection between the oxidative phosphorylation and oxalate production and this was further substantiated through transcription analysis.

  12. Mutants of alfalfa mosaic virus

    International Nuclear Information System (INIS)

    Roosien, J.

    1983-01-01

    In this thesis the isolation and characterization of a number of mutants of alfalfa mosaic virus, a plant virus with a coat protein dependent genome, is described. Thermo-sensitive (ts) mutants were selected since, at least theoretically, ts mutations can be present in all virus coded functions. It was found that a high percentage of spontaneous mutants, isolated because of their aberrant symptoms, were ts. The majority of these isolates could grow at the non-permissive temperature in the presence of a single wild type (wt) component. To increase the mutation rate virus preparations were treated with several mutagens. After nitrous acid treatment or irradiation with ultraviolet light, an increase in the level of mutations was observed. UV irradiation was preferred since it did not require large amounts of purified viral components. During the preliminary characterization of potential ts mutants the author also obtained one structural and several symptom mutants which were analysed further (chapter 7, 8 and 9). The properties of the ts mutants are described in chapter 3-7. (Auth.)

  13. Evaluation of Yield and Chemical Characteristics of some Peanut Mutants Induced by Gamma Irradiation

    International Nuclear Information System (INIS)

    Abd El-daem, G.A.; Anwar, M.M.

    2013-01-01

    This study was conducted to evaluate some promising mutants in peanut for yielding ability over three generation (M5, M6 and M7) and to evaluate yield attributes as will as chemical characteristics of these mutants in M7 generation induced by 100 Gy gamma radiation. The obtained results showed that the increase of yield / plot over three generation as a percentage of control was 5% for mutant 7, 10.2 % for mutant 10; 22% for mutant 9 and 22.9% for mutant 8. This increase in yield may be due to increase of one or more of yield attributes for most mutant lines. The significant increase for. No .of pods and seeds/ plant, weight of pods and seeds/ plant and 100- seed weight in M7 as compared to the control. For saturated fatty acid composition, results revealed that total saturated fatty acids ranged from 17.79% for mutant 8 to 21.75 for mutant 2 compared to 24.21% for control. Reduction of total saturated fatty acid was noticed for different mutants compared to that of the original variety. However, for total unsaturated fatty acids, results indicated that total unsaturated fatty acid composition ranged from 77.95% for mutant 9 to 82.09% for mutant 8 compared to 75.49% for control. Higher total unsaturated fatty acids for all mutant lines were obtained than that of the control, however, total saturated (TS)/ total unsaturated (TU) ratio was decreased for all mutants compared to control. The physical and chemical contents of Peanut oils showed that the refractive indices were ranged from 1.4620 to 1.4718 specific gravity were in range of 0.9146 to 0.9177. Acid value was range from 0.54 to 0.89% lodine value was ranged from 94.56 to 101.85. Saponification value was ranged from 185.2 to 190.7 and unsaponifiable matter was ranged from 0.98 to 1.33. The peroxide values ranged from 1.15 to 2.33 meq/kg oil. Also, fortified yoghurt made with replaced mutant peanut oil by 50% as milk fat substitute. Data showed that chemical composition and organolyptic properties had the

  14. Respiration responses of a polA1 and a tif-1 mutant of Escherichia coli to far-ultraviolet irradiation

    International Nuclear Information System (INIS)

    Swenson, P.A.

    1981-01-01

    Cessation of respiration in Escherichia coli 60 min after far - ultra-violet (254 nm) irradiation is dependent upon the recA and lexA gene products and is regulated by cyclic 3', 5'-adenosine monophosphate (cAMP) and its receptor protein. Two E. coli B/r mutants were studied, polA1 and tif-1, both of which express other rec/lex functions after UV irradiation. After receiving a relatively high UV fluence, the polA1 mutant, deficient in DNA polymerase 1, showed a respiration shutoff response like the wild type cells. 5-Fluorouracil and rifampin, an RNA synthesis inhibitor, did not prevent respiration shutoff in the mutant cells as they did in the wild type cells. At lower fluences which did not shut off respiration of polA1 cells, cAMP did not cause a more complete shutoff as it did for the wild type cells. The tif-1 mutant has a modified recA protein, and when unirradiated cells are incubated at 42 0 C they form filaments, mutate, and show other rec/lex responses. This mutant did not shut off its respiration at either 30 or 42 0 C, and the response was not modified by cAMP. In an E. coli K12 strain, W3110, 52 J/m 2 UV did not shut off respiration and cAMP had no effect. (author)

  15. Impact of Branched-Chain Amino Acid Catabolism on Fatty Acid and Alkene Biosynthesis in Micrococcus luteus.

    Science.gov (United States)

    Surger, Maximilian J; Angelov, Angel; Stier, Philipp; Übelacker, Maria; Liebl, Wolfgang

    2018-01-01

    Micrococcus luteus naturally produces alkenes, unsaturated aliphatic hydrocarbons, and represents a promising host to produce hydrocarbons as constituents of biofuels and lubricants. In this work, we identify the genes for key enzymes of the branched-chain amino acid catabolism in M. luteus , whose first metabolic steps lead also to the formation of primer molecules for branched-chain fatty acid and olefin biosynthesis, and demonstrate how these genes can be used to manipulate the production of specific olefins in this organism. We constructed mutants of several gene candidates involved in the branched-chain amino acid metabolism or its regulation and investigated the resulting changes in the cellular fatty acid and olefin profiles by GC/MS. The gene cluster encoding the components of the branched-chain α-keto acid dehydrogenase (BCKD) complex was identified by deletion and promoter exchange mutagenesis. Overexpression of the BCKD gene cluster resulted in about threefold increased olefin production whereas deletion of the cluster led to a drastic reduction in branched-chain fatty acid content and a complete loss of olefin production. The specificities of the acyl-CoA dehydrogenases of the branched amino acid degradation pathways were deduced from the fatty acid and olefin profiles of the respective deletion mutant strains. In addition, growth experiments with branched amino acids as the only nitrogen source were carried out with the mutants in order to confirm our annotations. Both the deletion mutant of the BCKD complex, responsible for the further degradation of all three branched-chain amino acids, as well as the deletion mutant of the proposed isovaleryl-CoA dehydrogenase (specific for leucine degradation) were not able to grow on leucine in contrast to the parental strain. In conclusion, our experiments allow the unambigous assignment of specific functions to the genes for key enzymes of the branched-chain amino acid metabolism of M. luteus . We also show how

  16. Identification of a classical mutant in the industrial host Aspergillus niger by systems genetics: LaeA is required for citric acid production and regulates the formation of some secondary metabolites

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Jing; Arentshorst, Mark; Nair, Deepa; Dai, Ziyu; Baker, Scott E.; Frisvad, Jens; Nielsen, Kristian F.; Punt, Peter J.; Ram, Arthur F.

    2016-01-11

    Rapid acidification of the culture medium by the production of organic acids and the production of acid-induced proteases are key characteristics of the filamentous fungus Aspergillus niger. The D15 mutant of A. niger is non-acidifying mutant and used often for the expression of recombinant proteins in A. niger, because of its reduced production of extracellular proteases under non-acidic conditions. In this study, the D15 mutant is characterized in detail. Strongly reduced levels of citric and oxalic acid were observed in the D15 mutant both in shake flask cultures and in controlled batch cultivations. To identify the mutation in the D15 mutant, we successfully combined high-throughput sequencing (Illumina) with bulk segregant analysis. Because of the lack of a sexual cycle for A. niger, the parasexual cycle was used to generate a pool of segregants. From the 52 single nucleotide polymorphisms (SNPs) between the parental strains, three SNPs were homozygous in the genomic DNA of pool of segregants. These three SNPs mapped to all the right arm of chromosome II, indicating that this region contains the genetic locus affecting the phenotype related to acid production. Of the three SNPs, one mutation resulted in a missense mutation in the gene encoding the A. niger homologue of the A. nidulans methyltransferase gene laeA. Complementation analysis of the original mutant with the laeA gene and targeted disruption of laeA further confirmed that LaeA is involved in citric acid production in A. niger lab (N402) and citric acid production strains (ATCC 11414). Analysis of the secondary metabolite (SM) profile of the laeA mutants indicate that LaeA is required for the production of several SMs (asperrubrol, atromentin and JBIR86), but deletion of laeA also resulted in the presence of SMs (aspernigrin A/B and BMS-192548) that were not detected in the wild-type strain. The levels of ten other SMs were not strongly affected as a result of laeA deletion indicating that only a

  17. Identification of key uric acid synthesis pathway in a unique mutant silkworm Bombyx mori model of Parkinson's disease.

    Directory of Open Access Journals (Sweden)

    Hiroko Tabunoki

    Full Text Available Plasma uric acid (UA levels decrease following clinical progression and stage development of Parkinson's disease (PD. However, the molecular mechanisms underlying decreases in plasma UA levels remain unclear, and the potential to apply mutagenesis to a PD model has not previously been discovered. We identified a unique mutant of the silkworm Bombyx mori (B.mori op. Initially, we investigated the causality of the phenotypic "op" by microarray analysis using our constructed KAIKO functional annotation pipeline. Consequently, we found a novel UA synthesis-modulating pathway, from DJ-1 to xanthine oxidase, and established methods for large-scale analysis of gene expression in B. mori. We found that the mRNA levels of genes in this pathway were significantly lower in B. mori op mutants, indicating that downstream events in the signal transduction cascade might be prevented. Additionally, levels of B.mori tyrosine hydroxylase (TH and DJ-1 mRNA were significantly lower in the brain of B. mori op mutants. UA content was significantly lower in the B. mori op mutant tissues and hemolymph. The possibility that the B. mori op mutant might be due to loss of DJ-1 function was supported by the observed vulnerability to oxidative stress. These results suggest that UA synthesis, transport, elimination and accumulation are decreased by environmental oxidative stress in the B. mori op mutant. In the case of B. mori op mutants, the relatively low availability of UA appears to be due both to the oxidation of DJ-1 and to its expenditure to mitigate the effects of environmental oxidative stress. Our findings are expected to provide information needed to elucidate the molecular mechanism of decreased plasma UA levels in the clinical stage progression of PD.

  18. Mechanisms responsible for imipenem resistance among Pseudomonas aeruginosa clinical isolates exposed to imipenem concentrations within the mutant selection window.

    Science.gov (United States)

    Vassilara, Foula; Galani, Irene; Souli, Maria; Papanikolaou, Konstantinos; Giamarellou, Helen; Papadopoulos, Antonios

    2017-07-01

    The aim of this study was to determine the propensities of imipenem to select for resistant Pseudomonas aeruginosa mutants by determining the mutant prevention concentrations (MPCs) for 9 unrelated clinical isolates and the accession of any relationship with mechanisms of resistance development. The MPC/MIC ratios ranged from 4 to 16. Detection of resistance mechanisms in the mutant derivatives of the nine isolates mainly revealed inactivating mutations in the gene coding for outer membrane protein OprD. Point mutations leading to premature stop codons or amino acid substitution S278P, ≥1bp deletion leading to frameshift mutations and interruption of the oprD by an insertion sequence, were observed. MPC and mutant selection window (MSW) are unique parameters that may guide the implementation of antimicrobial treatment, providing useful information about the necessary imipenem concentration needed in the infection area, in order to avoid the emergence of resistance, especially in clinical situations with high bacterial load. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Interactive Effects of Jasmonic Acid, Salicylic Acid, and Gibberellin on Induction of Trichomes in Arabidopsis1

    Science.gov (United States)

    Traw, M. Brian; Bergelson, Joy

    2003-01-01

    Leaf trichomes protect plants from attack by insect herbivores and are often induced following damage. Hormonal regulation of this plant induction response has not been previously studied. In a series of experiments, we addressed the effects of artificial damage, jasmonic acid, salicylic acid, and gibberellin on induction of trichomes in Arabidopsis. Artificial damage and jasmonic acid caused significant increases in trichome production of leaves. The jar1-1 mutant exhibited normal trichome induction following treatment with jasmonic acid, suggesting that adenylation of jasmonic acid is not necessary. Salicylic acid had a negative effect on trichome production and consistently reduced the effect of jasmonic acid, suggesting negative cross-talk between the jasmonate and salicylate-dependent defense pathways. Interestingly, the effect of salicylic acid persisted in the nim1-1 mutant, suggesting that the Npr1/Nim1 gene is not downstream of salicylic acid in the negative regulation of trichome production. Last, we found that gibberellin and jasmonic acid had a synergistic effect on the induction of trichomes, suggesting important interactions between these two compounds. PMID:14551332

  20. A cataract-causing connexin 50 mutant is mislocalized to the ER due to loss of the fourth transmembrane domain and cytoplasmic domain.

    Science.gov (United States)

    Somaraju Chalasani, Madhavi Latha; Muppirala, Madhavi; G Ponnam, Surya Prakash; Kannabiran, Chitra; Swarup, Ghanshyam

    2013-01-01

    Mutations in the eye lens gap junction protein connexin 50 cause cataract. Earlier we identified a frameshift mutant of connexin 50 (c.670insA; p.Thr203AsnfsX47) in a family with autosomal recessive cataract. The mutant protein is smaller and contains 46 aberrant amino acids at the C-terminus after amino acid 202. Here, we have analysed this frameshift mutant and observed that it localized to the endoplasmic reticulum (ER) but not in the plasma membrane. Moreover, overexpression of the mutant resulted in disintegration of the ER-Golgi intermediate compartment (ERGIC), reduction in the level of ERGIC-53 protein and breakdown of the Golgi in many cells. Overexpression of the frameshift mutant partially inhibited the transport of wild type connexin 50 to the plasma membrane. A deletion mutant lacking the aberrant sequence showed predominant localization in the ER and inhibited anterograde protein transport suggesting, therefore, that the aberrant sequence is not responsible for improper localization of the frameshift mutant. Further deletion analysis showed that the fourth transmembrane domain and a membrane proximal region (231-294 amino acids) of the cytoplasmic domain are needed for transport from the ER and localization to the plasma membrane. Our results show that a frameshift mutant of connexin 50 mislocalizes to the ER and causes disintegration of the ERGIC and Golgi. We have also identified a sequence of connexin 50 crucial for transport from the ER and localization to the plasma membrane.

  1. Characterization of Brucella abortus mutant strain Δ22915, a potential vaccine candidate.

    Science.gov (United States)

    Bao, Yanqing; Tian, Mingxing; Li, Peng; Liu, Jiameng; Ding, Chan; Yu, Shengqing

    2017-04-04

    Brucellosis, caused by Brucella spp., is an important zoonosis worldwide. Vaccination is an effective strategy for protection against Brucella infection in livestock in developing countries and in wildlife in developed countries. However, current vaccine strains including S19 and RB51 are pathogenic to humans and pregnant animals, limiting their use. In this study, we constructed the Brucella abortus (B. abortus) S2308 mutant strain Δ22915, in which the putative lytic transglycosylase gene BAB_RS22915 was deleted. The biological properties of mutant strain Δ22915 were characterized and protection of mice against virulent S2308 challenge was evaluated. The mutant strain Δ22915 showed reduced survival within RAW264.7 cells and survival in vivo in mice. In addition, the mutant strain Δ22915 failed to escape fusion with lysosomes within host cells, and caused no observable pathological damage. RNA-seq analysis indicated that four genes associated with amino acid/nucleotide transport and metabolism were significantly upregulated in mutant strain Δ22915. Furthermore, inoculation of ∆22915 at 10 5 colony forming units induced effective host immune responses and long-term protection of BALB/c mice. Therefore, mutant strain ∆22915 could be used as a novel vaccine candidate in the future to protect animals against B. abortus infection.

  2. Genetics of dwarfness in induced mutants of hexaploid triticale and its response to exogenous GA3

    International Nuclear Information System (INIS)

    Reddy, V.D.; Reddy, G.M.

    1991-01-01

    Genetics of dwarfism in two induced mutant (d 1 and d 2 ) of hexaploid triticale, DTS 330, revealed that this trait is governed by single recessive gene. Both d 1 and d 2 were allelic to each other and d 1 was dominant over d 2 . Both d 1 , d 2 and their F 1 showed no response to exogenous GA 3 , whereas, DTS 330, d 1 x DTS 330 and d 2 x DTS 330 were responsive. The endogenous levels of GA 3 were more in the dwarf mutants than control, suggesting that dwarfness in these may be due to a partial block in the GA utilizing mechanism, rather than a block in GA biosynthesis. (author). 5 refs., 2 tabs

  3. Antagonism between salicylic and abscisic acid reflects early host-pathogen conflict and moulds plant defence responses.

    Science.gov (United States)

    de Torres Zabala, Marta; Bennett, Mark H; Truman, William H; Grant, Murray R

    2009-08-01

    The importance of phytohormone balance is increasingly recognized as central to the outcome of plant-pathogen interactions. Recently it has been demonstrated that abscisic acid signalling pathways are utilized by the bacterial phytopathogen Pseudomonas syringae to promote pathogenesis. In this study, we examined the dynamics, inter-relationship and impact of three key acidic phytohormones, salicylic acid, abscisic acid and jasmonic acid, and the bacterial virulence factor, coronatine, during progression of P. syringae infection of Arabidopsis thaliana. We show that levels of SA and ABA, but not JA, appear to play important early roles in determining the outcome of the infection process. SA is required in order to mount a full innate immune responses, while bacterial effectors act rapidly to activate ABA biosynthesis. ABA suppresses inducible innate immune responses by down-regulating SA biosynthesis and SA-mediated defences. Mutant analyses indicated that endogenous ABA levels represent an important reservoir that is necessary for effector suppression of plant-inducible innate defence responses and SA synthesis prior to subsequent pathogen-induced increases in ABA. Enhanced susceptibility due to loss of SA-mediated basal resistance is epistatically dominant over acquired resistance due to ABA deficiency, although ABA also contributes to symptom development. We conclude that pathogen-modulated ABA signalling rapidly antagonizes SA-mediated defences. We predict that hormonal perturbations, either induced or as a result of environmental stress, have a marked impact on pathological outcomes, and we provide a mechanistic basis for understanding priming events in plant defence.

  4. Biosynthetic pathways to delta-aminolevulinic acid induced by blue light in the pigment mutant C-2A' of Scenedesmus obliquus

    International Nuclear Information System (INIS)

    Klein, O.; Senger, H.

    1978-01-01

    The X-ray induced mutant C-2A' of Scenedesmus obliquus grows heterotrophically but forms only traces of chlorophyll in the dark. Upon illumination, delta-aminolevulinic acid (ALA) is synthesized and chlorophyll is formed. These processes are blue light dependent and ceased immediately when the cells were transferred back into darkness. Addition of levulinic acid (LA) inhibited the light-dependent formation of chlorophyll and caused accumulation of ALA by competitive inhibition of the ALA dehydratase (EC. 4.2.1.24). By feeding specifically labelled 14 C precursors to the pigment mutant, inhibiting the ALA dehydratase with LA, accumulating, extracting and analyzing the ALA, two pathways leading towards ALA could be established: glycine and succinyl CoA can be condensed to ALA and the 5 carbon skeleton of glutamate can completely be incorporated into ALA via a second pathway. The glycine-succinyl CoA pathway dominated over the glutamate pathway, but both led to chlorophyll formation. (author)

  5. Mechanical touch responses of Arabidopsis TCH1-3 mutant roots on inclined hard-agar surface

    Science.gov (United States)

    Zha, Guodong; Wang, Bochu; Liu, Junyu; Yan, Jie; Zhu, Liqing; Yang, Xingyan

    2016-01-01

    The gravity-induced mechanical touch stimulus can affect plant root architecture. Mechanical touch responses of plant roots are an important aspect of plant root growth and development. Previous studies have reported that Arabidopsis TCH1-3 genes are involved in mechano-related events, how-ever, the physiological functions of TCH1-3 genes in Arabidopsis root mechanoresponses remain unclear. In the present study, we applied an inclined hard agar plate method to produce mechanical touch stimulus, and provided evidence that altered mechanical environment could influence root growth. Furthermore, tch1-3 Arabidopsis mutants were investigated on inclined agar surfaces to explore the functions of TCH1-3 genes on Arabidopsis root mechanoresponses. The results showed that two tch2 mutants, cml24-2 and cml24-4, exhibited significantly reduced root length, biased skewing, and decreased density of lateral root. In addition, primary root length and density of lateral root of tch3 (cml12-2) was significantly decreased on inclined agar surfaces. This study indicates that the tch2 and tch3 mutants are hypersensitive to mechanical touch stimulus, and TCH2 (CML24-2 and CML24-4) and TCH3 (CML12-2) genes may participate in the mechanical touch response of Arabidopsis roots.

  6. A study of eukaryotic response mechanisms to atmospheric pressure cold plasma by using Saccharomyces cerevisiae single gene mutants

    International Nuclear Information System (INIS)

    Feng Hongqing; Wang Ruixue; Sun Peng; Wu Haiyan; Liu Qi; Li Fangting; Fang Jing; Zhang Jue; Zhu Weidong

    2010-01-01

    The mechanisms of eukaryotic cell response to cold plasma are studied. A series of single gene mutants of eukaryotic model organism Saccharomyces cerevisiae are used to compare their sensitivity to plasma treatment with the wild type. We examined 12 mutants in the oxidative stress pathway and the cell cycle pathway, in which 8 are found to be hypersensitive to plasma processing. The mutated genes' roles in the two pathways are analyzed to understand the biological response mechanisms of plasma treatment. The results demonstrate that genes from both pathways are needed for the eukaryotic cells to survive the complex plasma treatment.

  7. Reduced heme levels underlie the exponential growth defect of the Shewanella oneidensis hfq mutant.

    Directory of Open Access Journals (Sweden)

    Christopher M Brennan

    Full Text Available The RNA chaperone Hfq fulfills important roles in small regulatory RNA (sRNA function in many bacteria. Loss of Hfq in the dissimilatory metal reducing bacterium Shewanella oneidensis strain MR-1 results in slow exponential phase growth and a reduced terminal cell density at stationary phase. We have found that the exponential phase growth defect of the hfq mutant in LB is the result of reduced heme levels. Both heme levels and exponential phase growth of the hfq mutant can be completely restored by supplementing LB medium with 5-aminolevulinic acid (5-ALA, the first committed intermediate synthesized during heme synthesis. Increasing expression of gtrA, which encodes the enzyme that catalyzes the first step in heme biosynthesis, also restores heme levels and exponential phase growth of the hfq mutant. Taken together, our data indicate that reduced heme levels are responsible for the exponential growth defect of the S. oneidensis hfq mutant in LB medium and suggest that the S. oneidensis hfq mutant is deficient in heme production at the 5-ALA synthesis step.

  8. Physiological and Proteomic Analysis of the Rice Mutant cpm2 Suggests a Negative Regulatory Role of Jasmonic Acid in Drought Tolerance

    Directory of Open Access Journals (Sweden)

    Rohit Dhakarey

    2017-11-01

    Full Text Available It is widely known that numerous adaptive responses of drought-stressed plants are stimulated by chemical messengers known as phytohormones. Jasmonic acid (JA is one such phytohormone. But there are very few reports revealing its direct implication in drought related responses or its cross-talk with other phytohormones. In this study, we compared the morpho-physiological traits and the root proteome of a wild type (WT rice plant with its JA biosynthesis mutant coleoptile photomorphogenesis 2 (cpm2, disrupted in the allene oxide cyclase (AOC gene, for insights into the role of JA under drought. The mutant had higher stomatal conductance, higher water use efficiency and higher shoot ABA levels under severe drought as compared to the WT. Notably, roots of cpm2 were better developed compared to the WT under both, control and drought stress conditions. Root proteome was analyzed using the Tandem Mass Tag strategy to better understand this difference at the molecular level. Expectedly, AOC was unique but notably highly abundant under drought in the WT. Identification of other differentially abundant proteins (DAPs suggested increased energy metabolism (i.e., increased mobilization of resources and reactive oxygen species scavenging in cpm2 under drought. Additionally, various proteins involved in secondary metabolism, cell growth and cell wall synthesis were also more abundant in cpm2 roots. Proteome-guided transcript, metabolite, and histological analyses provided further insights into the favorable adaptations and responses, most likely orchestrated by the lack of JA, in the cpm2 roots. Our results in cpm2 are discussed in the light of JA crosstalk to other phytohormones. These results together pave the path for understanding the precise role of JA during drought stress in rice.

  9. Flowering responses to light-breaks in photomorphogenic mutants of Arabidopsis thaliana, a long-day plant

    International Nuclear Information System (INIS)

    Goto, N.; Kumagai, T.; Koornneef, M.

    1991-01-01

    Flowering response and plant form of photomorphogenic mutants (hy1, hy2, hy3, hy4 and hy5) of Arabidopsis thaliana (L.), a long-day plant, were examined in long and short days. There were only slight differences among genotypes including Landsberg wild type with respect to the flowering time under long days. The effect of 1 h light-(night)-breaks of far-red, red, blue and white light given in the middle of the dark period of plants grown under short days, was studied. Effects of far-red light applied at the end or the beginning of the main photoperiod on flowering and plant form were also examined. The light-breaks with all the above mentioned light qualities promoted floral initiation of all the genotypes including the wild type in terms of both the flowering time and the number of rosette leaves. In general, far-red light was most effective. It is possible to classify the hy-mutants into 3 groups by their responses to light-breaks under short day conditions: (a) Mutants hy2 and hy3, which have a reduced number of rosette leaves, and flower early. Red light is as effective as far-red light. The wavelength of light-breaks is relatively unimportant for flowering response. (b) Mutants hy4, hy5 and Landsberg wild type, which have a greater number of rosette leaves, and flower relatively late. The effectiveness of light-breaks is in the following order, far-red, blue, and red light, which is in reverse order to the transformation of phytochrome to the P fr form. (c) Mutant hy1, which behaves anomalously with respect to relations between flowering time and number of rosette leaves; late flowering with reduced number of rosette leaves. Red, blue and far-red light are effective, but white light is ineffective for reducing the number of rosette leaves. When far-red light was given in the middle of the night or at the end of the main photoperiod, it markedly reduced the number of rosette leaves compared to those grown under short days for all the genotypes, while when

  10. Transcriptional regulation of Arabidopsis MIR168a and argonaute1 homeostasis in abscisic acid and abiotic stress responses.

    Science.gov (United States)

    Li, Wei; Cui, Xiao; Meng, Zhaolu; Huang, Xiahe; Xie, Qi; Wu, Heng; Jin, Hailing; Zhang, Dabing; Liang, Wanqi

    2012-03-01

    The accumulation of a number of small RNAs in plants is affected by abscisic acid (ABA) and abiotic stresses, but the underlying mechanisms are poorly understood. The miR168-mediated feedback regulatory loop regulates ARGONAUTE1 (AGO1) homeostasis, which is crucial for gene expression modulation and plant development. Here, we reveal a transcriptional regulatory mechanism by which MIR168 controls AGO1 homeostasis during ABA treatment and abiotic stress responses in Arabidopsis (Arabidopsis thaliana). Plants overexpressing MIR168a and the AGO1 loss-of-function mutant ago1-27 display ABA hypersensitivity and drought tolerance, while the mir168a-2 mutant shows ABA hyposensitivity and drought hypersensitivity. Both the precursor and mature miR168 were induced under ABA and several abiotic stress treatments, but no obvious decrease for the target of miR168, AGO1, was shown under the same conditions. However, promoter activity analysis indicated that AGO1 transcription activity was increased under ABA and drought treatments, suggesting that transcriptional elevation of MIR168a is required for maintaining a stable AGO1 transcript level during the stress response. Furthermore, we showed both in vitro and in vivo that the transcription of MIR168a is directly regulated by four abscisic acid-responsive element (ABRE) binding factors, which bind to the ABRE cis-element within the MIR168a promoter. This ABRE motif is also found in the promoter of MIR168a homologs in diverse plant species. Our findings suggest that transcriptional regulation of miR168 and posttranscriptional control of AGO1 homeostasis may play an important and conserved role in stress response and signal transduction in plants.

  11. Abscisic acid and transpiration rate are involved in the response to boron toxicity in Arabidopsis plants.

    Science.gov (United States)

    Macho-Rivero, Miguel Ángel; Camacho-Cristóbal, Juan José; Herrera-Rodríguez, María Begoña; Müller, Maren; Munné-Bosch, Sergi; González-Fontes, Agustín

    2017-05-01

    Boron (B) is an essential microelement for vascular plant development, but its toxicity is a major problem affecting crop yields in arid and semi-arid areas of the world. In the literature, several genes involved in abscisic acid (ABA) signalling and responses are upregulated in Arabidopsis roots after treatment with excess B. It is known that the AtNCED3 gene, which encodes a crucial enzyme for ABA biosynthesis, plays a key role in the plant response to drought stress. In this study, root AtNCED3 expression and shoot ABA content were rapidly increased in wild-type plants upon B-toxicity treatment. The Arabidopsis ABA-deficient nced3-2 mutant had higher transpiration rate, stomatal conductance and accumulated more B in their shoots than wild-type plants, facts that were associated with the lower levels of ABA in this mutant. However, in wild-type plants, B toxicity caused a significant reduction in stomatal conductance, resulting in a decreased transpiration rate. This response could be a mechanism to limit the transport of excess B from the roots to the leaves under B toxicity. In agreement with the higher transpiration rate of the nced3-2 mutant, this genotype showed an increased leaf B concentration and damage upon exposure to 5 mM B. Under B toxicity, ABA application decreased B accumulation in wild-type and nced3-2 plants. In summary, this work shows that excess B applied to the roots leads to rapid changes in AtNCED3 expression and gas exchange parameters that would contribute to restrain the B entry into the leaves, this effect being mediated by ABA. © 2016 Scandinavian Plant Physiology Society.

  12. Evaluation of Chemical Changes in Some Soybean Mutants Induced by Gamma Irradiation

    International Nuclear Information System (INIS)

    Abd-Elkalik, K.; Mekkawy, S.H.; El-Demerdash, H.M.

    2010-01-01

    The Egyptian soybean cultivar Giza-22 was used to induce genetic variability by using gamma rays at dose levels of 100, 150 and 200 Gy. Sixteen mutants (including parental cultivar) were evaluated in M3 generation for their agronomic traits and chemical analysis was done in seeds of M3 generation. Four mutants A21 (150 Gy), A22, A23 and A24 (200 Gy) showed superiority in their agronomic traits compared with parental cultivar. The results of chemical analysis of seeds of M3 generation showed that, oil and energy contents were unaffected by irradiation treatments while protein contents were significantly increased at doses 150 and 200 Gy. Phenolic and tannin contents in seeds of M3 generation showed no significant changes in their percentages due to irradiation treatments. Gamma irradiation significantly increased in linoleic acid content in most of the mutants compared with the control (Giza-22), whereas, there were decreases in linolenic acid content. Investigation of amino acid composition of mutants of M3 generation revealed significant increases in the essential amino acids in most mutants induced by gamma irradiation at 150 and 200 Gy. It could be suggested that the use of gamma irradiation can induce an improvement of the oil and protein composition in soybean

  13. Clavulanic acid production by the MMS 150 mutant obtained from wild type Streptomyces clavuligerus ATCC 27064

    Directory of Open Access Journals (Sweden)

    Eliton da Silva Vasconcelos

    2013-12-01

    Full Text Available Clavulanic acid (CA is a powerful inhibitor of the beta-lactamases, enzymes produced by bacteria resistants to penicillin and cefalosporin. This molecule is produced industrially by strains of Streptomyces clavuligerus in complex media which carbon and nitrogen resources are supplied by inexpensive compounds still providing high productivity. The genetic production improvement using physical and chemical mutagenic agents is an important strategy in programs of industrial production development of bioactive metabolites. However, parental strains are susceptible to loss of their original productivity due genetic instability phenomenona. In this work, some S. clavuligerus mutant strains obtained by treatment with UV light and with MMS are compared with the wild type (Streptomyces clavuligerus ATCC 27064. The results indicated that the random mutations originated some strains with different phenotypes, most divergent demonstrated by the mutants strains named AC116, MMS 150 and MMS 54, that exhibited lack of pigmentation in their mature spores. Also, the strain MMS 150 presented a larger production of CA when cultivated in semi-synthetics media. Using other media, the wild type strain obtained a larger CA production. Besides, using the modifed complex media the MMS 150 strain showed changes in its lipolitic activity and a larger production of CA. The studies also allowed finding the best conditions for a lipase activity exhibited by wild type S. clavuligerus and the MMS150 mutant.

  14. Clavulanic acid production by the MMS 150 mutant obtained from wild type Streptomyces clavuligerus ATCC 27064.

    Science.gov (United States)

    da Silva Vasconcelos, Eliton; de Lima, Vanderlei Aparecido; Goto, Leandro Seiji; Cruz-Hernández, Isara Lourdes; Hokka, Carlos Osamu

    2013-12-01

    Clavulanic acid (CA) is a powerful inhibitor of the beta-lactamases, enzymes produced by bacteria resistants to penicillin and cefalosporin. This molecule is produced industrially by strains of Streptomyces clavuligerus in complex media which carbon and nitrogen resources are supplied by inexpensive compounds still providing high productivity. The genetic production improvement using physical and chemical mutagenic agents is an important strategy in programs of industrial production development of bioactive metabolites. However, parental strains are susceptible to loss of their original productivity due genetic instability phenomenona. In this work, some S. clavuligerus mutant strains obtained by treatment with UV light and with MMS are compared with the wild type (Streptomyces clavuligerus ATCC 27064). The results indicated that the random mutations originated some strains with different phenotypes, most divergent demonstrated by the mutants strains named AC116, MMS 150 and MMS 54, that exhibited lack of pigmentation in their mature spores. Also, the strain MMS 150 presented a larger production of CA when cultivated in semi-synthetics media. Using other media, the wild type strain obtained a larger CA production. Besides, using the modifed complex media the MMS 150 strain showed changes in its lipolitic activity and a larger production of CA. The studies also allowed finding the best conditions for a lipase activity exhibited by wild type S. clavuligerus and the MMS150 mutant.

  15. Evaluate The Fluctuation Of Phytic Acid Content In Seed From Mutant Lines By Gamma Ray

    International Nuclear Information System (INIS)

    Nguyen Thi Lang; Pham Van Ut

    2011-01-01

    Phytic acid is a molecule composed of myo-inositol 1,2,3,4,5,6 hexakis dihydrogen phosphate (Ins P6), a major component in the source of phosphorus (P) reserves of about 50 plants - 80% of total seed phosphorus (Lott, 1984). At physiological pH in the form of phytic acid have negatively charged ions hold together the complex mineral nutrition creates indigestion. Moreover, phosphorus in the form of phytate or phytic humans and monogastric animals can not absorb, are all discharged polluted environment transitions. In rice OM819, OM4900, OM3536, D4 and D8 are irradiated with gamma rays at 5 doses: 100, 200, 300, 400 and 500 Gy to create mutant strains with low levels of phytic acid. Results in radiation levels may appear 100 Gy line grain phytic acid expression is low. At the level 200 Gy of radiation is three populations OM819, OM4900 and OM3536 with 8 lines for grain phytic acid expression is low. At 300 Gy extent, appeared seven lines with low nuclear expression of phytic acid 4 populations OM819, OM4900, OM3536 and D4. At the level 400 Gy of radiation there are 4 populations appear only 5 lines expressed low phytic acid, with 3-line expression levels 3 and 2 lines with level 4 expression. At the level of radiation 500 Gy only one line appears at level 3 of phytic acid this is OM819 populations. For genotype analysis using marker RM 261 with 66.67% of the rice low phytic acid content of the expression analysis of biochemical polymorphisms. (author)

  16. Probiotic features of Lactobacillus plantarum mutant strains.

    Science.gov (United States)

    Bove, Pasquale; Gallone, Anna; Russo, Pasquale; Capozzi, Vittorio; Albenzio, Marzia; Spano, Giuseppe; Fiocco, Daniela

    2012-10-01

    In this study, the probiotic potential of Lactobacillus plantarum wild-type and derivative mutant strains was investigated. Bacterial survival was evaluated in an in vitro system, simulating the transit along the human oro-gastro-intestinal tract. Interaction with human gut epithelial cells was studied by assessing bacterial adhesive ability to Caco-2 cells and induction of genes involved in innate immunity. L. plantarum strains were resistant to the combined stress at the various steps of the simulated gastrointestinal tract. Major decreases in the viability of L. plantarum cells were observed mainly under drastic acidic conditions (pH ≤ 2.0) of the gastric compartment. Abiotic stresses associated to small intestine poorly affected bacterial viability. All the bacterial strains significantly adhered to Caco-2 cells, with the ΔctsR mutant strain exhibiting the highest adhesion. Induction of immune-related genes resulted higher upon incubation with heat-inactivated bacteria rather than with live ones. For specific genes, a differential transcriptional pattern was observed upon stimulation with different L. plantarum strains, evidencing a possible role of the knocked out bacterial genes in the modulation of host cell response. In particular, cells from Δhsp18.55 and ΔftsH mutants strongly triggered immune defence genes. Our study highlights the relevance of microbial genetic background in host-probiotic interaction and might contribute to identify candidate bacterial genes and molecules involved in probiosis.

  17. Conjugated Fatty Acid Synthesis

    Science.gov (United States)

    Rawat, Richa; Yu, Xiao-Hong; Sweet, Marie; Shanklin, John

    2012-01-01

    Conjugated linolenic acids (CLNs), 18:3 Δ9,11,13, lack the methylene groups found between the double bonds of linolenic acid (18:3 Δ9,12,15). CLNs are produced by conjugase enzymes that are homologs of the oleate desaturases FAD2. The goal of this study was to map the domain(s) within the Momordica charantia conjugase (FADX) responsible for CLN formation. To achieve this, a series of Momordica FADX-Arabidopsis FAD2 chimeras were expressed in the Arabidopsis fad3fae1 mutant, and the transformed seeds were analyzed for the accumulation of CLN. These experiments identified helix 2 and the first histidine box as a determinant of conjugase product partitioning into punicic acid (18:3 Δ9cis,11trans,13cis) or α-eleostearic acid (18:3 Δ9cis,11trans,13trans). This was confirmed by analysis of a FADX mutant containing six substitutions in which the sequence of helix 2 and first histidine box was converted to that of FAD2. Each of the six FAD2 substitutions was individually converted back to the FADX equivalent identifying residues 111 and 115, adjacent to the first histidine box, as key determinants of conjugase product partitioning. Additionally, expression of FADX G111V and FADX G111V/D115E resulted in an approximate doubling of eleostearic acid accumulation to 20.4% and 21.2%, respectively, compared with 9.9% upon expression of the native Momordica FADX. Like the Momordica conjugase, FADX G111V and FADX D115E produced predominantly α-eleostearic acid and little punicic acid, but the FADX G111V/D115E double mutant produced approximately equal amounts of α-eleostearic acid and its isomer, punicic acid, implicating an interactive effect of residues 111 and 115 in punicic acid formation. PMID:22451660

  18. Is auxin involved in the induction of genetic instability in barley homeotic double mutants?

    Science.gov (United States)

    Šiukšta, Raimondas; Vaitkūnienė, Virginija; Rančelis, Vytautas

    2018-02-01

    The triggers of genetic instability in barley homeotic double mutants are tweaky spike -type mutations associated with an auxin imbalance in separate spike phytomeres. Barley homeotic tweaky spike;Hooded (tw;Hd) double mutants are characterized by an inherited instability of spike and flower development, which is absent in the single parental constituents. The aim of the present study was to show that the trigger of genetic instability in the double mutants is the tw mutations, which are associated with an auxin imbalance in the developing spikes. Their pleiotropic effects on genes related to spike/flower development may cause the genetic instability of double mutants. The study of four double-mutant groups composed of different mutant alleles showed that the instability arose only if the mutant allele tw was a constituent of the double mutants. Application of auxin inhibitors and 2,4-dichlorophenoxyacetic acid (2,4-D) demonstrated the relationship of the instability of the double mutants and the phenotype of the tw mutants to auxin imbalance. 2,4-D induced phenocopies of the tw mutation in wild-type plants and rescued the phenotypes of three allelic tw mutants. The differential display (dd-PCR) method allowed the identification of several putative candidate genes in tw that may be responsible for the initiation of instability in the double mutants by pleiotropic variations of their expression in the tw mutant associated with auxin imbalance in the developing spikes. The results of the present study linked the genetic instability of homeotic double mutants with an auxin imbalance caused by one of the constituents (tw). The genetic instability of the double mutants in relation to auxin imbalance was studied for the first time. A matrocliny on instability expression was also observed.

  19. Arabidopsis mutants lacking phenolic sunscreens exhibit enhanced ultraviolet-B injury and oxidative damage

    International Nuclear Information System (INIS)

    Landry, L.G.; Last, R.L.; Chapple, C.C.S.

    1995-01-01

    We have assessed ultraviolet-B (UV-B)-induced injury in wild-type Arabidopsis thaliana and two mutants with altered aromatic secondary product biosynthesis. Arabidopsis mutants defective in the ability to synthesize UV-B-absorbing compounds (flavonoids in transparent testa 5 [tt5] and sinapate esters in ferulic acid hydroxylase 1 [fah 1]) are more sensitive to UV-B than is the wild-type Landsberg erecta. Despite its ability to accumulate UV-absorptive flavonoid compounds, the ferulic acid hydroxylase mutant fah1 exhibits more physiological injury (growth inhibition and foliar lesions) than either wild type or tt5. The extreme UV-B sensitivity of fah1 demonstrates the importance of hydroxycinnamate esters as UV-B protectants. Consistent with the whole-plant response, the highest levels of lipid and protein oxidation products were seen in fah1. Ascorbate peroxidase enzyme activity was also increased in the leaves of UV-B-treated plants in a dose- and genotype-dependent manner. These results demonstrate that, in A. thaliana, hydryoxycinnamates are more effective UV-B protectants than flavonoids. The data also indicate that A. thaliana responds to UV-B as an oxidative stress, and sunscreen compounds reduce the oxidative damage caused by UV-B. 36 refs., 6 figs

  20. Melatonin enhances cold tolerance in drought-primed wild-type and abscisic acid-deficient mutant barley.

    Science.gov (United States)

    Li, Xiangnan; Tan, Dun-Xian; Jiang, Dong; Liu, Fulai

    2016-10-01

    Melatonin is involved in multiple plant developmental processes and various stress responses. To explore the roles of melatonin played as well as its association with abscisic acid (ABA) in a process of drought priming-induced cold tolerance (DPICT), a wild-type barley and its ABA-deficient mutant Az34 counterpart were selected for comparison, in which the effects of melatonin application (either foliarly or rhizospherically) and/or drought priming on the cold tolerance of both types of barleys were systematically investigated. It was demonstrated that the early drought priming induced an increase of endogenous melatonin production, which is not ABA dependent. In addition, exogenously applied melatonin resulted in higher ABA concentration in the drought-primed plants than in the nonprimed plants when exposed to cold stress, indicating that ABA responded in a drought-dependent manner. The interplay of melatonin and ABA leads to plants maintaining better water status. Drought priming-induced melatonin accumulation enhanced the antioxidant capacity in both chloroplasts and mitochondria, which sustained the photosynthetic electron transport in photosynthetic apparatus of the plants under cold stress. These results suggest that the exogenous melatonin application enhances the DPICT by modulating subcellular antioxidant systems and ABA levels in barley. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  1. Analysis of Lysophospholipid Content in Low Phytate Rice Mutants.

    Science.gov (United States)

    Tong, Chuan; Chen, Yaling; Tan, Yuanyuan; Liu, Lei; Waters, Daniel L E; Rose, Terry J; Shu, Qingyao; Bao, Jinsong

    2017-07-05

    As a fundamental component of nucleic acids, phospholipids, and adenosine triphosphate, phosphorus (P) is critical to all life forms, however, the molecular mechanism of P translocation and distribution in rice grains are still not understood. Here, with the use of five different low phytic acid (lpa) rice mutants, the redistribution in the main P-containing compounds in rice grain, phytic acid (PA), lysophospholipid (LPL), and inorganic P (Pi), was investigated. The lpa mutants showed a significant decrease in PA and phytate-phosphorus (PA-P) concentration with a concomitant increase in Pi concentration. Moreover, defects in the OsST and OsMIK genes result in a great reduction of specific LPL components and LPL-phosphorus (LPL-P) contents in rice grain. In contrast, defective OsMRP5 and Os2-PGK genes led to a significant increase in individual LPL components. The effect of the Os2-PGK gene on the LPL accumulation was validated using breeding lines derived from a cross between KBNT-lpa (Os2-PGK mutation) and Jiahe218. This study demonstrates that these rice lpa mutants lead to the redistribution of Pi in endosperm and modify LPL biosynthesis. Increase LPLs in the endosperm in the lpa mutants may have practical applications in rice breeding to produce "healthier" rice.

  2. Adaptive response in Drosophila melanogaster heat shock proteins mutant strains

    International Nuclear Information System (INIS)

    Shaposhnikov, M.V.; Moskalev, A.A.; Turysheva, E.V.

    2007-01-01

    Complete text of publication follows. The members of the heat shock proteins (Hsp) family function as molecular chaperones and assist intracellular folding of newly synthesized proteins. Also it is possible that molecular chaperones are induced during adaptive response to oxidative stress and radiation. The aim of our research was to exam the role of heat shock proteins in adaptive response to oxidative stress after low dose rate gamma-irradiation in Drosophila melanogaster. Drosophilamelanogaster strains were kindly provided by Bloomington Drosophila Stock Center (University of state of Indiana, Bloomington, USA). We used wild type strain (CS), heat shock protein mutant strains (Hsp22, Hsp70, Hsp83), and heat shock factor mutant strain (Hsf). Strains were chronically exposured to adaptive dose of gamma-irradiation in dose rate of 0.17 cGy/h during all stages of life history (from the embrional stage to the stage of matured imago). The rate of absorbed dose was 60 cGy. For oxidative-stress challenge twodays old flies were starved in empty vials for 6 h and then transferred to vials containing only filter paper soaked with 20 mM paraquat in 5% sucrose solution. Survival data were collected after 26 h of treatment. Dead flies were counted daily. The obtained data were subjected to survival analysis by Kaplan and Meier method and presented as survival curves. Statistical analysis was held by non-parametric methods. To test the significance of the difference between the two age distributions Kolmogorov-Smirnov test was applied. Gehan-Braslow- Wilcoxon and Cox-Mantel tests were used for estimation of median life span differences. In addition the minimal and maximal life span, time of 90% death, and mortality rate doubling time (MRDT) were estimated. The obtained results will be discussed in presentation.

  3. Arabidopsis GLUTATHIONE REDUCTASE1 plays a crucial role in leaf responses to intracellular hydrogen peroxide and in ensuring appropriate gene expression through both salicylic acid and jasmonic acid signaling pathways.

    Science.gov (United States)

    Mhamdi, Amna; Hager, Jutta; Chaouch, Sejir; Queval, Guillaume; Han, Yi; Taconnat, Ludivine; Saindrenan, Patrick; Gouia, Houda; Issakidis-Bourguet, Emmanuelle; Renou, Jean-Pierre; Noctor, Graham

    2010-07-01

    Glutathione is a major cellular thiol that is maintained in the reduced state by glutathione reductase (GR), which is encoded by two genes in Arabidopsis (Arabidopsis thaliana; GR1 and GR2). This study addressed the role of GR1 in hydrogen peroxide (H(2)O(2)) responses through a combined genetic, transcriptomic, and redox profiling approach. To identify the potential role of changes in glutathione status in H(2)O(2) signaling, gr1 mutants, which show a constitutive increase in oxidized glutathione (GSSG), were compared with a catalase-deficient background (cat2), in which GSSG accumulation is conditionally driven by H(2)O(2). Parallel transcriptomics analysis of gr1 and cat2 identified overlapping gene expression profiles that in both lines were dependent on growth daylength. Overlapping genes included phytohormone-associated genes, in particular implicating glutathione oxidation state in the regulation of jasmonic acid signaling. Direct analysis of H(2)O(2)-glutathione interactions in cat2 gr1 double mutants established that GR1-dependent glutathione status is required for multiple responses to increased H(2)O(2) availability, including limitation of lesion formation, accumulation of salicylic acid, induction of pathogenesis-related genes, and signaling through jasmonic acid pathways. Modulation of these responses in cat2 gr1 was linked to dramatic GSSG accumulation and modified expression of specific glutaredoxins and glutathione S-transferases, but there is little or no evidence of generalized oxidative stress or changes in thioredoxin-associated gene expression. We conclude that GR1 plays a crucial role in daylength-dependent redox signaling and that this function cannot be replaced by the second Arabidopsis GR gene or by thiol systems such as the thioredoxin system.

  4. Application of gamma rays for induction of tolerance mutants to environmental stress conditions in canola

    International Nuclear Information System (INIS)

    Mansour, M.E.S.F.

    2013-01-01

    The present study aimed to induce useful mutations in canola possess high seed yield and oil content under new reclamation desert land at Ras-Suder-Sina (saline) and Inshas (harsh and poor fertility). Canola seeds of four varieties (Serow 4, Serow 6, Pactol as local cultivars and Evita as exotic variety) were treated with gamma rays at four doses (0, 100, 400 and 600 Gy). Thirty mutant plants for number of pods/plant and changes in morphological criteria were selected at M 2 generation. The mutants at M 3 generation confirmed that induction of mutant lines possessed higher number of pods and seed yield/plant than the mother varieties. The mutant lines possessed homogeneity at M 3 generation were 5, 8,10, 11, 18 and 22 at serow 4, 38 and 45 at serow 6, 63 and 66 at Pactol and mutant lines 74,75, 78,92 at Evita. Highest number of pods/plant (110) was recorded at line 74 derived from Evita variety. The results were appeared the same trend for seed yield/plant with number of pods/plant, the lines which possessed high number of pods/plant were had high seed yield/plant. The results at M 4 and M 5 generations for 13 homogeneity mutant lines selected from M 3 generation contained different response of mutant genotypes for different conditions on the bases of number of pods and seed yield/plant. Promising mutant lines were detected under both conditions possessed significant increases at both M 4 and M 5 generations. Oil percent as well as acid value at M 4 and M 5 were recorded the highest mean value was found at Inshas in line 75 and the lowest acid value was noticed at line 5. Finally nine mutant lines possessed promising traits of this study, lines 11, 66 and 87 under both conditions (Suder and Inshas), lines 8, 38 and 63 under Ras-Sudr and lines 74, 75 and 92 under Inshas condition.

  5. Optimization of L(+)-Lactic Acid Fermentation Without Neutralisation of Rhizopus Oryzae Mutant RK02 by Low-Energy Ion Implantation

    International Nuclear Information System (INIS)

    Li Wen; Wang Tao; Yang Yingge; Liu Dan; Fan Yonghong; Wang Dongmei; Yang Qian; Yao Jianming; Zheng Zhiming; Yu Zengliang

    2008-01-01

    In order to get an industrial strain which can yield a high concentration of lactic acid for ISPR (in situ product removal), the original strain Rhizopus oryzae RE3303 was mutated by low-energy ion beam implantation. A mutant RK02 was screened, and the factors such as the substrate concentration, nitrogen source concentration, inoculum size, seed age, aeration and temperature that affect the production of lactic acid were studied in detail. Under optimal conditions, the maximum concentration of L(+)-lactic acid reached 34.85 g/L after 30 h shake-flask cultivation without adding any neutralisation (5% Glucose added), which was a 146% increase in lactic acid production after ion implantation compared with the original strain. It was also shown that RK02 can be used in ISPR to reduce the number of times of separation.

  6. Isolation and characterization of Candida albicans morphological mutants derepressed for the formation of filamentous hypha-type structures

    International Nuclear Information System (INIS)

    Gil, C.; Pomes, R.; Nombela, C.

    1990-01-01

    Several Candida albicans morphological mutants were obtained by a procedure based on a combined treatment with nitrous acid plus UV irradiation and a double-enrichment step to increase the proportion of mutants growing as long filamentous structures. Altered cell morphogenesis in these mutants correlated with an altered colonial phenotype. Two of these mutants, C. albicans NEL102 and NEL103, were selected and characterized. Mutant blastoconidia initiated budding but eventually gave rise to filamentous hypha-type formations. These filaments were long and septate, and they branched very regularly at positions near septa. Calcofluor white (which is known to bind chitin-rich areas) stained septa, branching zones, and filament tips very intensely, as observed under the fluorescence microscope. Wild-type hybrids were obtained by fusing protoplasts of strain NEL102 with B14, another morphological mutant previously described as being permanently pseudomycelial, indicating that genetic determinants responsible for the two altered phenotypes are different. The mutants characterized in this work seemed to sequentially express the morphogenic characteristics of C. albicans, from blastoconidia to hyphae, in the absence of any inducer. Further characterization of these strains could be relevant to gain understanding of the genetic control of dimorphism in this species

  7. An UPLC-ESI-MS/MS Assay Using 6-Aminoquinolyl-N-Hydroxysuccinimidyl Carbamate Derivatization for Targeted Amino Acid Analysis: Application to Screening of Arabidopsis thaliana Mutants

    Directory of Open Access Journals (Sweden)

    Carolina Salazar

    2012-07-01

    Full Text Available In spite of the large arsenal of methodologies developed for amino acid assessment in complex matrices, their implementation in metabolomics studies involving wide-ranging mutant screening is hampered by their lack of high-throughput, sensitivity, reproducibility, and/or wide dynamic range. In response to the challenge of developing amino acid analysis methods that satisfy the criteria required for metabolomic studies, improved reverse-phase high-performance liquid chromatography-mass spectrometry (RPHPLC-MS methods have been recently reported for large-scale screening of metabolic phenotypes. However, these methods focus on the direct analysis of underivatized amino acids and, therefore, problems associated with insufficient retention and resolution are observed due to the hydrophilic nature of amino acids. It is well known that derivatization methods render amino acids more amenable for reverse phase chromatographic analysis by introducing highly-hydrophobic tags in their carboxylic acid or amino functional group. Therefore, an analytical platform that combines the 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC pre-column derivatization method with ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS is presented in this article. For numerous reasons typical amino acid derivatization methods would be inadequate for large scale metabolic projects. However, AQC derivatization is a simple, rapid and reproducible way of obtaining stable amino acid adducts amenable for UPLC-ESI-MS/MS and the applicability of the method for high-throughput metabolomic analysis in Arabidopsis thaliana is demonstrated in this study. Overall, the major advantages offered by this amino acid analysis method include high-throughput, enhanced sensitivity and selectivity; characteristics that showcase its utility for the rapid screening of the preselected plant metabolites without compromising the quality of the

  8. An UPLC-ESI-MS/MS Assay Using 6-Aminoquinolyl-N-Hydroxysuccinimidyl Carbamate Derivatization for Targeted Amino Acid Analysis: Application to Screening of Arabidopsis thaliana Mutants.

    Science.gov (United States)

    Salazar, Carolina; Armenta, Jenny M; Shulaev, Vladimir

    2012-07-06

    In spite of the large arsenal of methodologies developed for amino acid assessment in complex matrices, their implementation in metabolomics studies involving wide-ranging mutant screening is hampered by their lack of high-throughput, sensitivity, reproducibility, and/or wide dynamic range. In response to the challenge of developing amino acid analysis methods that satisfy the criteria required for metabolomic studies, improved reverse-phase high-performance liquid chromatography-mass spectrometry (RPHPLC-MS) methods have been recently reported for large-scale screening of metabolic phenotypes. However, these methods focus on the direct analysis of underivatized amino acids and, therefore, problems associated with insufficient retention and resolution are observed due to the hydrophilic nature of amino acids. It is well known that derivatization methods render amino acids more amenable for reverse phase chromatographic analysis by introducing highly-hydrophobic tags in their carboxylic acid or amino functional group. Therefore, an analytical platform that combines the 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) pre-column derivatization method with ultra performance liquid chromatography-electrospray ionization-tandem mass spectrometry (UPLC-ESI-MS/MS) is presented in this article. For numerous reasons typical amino acid derivatization methods would be inadequate for large scale metabolic projects. However, AQC derivatization is a simple, rapid and reproducible way of obtaining stable amino acid adducts amenable for UPLC-ESI-MS/MS and the applicability of the method for high-throughput metabolomic analysis in Arabidopsis thaliana is demonstrated in this study. Overall, the major advantages offered by this amino acid analysis method include high-throughput, enhanced sensitivity and selectivity; characteristics that showcase its utility for the rapid screening of the preselected plant metabolites without compromising the quality of the metabolic data. The

  9. Integrative proteomics, genomics, and translational immunology approaches reveal mutated forms of Proteolipid Protein 1 (PLP1) and mutant-specific immune response in multiple sclerosis.

    Science.gov (United States)

    Qendro, Veneta; Bugos, Grace A; Lundgren, Debbie H; Glynn, John; Han, May H; Han, David K

    2017-03-01

    In order to gain mechanistic insights into multiple sclerosis (MS) pathogenesis, we utilized a multi-dimensional approach to test the hypothesis that mutations in myelin proteins lead to immune activation and central nervous system autoimmunity in MS. Mass spectrometry-based proteomic analysis of human MS brain lesions revealed seven unique mutations of PLP1; a key myelin protein that is known to be destroyed in MS. Surprisingly, in-depth genomic analysis of two MS patients at the genomic DNA and mRNA confirmed mutated PLP1 in RNA, but not in the genomic DNA. Quantification of wild type and mutant PLP RNA levels by qPCR further validated the presence of mutant PLP RNA in the MS patients. To seek evidence linking mutations in abundant myelin proteins and immune-mediated destruction of myelin, specific immune response against mutant PLP1 in MS patients was examined. Thus, we have designed paired, wild type and mutant peptide microarrays, and examined antibody response to multiple mutated PLP1 in sera from MS patients. Consistent with the idea of different patients exhibiting unique mutation profiles, we found that 13 out of 20 MS patients showed antibody responses against specific but not against all the mutant-PLP1 peptides. Interestingly, we found mutant PLP-directed antibody response against specific mutant peptides in the sera of pre-MS controls. The results from integrative proteomic, genomic, and immune analyses reveal a possible mechanism of mutation-driven pathogenesis in human MS. The study also highlights the need for integrative genomic and proteomic analyses for uncovering pathogenic mechanisms of human diseases. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Applications of Protein Thermodynamic Database for Understanding Protein Mutant Stability and Designing Stable Mutants.

    Science.gov (United States)

    Gromiha, M Michael; Anoosha, P; Huang, Liang-Tsung

    2016-01-01

    Protein stability is the free energy difference between unfolded and folded states of a protein, which lies in the range of 5-25 kcal/mol. Experimentally, protein stability is measured with circular dichroism, differential scanning calorimetry, and fluorescence spectroscopy using thermal and denaturant denaturation methods. These experimental data have been accumulated in the form of a database, ProTherm, thermodynamic database for proteins and mutants. It also contains sequence and structure information of a protein, experimental methods and conditions, and literature information. Different features such as search, display, and sorting options and visualization tools have been incorporated in the database. ProTherm is a valuable resource for understanding/predicting the stability of proteins and it can be accessed at http://www.abren.net/protherm/ . ProTherm has been effectively used to examine the relationship among thermodynamics, structure, and function of proteins. We describe the recent progress on the development of methods for understanding/predicting protein stability, such as (1) general trends on mutational effects on stability, (2) relationship between the stability of protein mutants and amino acid properties, (3) applications of protein three-dimensional structures for predicting their stability upon point mutations, (4) prediction of protein stability upon single mutations from amino acid sequence, and (5) prediction methods for addressing double mutants. A list of online resources for predicting has also been provided.

  11. Partial diversion of a mutant proinsulin (B10 aspartic acid) from the regulated to the constitutive secretory pathway in transfected AtT-20 cells.

    OpenAIRE

    Gross, D J; Halban, P A; Kahn, C R; Weir, G C; Villa-Komaroff, L

    1989-01-01

    A patient with type II diabetes associated with hyperproinsulinemia has been shown to have a point mutation in one insulin gene allele, resulting in replacement of histidine with aspartic acid at position 10 of the B-chain. To investigate the basis of the proinsulin processing defect, we introduced an identical mutation in the rat insulin II gene and expressed both the normal and the mutant genes in the AtT-20 pituitary corticotroph cell line. Cells expressing the mutant gene showed increased...

  12. Nicotiana plumbaginifolia hlg mutants have a mutation in a PHYB-type phytochrome gene: they have elongated hypocotyls in red light, but are not elongated as adult plants.

    Science.gov (United States)

    Hudson, M; Robson, P R; Kraepiel, Y; Caboche, M; Smith, H

    1997-11-01

    Two new allelic mutants of Nicotiana plumbaginifolia have been isolated which display a hypocotyl which is long (hlg) when seedlings are grown in continuous white light (W). This can be accounted for by the decreased response to red light (R) of the hypocotyl elongation rate in these mutants. Responses to other wavelengths are unaffected in the mutants. When grown in white light, mature hlg mutants are not elongated with respect to the wild-type; they also bolt and flower later. The shade-avoidance responses to red/far red ratio (R:FR) are intact in these mutants. Both mutants are deficient in phyB-like polypeptide that is immunodetectable in the wild-type; both have wild-type levels of a phyA-like polypeptide. These alleles are inherited in a partially dominant manner, and correspond to single-base missense mutations in a gene highly homologous to N. tabacum PHYB, which codes for a phytochrome B-type photoreceptor. One allele, hlg-1, has an introduced amino acid substitution; this may define a residue essential for phytochrome protein stability. The other allele, hlg-2, has a stop codon introduced C-terminal to the chromophore binding domain. As these phyB mutants are unaffected in shade-avoidance responses, but deficient in perception of R, it is concluded that the phyB absent in these mutants is responsible for R perception in the N. plumbaginifolia seedling, but is not a R:FR sensor in light-grown plants.

  13. R Factor-Controlled Restriction and Modification of Deoxyribonucleic Acid: Restriction Mutants

    Science.gov (United States)

    Yoshimori, Robert; Roulland-Dussoix, Daisy; Boyer, Herbert W.

    1972-01-01

    Restriction mutants of two different R factor-controlled host specificities (RI and RII) were isolated. All of the restriction mutants examined had a normal modification phenotype. No complementation was observed between the RI and RII host specificities. It is concluded that for each host specificity no protein subunit is shared by the restriction endonuclease and modification methylase. PMID:4565538

  14. Genetic analysis of diaminopimelic acid- and lysine-requiring mutants of Escherichia coli.

    Science.gov (United States)

    Bukhari, A I; Taylor, A L

    1971-03-01

    Several diaminopimelic acid (DAP)- and lysine-requiring mutants of Escherichia coli were isolated and studied by genetic, physiological, and biochemical means. The genes concerned with DAP-lysine synthesis map at several different sites on the E. coli chromosome and, therefore, do not constitute a single operon. Three separate loci affecting DAP synthesis are located in the 0 to 2.5 min region of the genetic map. The order of the loci in this region is thr-dapB-pyrA-ara-leu-pan-dapC-tonA-dapD. Two additional DAP genes map in the region between min 47 and 48, with the gene order being gua-dapA-dapE-ctr. The lys locus at min 55 determines the synthesis of the enzyme DAP decarboxylase, which catalyzes the conversion of DAP into lysine. The order of the genes in this region is serA-lysA-thyA.

  15. Quality characteristics of soybean pasted (Doenjang) manufactured with 2 soybean mutant lines derived from cv. baekwon

    International Nuclear Information System (INIS)

    Lee, Kyung Jun; Kang, Si Yong; Choi, Hong Il; Kim, Jin Baek

    2016-01-01

    In order to identification of the possibility of manufacturing soybean paste (doenjang) with soybean mutant lines induced from gamma-ray mutagenesis, this study was performed to investigate the quality characteristics of doenjang using two soybean mutant lines, Baekwon-1 (BW-1) and Baekwon-2 (BW-2) and their original cultivar (cv. Baekwon, BW) for 8 weeks. The BW and two mutant lines (BW-1 and BW-2) were showed higher content of amino type nitrogen than control (cv. Taegwang). The pH decreased and the titratable acidity increased all the samples during aging period. The lightness, redness and yellowness of doenjang were the lowest in BW. Total free sugar content of doenjang was the highest in control (10.43%) after 4 weeks and composed mainly fructose and glucose. The order of the free amino acid content was Glutamic acid>Leucine>Lysine>Phenylalanine>Aspartic acid in control, Glutamic acid>Leucine >Arginine>Lysine>Phenylalanine in BW, Glutamic acid>Lysine>Phenylalanine>Aspartic acid>Valine in BW-1 and Glutamic acid>Arginine>Lysine>Phenylalanine>Aspartic acid in BW-2, respectively. Our results showed that it is possible to increase the quality of doenjang using soybean mutant lines in manufacturing soybean paste

  16. Quality characteristics of soybean pasted (Doenjang) manufactured with 2 soybean mutant lines derived from cv. baekwon

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Kyung Jun; Kang, Si Yong; Choi, Hong Il; Kim, Jin Baek [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup (Korea, Republic of)

    2016-11-15

    In order to identification of the possibility of manufacturing soybean paste (doenjang) with soybean mutant lines induced from gamma-ray mutagenesis, this study was performed to investigate the quality characteristics of doenjang using two soybean mutant lines, Baekwon-1 (BW-1) and Baekwon-2 (BW-2) and their original cultivar (cv. Baekwon, BW) for 8 weeks. The BW and two mutant lines (BW-1 and BW-2) were showed higher content of amino type nitrogen than control (cv. Taegwang). The pH decreased and the titratable acidity increased all the samples during aging period. The lightness, redness and yellowness of doenjang were the lowest in BW. Total free sugar content of doenjang was the highest in control (10.43%) after 4 weeks and composed mainly fructose and glucose. The order of the free amino acid content was Glutamic acid>Leucine>Lysine>Phenylalanine>Aspartic acid in control, Glutamic acid>Leucine >Arginine>Lysine>Phenylalanine in BW, Glutamic acid>Lysine>Phenylalanine>Aspartic acid>Valine in BW-1 and Glutamic acid>Arginine>Lysine>Phenylalanine>Aspartic acid in BW-2, respectively. Our results showed that it is possible to increase the quality of doenjang using soybean mutant lines in manufacturing soybean paste.

  17. Enhanced L-lactic acid production from biomass-derived xylose by a mutant Bacillus coagulans.

    Science.gov (United States)

    Zheng, Zhaojuan; Cai, Cong; Jiang, Ting; Zhao, Mingyue; Ouyang, Jia

    2014-08-01

    Xylose effective utilization is crucial for production of bulk chemicals from low-cost lignocellulosic substrates. In this study, an efficient L-lactate production process from xylose by a mutant Bacillus coagulans NL-CC-17 was demonstrated. The nutritional requirements for L-lactate production by B. coagulans NL-CC-17 were optimized statistically in shake flask fermentations. Corn steep liquor powder and yeast exact were identified as the most significant factors by the two-level Plackett-Burman design. Steepest ascent experiments were applied to approach the optimal region of the two factors, and a central composite design was employed to determine their optimal levels. The optimal medium was used to perform batch fermentation in a 3-l bioreactor. A maximum of 90.29 g l(-1)  L-lactic acid was obtained from 100 g l(-1) xylose in 120 h. When using corn stove prehydrolysates as substrates, 23.49 g l(-1)  L-lactic acid was obtained in 36 h and the yield was 83.09 %.

  18. Characterization of a novel curled-cotyledons mutant in soybean ...

    African Journals Online (AJOL)

    ARL

    different stages. Protein and amino acid contents in seeds of mutant are higher than those of the wild .... curled shape, from seedling to maturation phase (Figure. 1D-1F). ..... Arabidopsis seed dermination by stimulating abscisic acid synthesis.

  19. Reduced de-etiolation of hypocotyl growth in a tomato mutant is associated with hypersensitivity to, and high endogenous levels of, abscisic acid

    Czech Academy of Sciences Publication Activity Database

    Fellner, Martin; Zhang, R.; Pharis, R.; Sawhney, V.

    2001-01-01

    Roč. 52, č. 357 (2001), s. 725-738 ISSN 0022-0957 R&D Projects: GA ČR GV521/96/K117 Institutional research plan: CEZ:AV0Z5038910 Keywords : Abscisic acid * elongated mutant * fluridone Subject RIV: EF - Botanics Impact factor: 2.433, year: 2001

  20. Structural analysis of site-directed mutants of cellular retinoic acid-binding protein II addresses the relationship between structural integrity and ligand binding

    International Nuclear Information System (INIS)

    Vaezeslami, Soheila; Jia, Xiaofei; Vasileiou, Chrysoula; Borhan, Babak; Geiger, James H.

    2008-01-01

    A water network stabilizes the structure of cellular retionic acid binding protein II. The structural integrity of cellular retinoic acid-binding protein II (CRABPII) has been investigated using the crystal structures of CRABPII mutants. The overall fold was well maintained by these CRABPII mutants, each of which carried multiple different mutations. A water-mediated network is found to be present across the large binding cavity, extending from Arg111 deep inside the cavity to the α2 helix at its entrance. This chain of interactions acts as a ‘pillar’ that maintains the integrity of the protein. The disruption of the water network upon loss of Arg111 leads to decreased structural integrity of the protein. A water-mediated network can be re-established by introducing the hydrophilic Glu121 inside the cavity, which results in a rigid protein with the α2 helix adopting an altered conformation compared with wild-type CRABPII

  1. High lysine and high yielding mutants in wheat (Triticum aestivum) L

    International Nuclear Information System (INIS)

    Mohammad, T.; Mahmood, F.; Ahmad, A.; Sattar, A.; Khan, I.

    1988-01-01

    The dry seeds of the variety Lu-26 were irradiated with 20 krad of gamma rays. In M 2 about 300 mutant plants were selected for short stature, rust resistance and other desirable traits. As a result of further selection, in M 6 , eight superior lines were finally identified. The agronomic characteristics of these mutants, the parent variety (Lu-26) and a standard check variety (Pak-81) are shown. The selected mutants and commercially grown cultivars (Lu-26 and Pak-81) were studied for total protein content and amino acid pattern. The mutants WM-89-1, WM-6-17 and WM-81-2 showing high yield also contained comparatively high amounts of methionine and lysine. The lysine contents were 565, 410, and 370 mg/100g in the mutants WM-89-1, WM-6-17 and WM-81-2, respectively against a range value of 210-370 mg/100g in other mutants and 250-320 in the commercial cultivars. The mutant WM-81-2 was comparable to WM-56-1-5 in lysine content. The results of these experiments show a possibility of developing varieties having high yield and high amounts of essential amino acids such as lysine and methionine

  2. Ethylene Responses in Rice Roots and Coleoptiles Are Differentially Regulated by a Carotenoid Isomerase-Mediated Abscisic Acid Pathway[OPEN

    Science.gov (United States)

    Yin, Cui-Cui; Ma, Biao; Collinge, Derek Phillip; Pogson, Barry James; He, Si-Jie; Xiong, Qing; Duan, Kai-Xuan; Chen, Hui; Yang, Chao; Lu, Xiang; Wang, Yi-Qin; Zhang, Wan-Ke; Chu, Cheng-Cai; Sun, Xiao-Hong; Fang, Shuang; Chu, Jin-Fang; Lu, Tie-Gang; Chen, Shou-Yi; Zhang, Jin-Song

    2015-01-01

    Ethylene and abscisic acid (ABA) act synergistically or antagonistically to regulate plant growth and development. ABA is derived from the carotenoid biosynthesis pathway. Here, we analyzed the interplay among ethylene, carotenoid biogenesis, and ABA in rice (Oryza sativa) using the rice ethylene response mutant mhz5, which displays a reduced ethylene response in roots but an enhanced ethylene response in coleoptiles. We found that MHZ5 encodes a carotenoid isomerase and that the mutation in mhz5 blocks carotenoid biosynthesis, reduces ABA accumulation, and promotes ethylene production in etiolated seedlings. ABA can largely rescue the ethylene response of the mhz5 mutant. Ethylene induces MHZ5 expression, the production of neoxanthin, an ABA biosynthesis precursor, and ABA accumulation in roots. MHZ5 overexpression results in enhanced ethylene sensitivity in roots and reduced ethylene sensitivity in coleoptiles. Mutation or overexpression of MHZ5 also alters the expression of ethylene-responsive genes. Genetic studies revealed that the MHZ5-mediated ABA pathway acts downstream of ethylene signaling to inhibit root growth. The MHZ5-mediated ABA pathway likely acts upstream but negatively regulates ethylene signaling to control coleoptile growth. Our study reveals novel interactions among ethylene, carotenogenesis, and ABA and provides insight into improvements in agronomic traits and adaptive growth through the manipulation of these pathways in rice. PMID:25841037

  3. Mutation of a vitelline membrane protein, BmEP80, is responsible for the silkworm "Ming" lethal egg mutant.

    Science.gov (United States)

    Chen, Anli; Gao, Peng; Zhao, Qiaoling; Tang, Shunming; Shen, Xingjia; Zhang, Guozheng; Qiu, Zhiyong; Xia, Dingguo; Huang, Yongping; Xu, Yunmin; He, Ningjia

    2013-02-25

    The egg stage is an important stage in the silkworm (Bombyx mori) life cycle. Normal silkworm eggs are usually short, elliptical, and laterally flattened, with a sometimes hollowed surface on the lateral side. However, the eggs laid by homozygous recessive "Ming" lethal egg mutants (l-e(m)) lose water and become concaved around 1h, ultimately exhibiting a triangular shape on the egg surfaces. We performed positional cloning, and narrowed down the region containing the gene responsible for the l-e(m) mutant to 360 kb on chromosome 10 using 2287 F(2) individuals. Using expression analysis and RNA interference, the best l-e(m) candidate gene was shown to be BmEP80. The results of the inverse polymerase chain reaction showed that an ~1.9 kb region from the 3' untranslated region of BmVMP23 to the forepart of BmEP80 was replaced by a >100 kb DNA fragment in the l-e(m) mutant. Several eggs laid by the normal moths injected with BmEP80 small interfering RNAs were evidently depressed and exhibited a triangular shape on the surface. The phenotype exhibited was consistent with the eggs laid by the l-e(m) mutant. Moreover, two-dimensional gel electrophoresis showed that the BmEP80 protein was expressed in the ovary from the 9th day of the pupa stage to eclosion in the wild-type silkworm, but was absent in the l-e(m) mutant. These results indicate that BmEP80 is responsible for the l-e(m) mutation. Copyright © 2012 Elsevier B.V. All rights reserved.

  4. MMS sensitivity of all amino acid-requiring mutants in aspergillus and its suppression by mutations in a single gene.

    Science.gov (United States)

    Käfer, E

    1987-04-01

    All available amino acid-requiring mutants of Aspergillus nidulans were found to be hypersensitive to MMS (methyl methanesulfonate) to various degrees. On MMS media, secondary mutations could be selected which suppress this MMS sensitivity but do not affect the requirement. Many such mutations were analyzed and found to be alleles of one gene, smsA (= suppressor of MMS sensitivity), which mapped distal on the right arm of chromosome V. This gene is more likely to be involved in general regulation of amino acid biosynthesis than MMS uptake, since a variety of pathway interactions were clearly modified by smsA suppressors in the absence of MMS.

  5. Arabidopsis plastid AMOS1/EGY1 integrates abscisic acid signaling to regulate global gene expression response to ammonium stress

    KAUST Repository

    Li, Baohai

    2012-10-12

    Ammonium (NH4 +) is a ubiquitous intermediate of nitrogen metabolism but is notorious for its toxic effects on most organisms. Extensive studies of the underlying mechanisms of NH4 + toxicity have been reported in plants, but it is poorly understood how plants acclimate to high levels of NH4 +. Here, we identified an Arabidopsis (Arabidopsis thaliana) mutant, ammonium overly sensitive1 (amos1), that displays severe chlorosis under NH4 + stress. Map-based cloning shows amos1 to carry a mutation in EGY1 (for ethylene-dependent, gravitropism-deficient, and yellow-green-like protein1), which encodes a plastid metalloprotease. Transcriptomic analysis reveals that among the genes activated in response to NH4 +, 90% are regulated dependent on AMOS1/ EGY1. Furthermore, 63% of AMOS1/EGY1-dependent NH4 +-activated genes contain an ACGTG motif in their promoter region, a core motif of abscisic acid (ABA)-responsive elements. Consistent with this, our physiological, pharmacological, transcriptomic, and genetic data show that ABA signaling is a critical, but not the sole, downstream component of the AMOS1/EGY1-dependent pathway that regulates the expression of NH4 +-responsive genes and maintains chloroplast functionality under NH4 + stress. Importantly, abi4 mutants defective in ABA-dependent and retrograde signaling, but not ABA-deficient mutants, mimic leaf NH4 + hypersensitivity of amos1. In summary, our findings suggest that an NH4 +-responsive plastid retrograde pathway, which depends on AMOS1/EGY1 function and integrates with ABA signaling, is required for the regulation of expression of the presence of high NH4 + levels. © 2012 American Society of Plant Biologists. All Rights Reserved.

  6. Arabidopsis plastid AMOS1/EGY1 integrates abscisic acid signaling to regulate global gene expression response to ammonium stress

    KAUST Repository

    Li, Baohai; Li, Qing; Xiong, Liming; Kronzucker, Herbert J.; Krä mer, Ute; Shi, Weiming

    2012-01-01

    Ammonium (NH4 +) is a ubiquitous intermediate of nitrogen metabolism but is notorious for its toxic effects on most organisms. Extensive studies of the underlying mechanisms of NH4 + toxicity have been reported in plants, but it is poorly understood how plants acclimate to high levels of NH4 +. Here, we identified an Arabidopsis (Arabidopsis thaliana) mutant, ammonium overly sensitive1 (amos1), that displays severe chlorosis under NH4 + stress. Map-based cloning shows amos1 to carry a mutation in EGY1 (for ethylene-dependent, gravitropism-deficient, and yellow-green-like protein1), which encodes a plastid metalloprotease. Transcriptomic analysis reveals that among the genes activated in response to NH4 +, 90% are regulated dependent on AMOS1/ EGY1. Furthermore, 63% of AMOS1/EGY1-dependent NH4 +-activated genes contain an ACGTG motif in their promoter region, a core motif of abscisic acid (ABA)-responsive elements. Consistent with this, our physiological, pharmacological, transcriptomic, and genetic data show that ABA signaling is a critical, but not the sole, downstream component of the AMOS1/EGY1-dependent pathway that regulates the expression of NH4 +-responsive genes and maintains chloroplast functionality under NH4 + stress. Importantly, abi4 mutants defective in ABA-dependent and retrograde signaling, but not ABA-deficient mutants, mimic leaf NH4 + hypersensitivity of amos1. In summary, our findings suggest that an NH4 +-responsive plastid retrograde pathway, which depends on AMOS1/EGY1 function and integrates with ABA signaling, is required for the regulation of expression of the presence of high NH4 + levels. © 2012 American Society of Plant Biologists. All Rights Reserved.

  7. Studies on reduced height mutants in rice

    International Nuclear Information System (INIS)

    Narahari, P.; Bhagwat, S.G.

    1984-01-01

    Two cross-bred derivatives of the mutant TR5xTR17 and TR21 continued to show promise and were advanced to wider scale testing. TR5 was found to carry a semi-dwarfing gene different from that in IR8. New semi-dwarf mutants were screened from M 2 through M 4 from two separate radiation experiments. The gibberellin response of seedlings of mutant and tester strains was evaluated and crosses of tester stocks and mutant semi-dwarfs were made for genetic analyses. (author)

  8. Chemical rescue of the post-translationally carboxylated lysine mutant of allantoinase and dihydroorotase by metal ions and short-chain carboxylic acids.

    Science.gov (United States)

    Ho, Ya-Yeh; Huang, Yen-Hua; Huang, Cheng-Yang

    2013-04-01

    Bacterial allantoinase (ALLase) and dihydroorotase (DHOase) are members of the cyclic amidohydrolase family. ALLase and DHOase possess similar binuclear metal centers in the active site in which two metals are bridged by a post-translationally carboxylated lysine. In this study, we determined the effects of carboxylated lysine and metal binding on the activities of ALLase and DHOase. Although DHOase is a metalloenzyme, purified DHOase showed high activity without additional metal supplementation in a reaction mixture or bacterial culture. However, unlike DHOase, ALLase had no activity unless some specific metal ions were added to the reaction mixture or culture. Substituting the metal binding sites H59, H61, K146, H186, H242, or D315 with alanine completely abolished the activity of ALLase. However, the K146C, K146D and K146E mutants of ALLase were still active with about 1-6% activity of the wild-type enzyme. These ALLase K146 mutants were found to have 1.4-1.7 mol metal per mole enzyme subunit, which may indicate that they still contained the binuclear metal center in the active site. The activity of the K146A mutant of the ALLase and the K103A mutant of DHOase can be chemically rescued by short-chain carboxylic acids, such as acetic, propionic, and butyric acids, but not by ethanol, propan-1-ol, and imidazole, in the presence of Co2+ or Mn2+ ions. However, the activity was still ~10-fold less than that of wild-type ALLase. Overall, these results indicated that the 20 natural basic amino acid residues were not sufficiently able to play the role of lysine. Accordingly, we proposed that during evolution, the post-translational modification of carboxylated lysine in the cyclic amidohydrolase family was selected for promoting binuclear metal center self-assembly and increasing the nucleophilicity of the hydroxide at the active site for enzyme catalysis. This kind of chemical rescue combined with site-directed mutagenesis may also be used to identify a binuclear metal

  9. Transcriptional Regulation of Arabidopsis MIR168a and ARGONAUTE1 Homeostasis in Abscisic Acid and Abiotic Stress Responses1[W

    Science.gov (United States)

    Li, Wei; Cui, Xiao; Meng, Zhaolu; Huang, Xiahe; Xie, Qi; Wu, Heng; Jin, Hailing; Zhang, Dabing; Liang, Wanqi

    2012-01-01

    The accumulation of a number of small RNAs in plants is affected by abscisic acid (ABA) and abiotic stresses, but the underlying mechanisms are poorly understood. The miR168-mediated feedback regulatory loop regulates ARGONAUTE1 (AGO1) homeostasis, which is crucial for gene expression modulation and plant development. Here, we reveal a transcriptional regulatory mechanism by which MIR168 controls AGO1 homeostasis during ABA treatment and abiotic stress responses in Arabidopsis (Arabidopsis thaliana). Plants overexpressing MIR168a and the AGO1 loss-of-function mutant ago1-27 display ABA hypersensitivity and drought tolerance, while the mir168a-2 mutant shows ABA hyposensitivity and drought hypersensitivity. Both the precursor and mature miR168 were induced under ABA and several abiotic stress treatments, but no obvious decrease for the target of miR168, AGO1, was shown under the same conditions. However, promoter activity analysis indicated that AGO1 transcription activity was increased under ABA and drought treatments, suggesting that transcriptional elevation of MIR168a is required for maintaining a stable AGO1 transcript level during the stress response. Furthermore, we showed both in vitro and in vivo that the transcription of MIR168a is directly regulated by four abscisic acid-responsive element (ABRE) binding factors, which bind to the ABRE cis-element within the MIR168a promoter. This ABRE motif is also found in the promoter of MIR168a homologs in diverse plant species. Our findings suggest that transcriptional regulation of miR168 and posttranscriptional control of AGO1 homeostasis may play an important and conserved role in stress response and signal transduction in plants. PMID:22247272

  10. The pht4;1-3 mutant line contains a loss of function allele in the Fatty Acid Desaturase 7 gene caused by a remnant inactivated selection marker-a cautionary tale.

    Science.gov (United States)

    Nilsson, Anders K; Andersson, Mats X

    2017-01-01

    A striking and unexpected biochemical phenotype was found in an insertion mutant line in the model plant Arabidopsis thaliana . One of two investigated insertion mutant lines in the gene encoding the phosphate transporter PHT4;1 demonstrated a prominent loss of trienoic fatty acids, whereas the other insertion line was indistinguishable from wild type in this aspect. We demonstrate that the loss of trienoic fatty acids was due to a remnant inactive negative selection marker gene in this particular transposon tagged line, pht4;1-3 . This constitutes a cautionary tale that warns of the importance to confirm the loss of this type of selection markers and the importance of verifying the relationship between a phenotype and genotype by more than one independent mutant line or alternatively genetic complementation.

  11. Saccharomyces cerevisiae mutants with enhanced induced mutation and altered mitotic gene conversion.

    Science.gov (United States)

    Ivanov, E L; Kovaltzova, S V; Korolev, V G

    1989-08-01

    We have developed a method to isolate yeast (Saccharomyces cerevisiae) mutants with enhanced induced mutagenesis based on nitrous acid-induced reversion of the ade2-42 allele. Six mutants have been isolated and designated him (high induced mutagenesis), and 4 of them were studied in more detail. The him mutants displayed enhanced reversion of the ade2-42 allele, either spontaneous or induced by nitrous acid, UV light, and the base analog 6-N-hydroxylaminopurine, but not by gamma-irradiation. It is worth noting that the him mutants turned out not to be sensitive to the lethal effects of the mutagens used. The enhancement in mutation induced by nitrous acid, UV light, and 6-N-hydroxylaminopurine has been confirmed in a forward-mutation assay (induction of mutations in the ADE1, ADE2 genes). The latter agent revealed the most apparent differences between the him mutants and the wild-type strain and was, therefore, chosen for the genetic analysis of mutants, him mutations analyzed behaved as a single Mendelian trait; complementation tests indicated 3 complementation groups (HIM1, HIM2, and HIM3), each containing 1 mutant allele. Uracil-DNA glycosylase activity was determined in crude cell extracts, and no significant differences between the wild-type and him strains were detected. Spontaneous mitotic gene conversion at the ADE2 locus is altered in him1 strains, either increased or decreased, depending on the particular heteroallelic combination. Genetic evidence strongly suggests him mutations to be involved in a process of mismatch correction of molecular heteroduplexes.

  12. Seed protein and nitrogen fixation in chickpea mutant variety Hyprosola

    Energy Technology Data Exchange (ETDEWEB)

    Schroeder, H E; Gibson, A H; Oram, R N [CSIRO, Division of Plant Industry, Canberra ACT (Australia); Shaikh, M A.Q. [Bangladesh Institute of Nuclear Agriculture, Mymensingh (Bangladesh)

    1989-01-01

    Full text: 'Hyprosola' is a high yielding, high protein mutant cultivar obtained after gamma irradiation from the variety 'Faridpur-1'. The mutant yields 45 % more protein per unit area. The essential amino acid index is unchanged. It is likely that the high nutritional value in 'Hyprosola' seed protein arises from an increase in the albumin:globulin ratio. Nitrogen fixation rates of the mutant during the first 7 weeks of growth were found to be similar to 'Faridpur-1'. Under field conditions, the mutant may be able to nodulate more rapidly and more extensively than the parent variety. (author)

  13. Fusion genetic analysis of jasmonate-signalling mutants in Arabidopsis

    DEFF Research Database (Denmark)

    Jensen, Anders Bøgh; Raventos, D.; Mundy, John Williams

    2002-01-01

    as two recessive mutants, designated joe1 and 2, that overexpress the reporter. Genetic analysis indicated that reporter overexpression in the joe mutants requires COI. joe1 responded to MeJA with increased anthocyanin accumulation, while joe2 responded with decreased root growth inhibition. In addition...... activity was also induced by the protein kinase inhibitor staurosporine and antagonized by the protein phosphatase inhibitor okadaic acid. FLUC bio-imaging, RNA gel-blot analysis and progeny analyses identified three recessive mutants that underexpress the FLUC reporter, designated jue1, 2 and 3, as well...

  14. Mutant uromodulin expression leads to altered homeostasis of the endoplasmic reticulum and activates the unfolded protein response.

    Directory of Open Access Journals (Sweden)

    Céline Schaeffer

    Full Text Available Uromodulin is the most abundant urinary protein in physiological conditions. It is exclusively produced by renal epithelial cells lining the thick ascending limb of Henle's loop (TAL and it plays key roles in kidney function and disease. Mutations in UMOD, the gene encoding uromodulin, cause autosomal dominant tubulointerstitial kidney disease uromodulin-related (ADTKD-UMOD, characterised by hyperuricemia, gout and progressive loss of renal function. While the primary effect of UMOD mutations, retention in the endoplasmic reticulum (ER, is well established, its downstream effects are still largely unknown. To gain insight into ADTKD-UMOD pathogenesis, we performed transcriptional profiling and biochemical characterisation of cellular models (immortalised mouse TAL cells of robust expression of wild type or mutant GFP-tagged uromodulin. In this model mutant uromodulin accumulation in the ER does not impact on cell viability and proliferation. Transcriptional profiling identified 109 genes that are differentially expressed in mutant cells relative to wild type ones. Up-regulated genes include several ER resident chaperones and protein disulphide isomerases. Consistently, pathway enrichment analysis indicates that mutant uromodulin expression affects ER function and protein homeostasis. Interestingly, mutant uromodulin expression induces the Unfolded Protein Response (UPR, and specifically the IRE1 branch, as shown by an increased splicing of XBP1. Consistent with UPR induction, we show increased interaction of mutant uromodulin with ER chaperones Bip, calnexin and PDI. Using metabolic labelling, we also demonstrate that while autophagy plays no role, mutant protein is partially degraded by the proteasome through ER-associated degradation. Our work demonstrates that ER stress could play a central role in ADTKD-UMOD pathogenesis. This sets the bases for future work to develop novel therapeutic strategies through modulation of ER homeostasis and

  15. Cerebrovascular defects in Foxc1 mutants correlate with aberrant WNT and VEGF-A pathways downstream of retinoic acid from the meninges.

    Science.gov (United States)

    Mishra, Swati; Choe, Youngshik; Pleasure, Samuel J; Siegenthaler, Julie A

    2016-12-01

    Growth and maturation of the cerebrovasculature is a vital event in neocortical development however mechanisms that control cerebrovascular development remain poorly understood. Mutations in or deletions that include the FOXC1 gene are associated with congenital cerebrovascular anomalies and increased stroke risk in patients. Foxc1 mutant mice display severe cerebrovascular hemorrhage at late gestational ages. While these data demonstrate Foxc1 is required for cerebrovascular development, its broad expression in the brain vasculature combined with Foxc1 mutant's complex developmental defects have made it difficult to pinpoint its function(s). Using global and conditional Foxc1 mutants, we find 1) significant cerebrovascular growth defects precede cerebral hemorrhage and 2) expression of Foxc1 in neural crest-derived meninges and brain pericytes, though not endothelial cells, is required for normal cerebrovascular development. We provide evidence that reduced levels of meninges-derived retinoic acid (RA), caused by defects in meninges formation in Foxc1 mutants, is a major contributing factor to the cerebrovascular growth defects in Foxc1 mutants. We provide data that suggests that meninges-derived RA ensures adequate growth of the neocortical vasculature via regulating expression of WNT pathway proteins and neural progenitor derived-VEGF-A. Our findings offer the first evidence for a role of the meninges in brain vascular development and provide new insight into potential causes of cerebrovascular defects in patients with FOXC1 mutations. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Cellular responses during morphological transformation in Azospirillum brasilense and Its flcA knockout mutant.

    Directory of Open Access Journals (Sweden)

    Xingsheng Hou

    Full Text Available FlcA is a response regulator controlling flocculation and the morphological transformation of Azospirillum cells from vegetative to cyst-like forms. To understand the cellular responses of Azospirillum to conditions that cause morphological transformation, proteins differentially expressed under flocculation conditions in A. brasilense Sp7 and its flcA knockout mutant were investigated. Comparison of 2-DE protein profiles of wild-type (Sp7 and a flcA deletion mutant (Sp7-flcAΔ revealed a total of 33 differentially expressed 2-DE gel spots, with 22 of these spots confidently separated to allow protein identification. Analysis of these spots by liquid chromatography-tandem mass spectrometry (LC-MS/MS and MASCOT database searching identified 48 proteins (≥10% emPAI in each spot. The functional characteristics of these proteins included carbon metabolism (beta-ketothiolase and citrate synthase, nitrogen metabolism (Glutamine synthetase and nitric oxide synthase, stress tolerance (superoxide dismutase, Alkyl hydroperoxidase and ATP-dependent Clp protease proteolytic subunit and morphological transformation (transducer coupling protein. The observed differences between Sp7 wild-type and flcA- strains enhance our understanding of the morphological transformation process and help to explain previous phenotypical observations. This work is a step forward in connecting the Azospirillum phenome and genome.

  17. Cellular responses during morphological transformation in Azospirillum brasilense and Its flcA knockout mutant.

    Science.gov (United States)

    Hou, Xingsheng; McMillan, Mary; Coumans, Joëlle V F; Poljak, Anne; Raftery, Mark J; Pereg, Lily

    2014-01-01

    FlcA is a response regulator controlling flocculation and the morphological transformation of Azospirillum cells from vegetative to cyst-like forms. To understand the cellular responses of Azospirillum to conditions that cause morphological transformation, proteins differentially expressed under flocculation conditions in A. brasilense Sp7 and its flcA knockout mutant were investigated. Comparison of 2-DE protein profiles of wild-type (Sp7) and a flcA deletion mutant (Sp7-flcAΔ) revealed a total of 33 differentially expressed 2-DE gel spots, with 22 of these spots confidently separated to allow protein identification. Analysis of these spots by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and MASCOT database searching identified 48 proteins (≥10% emPAI in each spot). The functional characteristics of these proteins included carbon metabolism (beta-ketothiolase and citrate synthase), nitrogen metabolism (Glutamine synthetase and nitric oxide synthase), stress tolerance (superoxide dismutase, Alkyl hydroperoxidase and ATP-dependent Clp protease proteolytic subunit) and morphological transformation (transducer coupling protein). The observed differences between Sp7 wild-type and flcA- strains enhance our understanding of the morphological transformation process and help to explain previous phenotypical observations. This work is a step forward in connecting the Azospirillum phenome and genome.

  18. Cell lines derived from a Medaka radiation-sensitive mutant have defects in DNA double-strand break responses

    International Nuclear Information System (INIS)

    Hidaka, Masayuki; Oda, Shoji; Mitani, Hiroshi; Kuwahara, Yoshikazu; Fukumoto, Manabu

    2010-01-01

    It was reported that the radiation-sensitive Medaka mutant 'ric1' has a defect in the repair of DNA double-strand breaks (DSBs) induced by γ-rays during early embryogenesis. To study the cellular response of a ric1 mutant to ionizing radiation (IR), we established the mutant embryonic cell lines RIC1-e9, RIC1-e42, RIC1-e43. Following exposure to γ-irradiation, the DSBs in wild-type cells were repaired within 1 h, while those in RIC1 cells were not rejoined even after 2 h. Cell death was induced in the wild-type cells with cell fragmentation, but only a small proportion of the RIC1 cells underwent cell death, and without cell fragmentation. Although both wild-type and RIC1 cells showed mitotic inhibition immediately after γ-irradiation, cell division was much slower to resume in the wild-type cells (20 h versus 12 h). In both wild-type and RIC1 cells, Ser139 phosphorylated H2AX (γH2AX) foci were formed after γ-irradiation, however, the γH2AX foci disappeared more quickly in the RIC1 cell lines. These results suggest that the instability of γH2AX foci in RIC1 cells cause an aberration of the DNA damage response. As RIC1 cultured cells showed similar defective DNA repair as ric1 embryos and RIC1 cells revealed defective cell death and cell cycle checkpoint, they are useful for investigating DNA damage responses in vitro. (author)

  19. Arabidopsis mutants lacking asparaginases develop normally but exhibit enhanced root inhibition by exogenous asparagine.

    Science.gov (United States)

    Ivanov, Ana; Kameka, Alexander; Pajak, Agnieszka; Bruneau, Luanne; Beyaert, Ronald; Hernández-Sebastià, Cinta; Marsolais, Frédéric

    2012-06-01

    Asparaginase catalyzes the degradation of L-asparagine to L-aspartic acid and ammonia, and is implicated in the catabolism of transported asparagine in sink tissues of higher plants. The Arabidopsis genome includes two genes, ASPGA1 and ASPGB1, belonging to distinct asparaginase subfamilies. Conditions of severe nitrogen limitation resulted in a slight decrease in seed size in wild-type Arabidopsis. However, this response was not observed in a homozygous T-DNA insertion mutant where ASPG genes had been inactivated. Under nitrogen-sufficient conditions, the ASPG mutant had elevated levels of free asparagine in mature seed. This phenotype was observed exclusively under conditions of low illumination, when a low ratio of carbon to nitrogen was translocated to the seed. Mutants deficient in one or both asparaginases were more sensitive than wild-type to inhibition of primary root elongation and root hair emergence by L-asparagine as a single nitrogen source. This enhanced inhibition was associated with increased accumulation of asparagine in the root of the double aspga1-1/-b1-1 mutant. This indicates that inhibition of root growth is likely elicited by asparagine itself or an asparagine-derived metabolite, other than the products of asparaginase, aspartic acid or ammonia. During germination, a fusion between the ASPGA1 promoter and beta-glucuronidase was expressed in endosperm cells starting at the micropylar end. Expression was initially high throughout the root and hypocotyl, but became restricted to the root tip after three days, which may indicate a transition to nitrogen-heterotrophic growth.

  20. Mycothiol-Deficient Mycobacterium smegmatis Mutants Are Hypersensitive to Alkylating Agents, Free Radicals, and Antibiotics

    Science.gov (United States)

    Rawat, Mamta; Newton, Gerald L.; Ko, Mary; Martinez, Gladys J.; Fahey, Robert C.; Av-Gay, Yossef

    2002-01-01

    Mycothiol (MSH; 1d-myo-inosityl 2-[N-acetyl-l-cysteinyl]amido-2-deoxy-α-d-glucopyranoside) is the major low-molecular-weight thiol produced by mycobacteria. Mutants of Mycobacterium smegmatis mc2155 deficient in MSH production were produced by chemical mutagenesis as well as by transposon mutagenesis. One chemical mutant (mutant I64) and two transposon mutants (mutants Tn1 and Tn2) stably deficient in MSH production were isolated by screening for reduced levels of MSH content. The MSH contents of transposon mutants Tn1 and Tn2 were found to be less than 0.1% that of the parent strain, and the MSH content of I64 was found to be 1 to 5% that of the parent strain. All three strains accumulated 1d-myo-inosityl 2-deoxy-α-d-glucopyranoside to levels 20- to 25-fold the level found in the parent strain. The cysteine:1d-myo-inosityl 2-amino-2-deoxy-α-d-glucopyranoside ligase (MshC) activities of the three mutant strains were ≤2% that of the parent strain. Phenotypic analysis revealed that these MSH-deficient mutants possess increased susceptibilities to free radicals and alkylating agents and to a wide range of antibiotics including erythromycin, azithromycin, vancomycin, penicillin G, rifamycin, and rifampin. Conversely, the mutants possess at least 200-fold higher levels of resistance to isoniazid than the wild type. We mapped the mutation in the chemical mutant by sequencing the mshC gene and showed that a single amino acid substitution (L205P) is responsible for reduced MSH production and its associated phenotype. Our results demonstrate that there is a direct correlation between MSH depletion and enhanced sensitivity to toxins and antibiotics. PMID:12384335

  1. Pleiotropic effects of hemagglutinin amino acid substitutions of H5 influenza escape mutants

    International Nuclear Information System (INIS)

    Rudneva, Irina A.; Timofeeva, Tatiana A.; Ignatieva, Anna V.; Shilov, Aleksandr A.; Krylov, Petr S.; Ilyushina, Natalia A.; Kaverin, Nikolai V.

    2013-01-01

    In the present study we assessed pleiotropic characteristics of the antibody-selected mutations. We examined pH optimum of fusion, temperatures of HA heat inactivation, and in vitro and in vivo replication kinetics of the previously obtained influenza H5 escape mutants. Our results showed that HA1 N142K mutation significantly lowered the pH of fusion optimum. Mutations of the escape mutants located in the HA lateral loop significantly affected H5 HA thermostability (P<0.05). HA changes at positions 131, 144, 145, and 156 and substitutions at positions 131, 142, 145, and 156 affected the replicative ability of H5 escape mutants in vitro and in vivo, respectively. Overall, a co-variation between antigenic specificity and different HA phenotypic properties has been demonstrated. We believe that the monitoring of pleiotropic effects of the HA mutations found in H5 escape mutants is essential for accurate prediction of mutants with pandemic potential. - Highlights: • HA1 N142K mutation significantly lowered the pH of fusion optimum. • Mutations located in the HA lateral loop significantly affected H5 HA thermostability. • HA changes at positions 131, 142, 144, 145, and 156 affected the replicative ability of H5 mutants. • Acquisition of glycosylation site could lead to the emergence of multiple pleiotropic effects

  2. Pleiotropic effects of hemagglutinin amino acid substitutions of H5 influenza escape mutants

    Energy Technology Data Exchange (ETDEWEB)

    Rudneva, Irina A.; Timofeeva, Tatiana A.; Ignatieva, Anna V.; Shilov, Aleksandr A.; Krylov, Petr S. [D.I. Ivanovsky Institute of Virology, 123098 Moscow (Russian Federation); Ilyushina, Natalia A., E-mail: Natalia.Ilyushina@fda.hhs.gov [FDA CDER, 29 Lincoln Drive, Bethesda, MD 20892 (United States); Kaverin, Nikolai V., E-mail: nik.kaverin@gmail.com [D.I. Ivanovsky Institute of Virology, 123098 Moscow (Russian Federation)

    2013-12-15

    In the present study we assessed pleiotropic characteristics of the antibody-selected mutations. We examined pH optimum of fusion, temperatures of HA heat inactivation, and in vitro and in vivo replication kinetics of the previously obtained influenza H5 escape mutants. Our results showed that HA1 N142K mutation significantly lowered the pH of fusion optimum. Mutations of the escape mutants located in the HA lateral loop significantly affected H5 HA thermostability (P<0.05). HA changes at positions 131, 144, 145, and 156 and substitutions at positions 131, 142, 145, and 156 affected the replicative ability of H5 escape mutants in vitro and in vivo, respectively. Overall, a co-variation between antigenic specificity and different HA phenotypic properties has been demonstrated. We believe that the monitoring of pleiotropic effects of the HA mutations found in H5 escape mutants is essential for accurate prediction of mutants with pandemic potential. - Highlights: • HA1 N142K mutation significantly lowered the pH of fusion optimum. • Mutations located in the HA lateral loop significantly affected H5 HA thermostability. • HA changes at positions 131, 142, 144, 145, and 156 affected the replicative ability of H5 mutants. • Acquisition of glycosylation site could lead to the emergence of multiple pleiotropic effects.

  3. Hyperpolarized 13C MR imaging detects no lactate production in mutant IDH1 gliomas: Implications for diagnosis and response monitoring

    Directory of Open Access Journals (Sweden)

    Myriam M. Chaumeil

    2016-01-01

    Full Text Available Metabolic imaging of brain tumors using 13C Magnetic Resonance Spectroscopy (MRS of hyperpolarized [1-13C] pyruvate is a promising neuroimaging strategy which, after a decade of preclinical success in glioblastoma (GBM models, is now entering clinical trials in multiple centers. Typically, the presence of GBM has been associated with elevated hyperpolarized [1-13C] lactate produced from [1-13C] pyruvate, and response to therapy has been associated with a drop in hyperpolarized [1-13C] lactate. However, to date, lower grade gliomas had not been investigated using this approach. The most prevalent mutation in lower grade gliomas is the isocitrate dehydrogenase 1 (IDH1 mutation, which, in addition to initiating tumor development, also induces metabolic reprogramming. In particular, mutant IDH1 gliomas are associated with low levels of lactate dehydrogenase A (LDHA and monocarboxylate transporters 1 and 4 (MCT1, MCT4, three proteins involved in pyruvate metabolism to lactate. We therefore investigated the potential of 13C MRS of hyperpolarized [1-13C] pyruvate for detection of mutant IDH1 gliomas and for monitoring of their therapeutic response. We studied patient-derived mutant IDH1 glioma cells that underexpress LDHA, MCT1 and MCT4, and wild-type IDH1 GBM cells that express high levels of these proteins. Mutant IDH1 cells and tumors produced significantly less hyperpolarized [1-13C] lactate compared to GBM, consistent with their metabolic reprogramming. Furthermore, hyperpolarized [1-13C] lactate production was not affected by chemotherapeutic treatment with temozolomide (TMZ in mutant IDH1 tumors, in contrast to previous reports in GBM. Our results demonstrate the unusual metabolic imaging profile of mutant IDH1 gliomas, which, when combined with other clinically available imaging methods, could be used to detect the presence of the IDH1 mutation in vivo.

  4. Gibberellic acid and cGMP-dependent transcriptional regulation in arabidopsis thaliana

    KAUST Repository

    Bastian, René

    2010-03-01

    An ever increasing amount of transcriptomic data and analysis tools provide novel insight into complex responses of biological systems. Given these resources we have undertaken to review aspects of transcriptional regulation in response to the plant hormone gibberellic acid (GA) and its second messenger guanosine 3\\',5\\'-cyclic monophosphate (cGMP) in Arabidopsis thaliana, both wild type and selected mutants. Evidence suggests enrichment of GA-responsive (GARE) elements in promoters of genes that are transcriptionally upregulated in response to cGMP but downregulated in a GA insensitive mutant (ga1-3). In contrast, in the genes upregulated in the mutant, no enrichment in the GARE is observed suggesting that GARE motifs are diagnostic for GA-induced and cGMP-dependent transcriptional upregulation. Further, we review how expression studies of GA-dependent transcription factors and transcriptional networks based on common promoter signatures derived from ab initio analyses can contribute to our understanding of plant responses at the systems level. © 2010 Landes Bioscience.

  5. Enhanced transpiration rate in the high pigment 1 tomato mutant and its physiological significance.

    Science.gov (United States)

    Carvalho, R F; Aidar, S T; Azevedo, R A; Dodd, I C; Peres, L E P

    2011-05-01

    Tomato high pigment (hp) mutants represent an interesting horticultural resource due to their enhanced accumulation of carotenoids, flavonoids and vitamin C. Since hp mutants are known for their exaggerated light responses, the molecules accumulated are likely to be antioxidants, recruited to deal with light and others stresses. Further phenotypes displayed by hp mutations are reduced growth and an apparent disturbance in water loss. Here, we examined the impact of the hp1 mutation and its near isogenic line cv Micro-Tom (MT) on stomatal conductance (gs), transpiration (E), CO(2) assimilation (A) and water use efficiency (WUE). Detached hp1 leaves lost water more rapidly than control leaves, but this behaviour was reversed by exogenous abscisic acid (ABA), indicating the ability of hp1 to respond to this hormone. Although attached hp1 leaves had enhanced gs, E and A compared to control leaves, genotypic differences were lost when water was withheld. Both instantaneous leaf-level WUE and long-term whole plant WUE did not differ between hp1 and MT. Our results indicate a link between exaggerated light response and water loss in hp1, which has important implications for the use of this mutant in both basic and horticultural research. © 2011 German Botanical Society and The Royal Botanical Society of the Netherlands.

  6. [Substrate specificities of bile salt hydrolase 1 and its mutants from Lactobacillus salivarius].

    Science.gov (United States)

    Bi, Jie; Fang, Fang; Qiu, Yuying; Yang, Qingli; Chen, Jian

    2014-03-01

    In order to analyze the correlation between critical residues in the catalytic centre of BSH and the enzyme substrate specificity, seven mutants of Lactobacillus salivarius bile salt hydrolase (BSH1) were constructed by using the Escherichia coli pET-20b(+) gene expression system, rational design and site-directed mutagenesis. These BSH1 mutants exhibited different hydrolytic activities against various conjugated bile salts through substrate specificities comparison. Among the residues being tested, Cys2 and Thr264 were deduced as key sites for BSH1 to catalyze taurocholic acid and glycocholic acid, respectively. Moreover, Cys2 and Thr264 were important for keeping the catalytic activity of BSH1. The high conservative Cys2 was not the only active site, other mutant amino acid sites were possibly involved in substrate binding. These mutant residues might influence the space and shape of the substrate-binding pockets or the channel size for substrate passing through and entering active site of BSH1, thus, the hydrolytic activity of BSH1 was changed to different conjugated bile salt.

  7. Increased sensitivity to salt stress in tocopherol-deficient Arabidopsis mutants growing in a hydroponic system

    Science.gov (United States)

    Ellouzi, Hasna; Hamed, Karim Ben; Cela, Jana; Müller, Maren; Abdelly, Chedly; Munné-Bosch, Sergi

    2013-01-01

    Recent studies suggest that tocopherols could play physiological roles in salt tolerance but the mechanisms are still unknown. In this study, we analyzed changes in growth, mineral and oxidative status in vte1 and vte4 Arabidopsis thaliana mutants exposed to salt stress. vte1 and vte4 mutants lack α-tocopherol, but only the vte1 mutant is additionally deficient in γ-tocopherol. Results showed that a deficiency in vitamin E leads to reduced growth and increased oxidative stress in hydroponically-grown plants. This effect was observed at early stages, not only in rosettes but also in roots. The vte1 mutant was more sensitive to salt-induced oxidative stress than the wild type and the vte4 mutant. Salt sensitivity was associated with (i) high contents of Na+, (ii) reduced efficiency of PSII photochemistry (Fv/Fm ratio) and (iii) more pronounced oxidative stress as indicated by increased hydrogen peroxide and malondialdeyde levels. The vte 4 mutant, which accumulates γ- instead of α-tocopherol showed an intermediate sensitivity to salt stress between the wild type and the vte1 mutant. Contents of abscisic acid, jasmonic acid and the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid were higher in the vte1 mutant than the vte4 mutant and wild type. It is concluded that vitamin E-deficient plants show an increased sensitivity to salt stress both in rosettes and roots, therefore indicating the positive role of tocopherols in stress tolerance, not only by minimizing oxidative stress, but also controlling Na+/K+ homeostasis and hormonal balance. PMID:23299430

  8. Precocious metamorphosis in the juvenile hormone-deficient mutant of the silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Takaaki Daimon

    Full Text Available Insect molting and metamorphosis are intricately governed by two hormones, ecdysteroids and juvenile hormones (JHs. JHs prevent precocious metamorphosis and allow the larva to undergo multiple rounds of molting until it attains the proper size for metamorphosis. In the silkworm, Bombyx mori, several "moltinism" mutations have been identified that exhibit variations in the number of larval molts; however, none of them have been characterized molecularly. Here we report the identification and characterization of the gene responsible for the dimolting (mod mutant that undergoes precocious metamorphosis with fewer larval-larval molts. We show that the mod mutation results in complete loss of JHs in the larval hemolymph and that the mutant phenotype can be rescued by topical application of a JH analog. We performed positional cloning of mod and found a null mutation in the cytochrome P450 gene CYP15C1 in the mod allele. We also demonstrated that CYP15C1 is specifically expressed in the corpus allatum, an endocrine organ that synthesizes and secretes JHs. Furthermore, a biochemical experiment showed that CYP15C1 epoxidizes farnesoic acid to JH acid in a highly stereospecific manner. Precocious metamorphosis of mod larvae was rescued when the wild-type allele of CYP15C1 was expressed in transgenic mod larvae using the GAL4/UAS system. Our data therefore reveal that CYP15C1 is the gene responsible for the mod mutation and is essential for JH biosynthesis. Remarkably, precocious larval-pupal transition in mod larvae does not occur in the first or second instar, suggesting that authentic epoxidized JHs are not essential in very young larvae of B. mori. Our identification of a JH-deficient mutant in this model insect will lead to a greater understanding of the molecular basis of the hormonal control of development and metamorphosis.

  9. Identification, cloning and characterization of sis7 and sis10 sugar-insensitive mutants of Arabidopsis

    Directory of Open Access Journals (Sweden)

    Biddle Kelly D

    2008-10-01

    Full Text Available Abstract Background The levels of soluble sugars, such as glucose and sucrose, help regulate many plant metabolic, physiological and developmental processes. Genetic screens are helping identify some of the loci involved in plant sugar response and reveal extensive cross-talk between sugar and phytohormone response pathways. Results A forward genetic screen was performed to identify mutants with increased resistance to the inhibitory effects of high levels of exogenous sugars on early Arabidopsis seedling development. The positional cloning and characterization of two of these sugar insensitive (sis mutants, both of which are also involved in abscisic acid (ABA biosynthesis or response, are reported. Plants carrying mutations in SIS7/NCED3/STO1 or SIS10/ABI3 are resistant to the inhibitory effects of high levels of exogenous Glc and Suc. Quantitative RT-PCR analyses indicate transcriptional upregulation of ABA biosynthesis genes by high concentrations of Glc in wild-type germinating seeds. Gene expression profiling revealed that a significant number of genes that are expressed at lower levels in germinating sis7-1/nced3-4/sto1-4 seeds than in wild-type seeds are implicated in auxin biosynthesis or transport, suggesting cross-talk between ABA and auxin response pathways. The degree of sugar insensitivity of different sis10/abi3 mutant seedlings shows a strong positive correlation with their level of ABA insensitivity during seed germination. Conclusion Mutations in the SIS7/NCED3/STO1 gene, which is primarily required for ABA biosynthesis under drought conditions, confer a sugar-insensitive phenotype, indicating that a constitutive role in ABA biosynthesis is not necessary to confer sugar insensitivity. Findings presented here clearly demonstrate that mutations in ABI3 can confer a sugar-insensitive phenotype and help explain previous, mixed reports on this topic by showing that ABA and sugar insensitivity exhibit a strong positive correlation in

  10. PHAGE RESISTANT LACTIC ACID BACTERIAL MUTANTS

    DEFF Research Database (Denmark)

    2001-01-01

    Method of obtaining mutated lactic acid bacteria having a reduced susceptibility towards attack by bacteriophages, the method comprising mutating a gene involved in the pyrimidine metabolism, including pyrG encoding CTP synthetase. Such lactic acid bacteria are useful in starter cultures...

  11. Proteomic analysis of the response of Escherichia coli to short-chain fatty acids.

    Science.gov (United States)

    Rodríguez-Moyá, María; Gonzalez, Ramon

    2015-06-03

    Given their simple and easy-to-manipulate chemical structures, short-chain fatty acids (SCFAs) are valuable feedstocks for many industrial applications. While the microbial production of SCFAs by engineered Escherichia coli has been demonstrated recently, productivity and yields are limited by their antimicrobial properties. In this work, we performed a comparative proteomic analysis of E. coli under octanoic acid stress (15 mM) and identified the underlying mechanisms of SCFA toxicity. Out of a total of 33 spots differentially expressed at a p-value ≤ 0.05, nine differentially expressed proteins involved in transport and structural roles (OmpF, HPr, and FliC), oxidative stress (SodA, SodB, and TrxA), protein synthesis (PPiB and RpsA) and metabolic functions (HPr, PflB) were selected for further investigation. Our studies suggest that membrane damage and oxidative stress are the main routes of inhibition by SCFAs in E. coli. The outer membrane porin OmpF had the greatest impact on SCFA tolerance. Intracellular pH analysis on ompF mutants grown under octanoic acid stress indicated that this porin facilitates transport of SCFAs into the cell. The same response was observed under hexanoic acid stress, further supporting the role of OmpF in response to the presence of SCFAs. Furthermore, analysis of membrane protein expression revealed that other outer membrane porins are also involved in the response of E. coli to SCFAs. This work covers the first known proteomic analysis to assess the inhibitory effect of SCFAs in E. coli. SCFAs are molecules of great interest in the industry, but their microbial production is limited by their antimicrobial properties. This work allowed identification of differentially expressed proteins in response to SCFA stress and demonstrated the relevance of short- and medium-chain FA transport across the cell membrane via outer membrane porins, providing valuable insights on the toxicity mechanism of SCFAs in E. coli. These results could

  12. vph6 mutants of Saccharomyces cerevisiae require calcineurin for growth and are defective in vacuolar H(+)-ATPase assembly.

    Science.gov (United States)

    Hemenway, C S; Dolinski, K; Cardenas, M E; Hiller, M A; Jones, E W; Heitman, J

    1995-11-01

    We have characterized a Saccharomyces cerevisiae mutant strain that is hypersensitive to cyclosporin A (CsA) and FK506, immunosuppressants that inhibit calcineurin, a serine-threonine-specific phosphatase (PP2B). A single nuclear mutation, designated cev1 for calcineurin essential for viability, is responsible for the CsA-FK506-sensitive phenotype. The peptidyl-prolyl cis-trans isomerases cyclophilin A and FKBP12, respectively, mediate CsA and FK506 toxicity in the cev1 mutant strain. We demonstrate that cev1 is an allele of the VPH6 gene and that vph6 mutant strains fail to assemble the vacuolar H(+)-ATPase (V-ATPase). The VPH6 gene was mapped on chromosome VIII and is predicted to encode a 181-amino acid (21 kD) protein with no identity to other known proteins. We find that calcineurin is essential for viability in many mutant strains with defects in V-ATPase function or vacuolar acidification. In addition, we find that calcineurin modulates extracellular acidification in response to glucose, which we propose occurs via calcineurin regulation of the plasma membrane H(+)-ATPase PMA1. Taken together, our findings suggest calcineurin plays a general role in the regulation of cation transport and homeostasis.

  13. Mms Sensitivity of All Amino Acid-Requiring Mutants in Aspergillus and Its Suppression by Mutations in a Single Gene

    OpenAIRE

    Käfer, Etta

    1987-01-01

    All available amino acid-requiring mutants of Aspergillus nidulans were found to be hypersensitive to MMS (methyl methanesulfonate) to various degrees. On MMS media, secondary mutations could be selected which suppress this MMS sensitivity but do not affect the requirement. Many such mutations were analyzed and found to be alleles of one gene, smsA (= suppressor of MMS sensitivity), which mapped distal on the right arm of chromosome V. This gene is more likely to be involved in general regula...

  14. Nanoformulated cell-penetrating survivin mutant and its dual actions

    Directory of Open Access Journals (Sweden)

    Sriramoju B

    2014-07-01

    Full Text Available Bhasker Sriramoju, Rupinder K Kanwar, Jagat R Kanwar Nanomedicine Laboratory of Immunology and Molecular Biomedical Research (NLIMBR, School of Medicine, Faculty of Health, Deakin University, Geelong, Australia Abstract: In this study, we investigated the differential actions of a dominant-negative survivin mutant (SurR9-C84A against cancerous SK-N-SH neuroblastoma cell lines and differentiated SK-N-SH neurons. In both the cases, the mutant protein displayed dual actions, where its effects were cytotoxic toward cancerous cells and proliferative toward the differentiated neurons. This can be explained by the fact that tumorous (undifferentiated SK-N-SH cells have a high endogenous survivin pool and upon treatment with mutant SuR9-C84A causes forceful survivin expression. These events significantly lowered the microtubule dynamics and stability, eventually leading to apoptosis. In the case of differentiated SK-N-SH neurons that express negligible levels of wild-type survivin, the mutant indistinguishably behaved in a wild-type fashion. It also favored cell-cycle progression, forming the chromosome-passenger complex, and stabilized the microtubule-organizing center. Therefore, mutant SurR9-C84A represents a novel therapeutic with its dual actions (cytotoxic toward tumor cells and protective and proliferative toward neuronal cells, and hence finds potential applications against a variety of neurological disorders. In this study, we also developed a novel poly(lactic-co-glycolic acid nanoparticulate formulation to surmount the hurdles associated with the delivery of SurR9-C84A, thus enhancing its effective therapeutic outcome. Keywords: survivin mutant, neurological disorders, protein therapeutics, inhibitor of apoptosis protein family, poly(lactic-co-glycolic acid

  15. Genetic Analysis of Diaminopimelic Acid- and Lysine-Requiring Mutants of Escherichia coli1

    Science.gov (United States)

    Bukhari, Ahmad I.; Taylor, Austin L.

    1971-01-01

    Several diaminopimelic acid (DAP)- and lysine-requiring mutants of Escherichia coli were isolated and studied by genetic, physiological, and biochemical means. The genes concerned with DAP-lysine synthesis map at several different sites on the E. coli chromosome and, therefore, do not constitute a single operon. Three separate loci affecting DAP synthesis are located in the 0 to 2.5 min region of the genetic map. The order of the loci in this region is thr-dapB-pyrA-ara-leu-pan-dapC-tonA-dapD. Two additional DAP genes map in the region between min 47 and 48, with the gene order being gua-dapA-dapE-ctr. The lys locus at min 55 determines the synthesis of the enzyme DAP decarboxylase, which catalyzes the conversion of DAP into lysine. The order of the genes in this region is serA-lysA-thyA. PMID:4926684

  16. Human COQ9 Rescues a coq9 Yeast Mutant by Enhancing Coenzyme Q Biosynthesis from 4-Hydroxybenzoic Acid and Stabilizing the CoQ-Synthome

    Directory of Open Access Journals (Sweden)

    Cuiwen H. He

    2017-07-01

    Full Text Available Coq9 is required for the stability of a mitochondrial multi-subunit complex, termed the CoQ-synthome, and the deamination step of Q intermediates that derive from para-aminobenzoic acid (pABA in yeast. In human, mutations in the COQ9 gene cause neonatal-onset primary Q10 deficiency. In this study, we determined whether expression of human COQ9 could complement yeast coq9 point or null mutants. We found that expression of human COQ9 rescues the growth of the temperature-sensitive yeast mutant, coq9-ts19, on a non-fermentable carbon source and increases the content of Q6, by enhancing Q biosynthesis from 4-hydroxybenzoic acid (4HB. To study the mechanism for the rescue by human COQ9, we determined the steady-state levels of yeast Coq polypeptides in the mitochondria of the temperature-sensitive yeast coq9 mutant expressing human COQ9. We show that the expression of human COQ9 significantly increased steady-state levels of yeast Coq4, Coq6, Coq7, and Coq9 at permissive temperature. Human COQ9 polypeptide levels persisted at non-permissive temperature. A small amount of the human COQ9 co-purified with tagged Coq6, Coq6-CNAP, indicating that human COQ9 interacts with the yeast Q-biosynthetic complex. These findings suggest that human COQ9 rescues the yeast coq9 temperature-sensitive mutant by stabilizing the CoQ-synthome and increasing Q biosynthesis from 4HB. This finding provides a powerful approach to studying the function of human COQ9 using yeast as a model.

  17. Assessing the Role of ETHYLENE RESPONSE FACTOR Transcriptional Repressors in Salicylic Acid-Mediated Suppression of Jasmonic Acid-Responsive Genes

    NARCIS (Netherlands)

    Caarls, Lotte; van der Does, Adriana; Hickman, Richard; Jansen, Wouter; van Verk, Marcel; Proietti, Silvia; Lorenzo, Oscar; Solano, Roberto; Pieterse, Corné M J; Van Wees, Saskia C M

    2017-01-01

    Salicylic acid (SA) and jasmonic acid (JA) cross-communicate in the plant immune signaling network to finely regulate induced defenses. In Arabidopsis, SA antagonizes many JA-responsive genes, partly by targeting the ETHYLENE RESPONSE FACTOR (ERF)-type transcriptional activator ORA59. Members of the

  18. Viral Escape Mutant Epitope Maintains TCR Affinity for Antigen yet Curtails CD8 T Cell Responses.

    Directory of Open Access Journals (Sweden)

    Shayla K Shorter

    Full Text Available T cells have the remarkable ability to recognize antigen with great specificity and in turn mount an appropriate and robust immune response. Critical to this process is the initial T cell antigen recognition and subsequent signal transduction events. This antigen recognition can be modulated at the site of TCR interaction with peptide:major histocompatibility (pMHC or peptide interaction with the MHC molecule. Both events could have a range of effects on T cell fate. Though responses to antigens that bind sub-optimally to TCR, known as altered peptide ligands (APL, have been studied extensively, the impact of disrupting antigen binding to MHC has been highlighted to a lesser extent and is usually considered to result in complete loss of epitope recognition. Here we present a model of viral evasion from CD8 T cell immuno-surveillance by a lymphocytic choriomeningitis virus (LCMV escape mutant with an epitope for which TCR affinity for pMHC remains high but where the antigenic peptide binds sub optimally to MHC. Despite high TCR affinity for variant epitope, levels of interferon regulatory factor-4 (IRF4 are not sustained in response to the variant indicating differences in perceived TCR signal strength. The CD8+ T cell response to the variant epitope is characterized by early proliferation and up-regulation of activation markers. Interestingly, this response is not maintained and is characterized by a lack in IL-2 and IFNγ production, increased apoptosis and an abrogated glycolytic response. We show that disrupting the stability of peptide in MHC can effectively disrupt TCR signal strength despite unchanged affinity for TCR and can significantly impact the CD8+ T cell response to a viral escape mutant.

  19. Isolation of New Gravitropic Mutants under Hypergravity Conditions.

    Science.gov (United States)

    Mori, Akiko; Toyota, Masatsugu; Shimada, Masayoshi; Mekata, Mika; Kurata, Tetsuya; Tasaka, Masao; Morita, Miyo T

    2016-01-01

    Forward genetics is a powerful approach used to link genotypes and phenotypes, and mutant screening/analysis has provided deep insights into many aspects of plant physiology. Gravitropism is a tropistic response in plants, in which hypocotyls and stems sense the direction of gravity and grow upward. Previous studies of gravitropic mutants have suggested that shoot endodermal cells in Arabidopsis stems and hypocotyls are capable of sensing gravity (i.e., statocytes). In the present study, we report a new screening system using hypergravity conditions to isolate enhancers of gravitropism mutants, and we also describe a rapid and efficient genome mapping method, using next-generation sequencing (NGS) and single nucleotide polymorphism (SNP)-based markers. Using the endodermal-amyloplast less 1 ( eal1 ) mutant, which exhibits defective development of endodermal cells and gravitropism, we found that hypergravity (10 g) restored the reduced gravity responsiveness in eal1 hypocotyls and could, therefore, be used to obtain mutants with further reduction in gravitropism in the eal1 background. Using the new screening system, we successfully isolated six ene ( enhancer of eal1 ) mutants that exhibited little or no gravitropism under hypergravity conditions, and using NGS and map-based cloning with SNP markers, we narrowed down the potential causative genes, which revealed a new genetic network for shoot gravitropism in Arabidopsis .

  20. Survival and mutant production induced by mutagenic agents in Metarhizium anisopliae Sobrevivência e obtenção de mutantes induzidos por agentes mutagênicos em Metarhizium anisopliae

    Directory of Open Access Journals (Sweden)

    V. Kava - Cordeiro

    1995-12-01

    Full Text Available A wild strain of Metarhizium anisopliae, an entomopathogenic fungus, was submitted to three mutagenic agents: gamma radiation, ultraviolet light and nitrous acid. Survival curves were obtained and mutants were selected using different mutagenic doses which gave 1 to 5% survival. Morphological and auxotrophic mutants were isolated. Morphological mutants were grouped in a class with yellow conidia and other with pale vinaceous conidia as opposed to the green wild type conidia. Auxotrophic mutants had requirements for vitamin and aminoacid biosynthesis. More than 58% of the total auxotrophk mutants required proline/aipnine. Gamma radiation showed to be the most efficient mutagenic agent giving 0.2% of auxotrophk mutants followed by ultraviolet light (0.12% and nitrous acid (0.06%.The conidial colour and auxotrophk mutants isolated until now from M. anisopliae were reviewed.Uma linhagem selvagem do fungo entomopatogênico Metarhizium anisopliae foi submetida à ação de três agentes mutagênicos: radiação gama, luz ultravioleta e ácido nitroso. Curvas de sobrevivência foram obtidas para cada mutagênicos utilizado e mutantes foram selecionados a partir de doses dos mutagênicos que proporcionassem de 1 a 5% de sobrevivência. Mutantes morfológicos para a coloração de conídios e mutantes auxotróficos foram isolados. Mutantes para coloração de conidios foram agrupados em duas classes, uma com conídios amarelos e outra com conídios vinho pálido. Os mutantes auxotróficos obtidos foram deficientes para aminoácidos e vitaminas e mais de 58% deles eram auxotróficos para prolina/argmina. Radiação gama foi o mutagênico mais eficiente com uma porcentagem de obtenção de mulantes auxotróficos de aproximadamente 0,2%, seguido pela luz ultravioleta (0.12% e pelo ácido nitroso (0.06%.Os mulantes morfológicos e auxotróficos obtidos até o momento em Metarhizium anisopliae foram revistos.

  1. Arabidopsis redox status in response to caterpillar herbivory

    Directory of Open Access Journals (Sweden)

    Jamuna ePaudel

    2013-05-01

    Full Text Available Plant responses to insect herbivory are regulated through complex, hormone-mediated interactions. Some caterpillar species have evolved strategies to manipulate this system by inducing specific pathways that suppress plant defense responses. Effectors in the labial saliva (LS secretions of Spodoptera exigua caterpillars are believed to induce the salicylic acid (SA pathway to interfere with the jasmonic acid (JA defense pathway; however, the mechanism underlying this subversion is unknown. Since Noctuid caterpillar LS contains enzymes that may affect cellular redox balance, this study investigated rapid changes in cellular redox metabolites within 45 min after herbivory. Caterpillar LS is involved in suppressing the increase in oxidative stress that was observed in plants fed upon by caterpillars with impaired LS secretions. To further understand the link between cellular redox balance and plant defense responses, marker genes of SA, JA and ethylene (ET pathways were compared in wildtype, the glutathione-compromised pad2-1 mutant and the tga2/5/6 triple mutant plants. AtPR1 and AtPDF1.2 showed LS-dependent expression that was alleviated in the pad2-1 and tga2/5/6 triple mutants. In comparison, the ET-dependent genes ERF1 expression showed LS-associated changes in both wildtype and pad2-1 mutant plants and the ORA 59 marker AtHEL had increased expression in response to herbivory, but a LS-dependent difference was not noted. These data support the model that there are SA/NPR1-, glutathione-dependent and ET-, glutathione-independent mechanisms leading to LS-associated suppression of plant induced defences.

  2. Low Ascorbic Acid in the vtc-1 Mutant of Arabidopsis Is Associated with Decreased Growth and Intracellular Redistribution of the Antioxidant System1

    Science.gov (United States)

    Veljovic-Jovanovic, Sonja D.; Pignocchi, Cristina; Noctor, Graham; Foyer, Christine H.

    2001-01-01

    Ascorbic acid has numerous and diverse roles in plant metabolism. We have used the vtc-1 mutant of Arabidopsis, which is deficient in ascorbate biosynthesis, to investigate the role of ascorbate concentration in growth, regulation of photosynthesis, and control of the partitioning of antioxidative enyzmes. The mutant possessed 70% less ascorbate in the leaves compared with the wild type. This lesion was associated with a slight increase in total glutathione but no change in the redox state of either ascorbate or glutathione. In vtc-1, total ascorbate in the apoplast was decreased to 23% of the wild-type value. The mutant displayed much slower shoot growth than the wild type when grown in air or at high CO2 (3 mL L−1), where oxidative stress is diminished. Leaves were smaller, and shoot fresh weight and dry weight were lower in the mutant. No significant differences in the light saturation curves for CO2 assimilation were found in air or at high CO2, suggesting that the effect on growth was not due to decreased photosynthetic capacity in the mutant. Analysis of chlorophyll a fluorescence quenching revealed only a slight effect on non-photochemical energy dissipation. Hydrogen peroxide contents were similar in the leaves of the vtc-1 mutant and the wild type. Total leaf peroxidase activity was increased in the mutant and compartment-specific differences in ascorbate peroxidase (APX) activity were observed. In agreement with the measurements of enzyme activity, the expression of cytosolic APX was increased, whereas that for chloroplast APX isoforms was either unchanged or slightly decreased. These data implicate ascorbate concentration in the regulation of the compartmentalization of the antioxidant system in Arabidopsis. PMID:11598218

  3. Radiation-sensitive mutant of hypertoxinogenic strain 569B of Vibro cholerae

    International Nuclear Information System (INIS)

    Das, G.; Das, J.

    1983-01-01

    A radiation-sensitive mutant of the hypertoxinogenic strain 569B of Vibrio cholerae was isolated and characterized. The mutant, designated V. cholerae 569Bsub(s), lacks both excision- and medium-dependent dark-repair mechanisms of UV-induced DNA damage while retaining the wild-type photoreactivating capability. Analysis of the UV-irradiated cell DNA by velocity sedimentation in alkaline sucrose gradient suggests that UV-induced pyrimidine dimers may not be incised in these cells. In contrast to the wild-type cells, the mutant cell DNA was degraded after treatment with nalidixic acid. The mutant cells failed to produce any detectable amount of cholera toxin as measured by ileal-loop assay. (orig.)

  4. SOS2-LIKE PROTEIN KINASE5, an SNF1-RELATED PROTEIN KINASE3-Type Protein Kinase, Is Important for Abscisic Acid Responses in Arabidopsis through Phosphorylation of ABSCISIC ACID-INSENSITIVE51[OPEN

    Science.gov (United States)

    Zhou, Xiaona; Hao, Hongmei; Zhang, Yuguo; Bai, Yili; Zhu, Wenbo; Qin, Yunxia; Yuan, Feifei; Zhao, Feiyi; Wang, Mengyao; Hu, Jingjiang; Xu, Hong; Guo, Aiguang; Zhao, Huixian; Zhao, Yang; Cao, Cuiling; Yang, Yongqing; Schumaker, Karen S.; Guo, Yan; Xie, Chang Gen

    2015-01-01

    Abscisic acid (ABA) plays an essential role in seed germination. In this study, we demonstrate that one SNF1-RELATED PROTEIN KINASE3-type protein kinase, SOS2-LIKE PROTEIN KINASE5 (PKS5), is involved in ABA signal transduction via the phosphorylation of an interacting protein, ABSCISIC ACID-INSENSITIVE5 (ABI5). We found that pks5-3 and pks5-4, two previously identified PKS5 superactive kinase mutants with point mutations in the PKS5 FISL/NAF (a conserved peptide that is necessary for interaction with SOS3 or SOS3-LIKE CALCIUM BINDING PROTEINs) motif and the kinase domain, respectively, are hypersensitive to ABA during seed germination. PKS5 was found to interact with ABI5 in yeast (Saccharomyces cerevisiae), and this interaction was further confirmed in planta using bimolecular fluorescence complementation. Genetic studies revealed that ABI5 is epistatic to PKS5. PKS5 phosphorylates a serine (Ser) residue at position 42 in ABI5 and regulates ABA-responsive gene expression. This phosphorylation was induced by ABA in vivo and transactivated ABI5. Expression of ABI5, in which Ser-42 was mutated to alanine, could not fully rescue the ABA-insensitive phenotypes of the abi5-8 and pks5-4abi5-8 mutants. In contrast, mutating Ser-42 to aspartate rescued the ABA insensitivity of these mutants. These data demonstrate that PKS5-mediated phosphorylation of ABI5 at Ser-42 is critical for the ABA regulation of seed germination and gene expression in Arabidopsis (Arabidopsis thaliana). PMID:25858916

  5. Lack of chemically induced mutation in repair-deficient mutants of yeast

    International Nuclear Information System (INIS)

    Prakash, L.

    1974-01-01

    Two genes, rad6 and rad9, that confer radiation sensitivity in the yeast Saccharomyces cerevisiae also greatly reduce the frequency of chemically-induced reversions of a tester mutant cyc1-131, which is a chain initiation mutant in the structural gene determining iso-1-cytochrome c. Mutations induced by ethyl methanesulfonate (EMS), diethyl sulfate (DES), methyl methanesulfonate (MMS), dimethyl sulfate (DMS), nitroquinoline oxide (NQO), nitrosoguanidine (NTG), nitrogen mustard (HN2), β-propiolactone, and tritiated uridine, as well as mutations induced by ultraviolet light (UV) and ionizing radiation were greatly diminished in strains homozygous for either the rad6 or rad9 gene. Nitrous acid and nitrosoimidazolidone (NIL), on the other hand, were highly mutagenic in these repair-deficient mutants, and at low doses, these mutagens acted with about the same efficiency as in the normal RAD strain. At high doses of either nitrous acid or NIL, however, reversion frequencies were significantly reduced in the two rad mutants compared to normal strains. Although both rad mutants are immutable to about the same extent, the rad9 strains tend to be less sensitive to the lethal effect of chemical mutagens than rad6 strains. It is concluded that yeast requires a functional repair system for mutation induction by chemical agents. (auth)

  6. Lack of chemically induced mutation in repair-deficient mutants of yeast.

    Science.gov (United States)

    Prakash, L

    1974-12-01

    Two genes, rad6 and rad9, that confer radiation sensitivity in the yeast Saccharomyces cerevisiae also greatly reduce the frequency of chemically-induced reversions of a tester mutant cyc1-131, which is a chain initiation mutant in the structural gene determining iso-1-cytochrome c. Mutations induced by ethyl methanesulfonate (EMS), diethyl sulfate (DES), methyl methanesulfonate (MMS), dimethyl sulfate (DMS), nitroquinoline oxide (NQO), nitrosoguanidine (NTG), nitrogen mustard (HN2), beta-propiolactone, and tritiated uridine, as well as mutations induced by ultraviolet light (UV) and ionizing radiation were greatly diminished in strains homozygous for either the rad6 or rad9 gene. Nitrous acid and nitrosoimidazolidone (NIL), on the other hand, were highly mutagenic in these repair-deficient mutants, and at low doses, these mutagens acted with about the same efficiency as in the normal RAD strain. At high doses of either nitrous acid or NIL, however, reversion frequencies were significantly reduced in the two rad mutants compared to normal strains. Although both rad mutants are immutable to about the same extent, the rad9 strains tend to be less sensitive to the lethal effect of chemical mutagens than rad6 strains. It is concluded that yeast requires a functional repair system for mutation induction by chemical agents.

  7. physical, chemical, technological and biological properties of some mutant oil seeds induced by gamma radiation

    International Nuclear Information System (INIS)

    Ali, H.G.M.

    2003-01-01

    The present study has been undertaken to evaluated sesame, sunflower and safflower seeds induced by gamma rays, as plant breeding unit, plant research department, radioisotope application division, nuclear research center, atomic energy authority Inshas. the obtained results indicate the following : chemical composition of mutant seeds: the radiation mutation caused a significant increase in both oil and ash content total carbohydrates showed a significant decreased in sesame seeds. radiation mutation induced significant increase in oil and protein content of sunflower and safflower seeds. while the total carbohydrate showed a significant decrease. physiochemical properties of oils extracted mutant seeds: the radiation mutation had no real effect on the refractive index and A.V of oils extracted from control and mutant sesame, sunflower and safflower seeds. while it caused a slight increase in red color and P.V. of sesame oil, the thiobarbituric acid (TBA) value of mutant sesame oil was not alter upon radiation mutation, but it induced a slight decrease in TBA of mutant sunflower and safflower oils. the unsaponifiable matter percentage of oils extracted from mutant sesame, sunflower and safflower seeds were slightly increased by radiation mutation .radiation mutation of seeds had no real effect on the total SFA and USFA of sesame oil. however, radiation mutation induced a remarkable changes in fatty acid profiles of sunflower and safflower oil as total SFA decreased, while USFA increased. Uric acid was only detected in oil extracted from mutant sunflower seeds

  8. Variations of L- and D-amino acid levels in the brain of wild-type and mutant mice lacking D-amino acid oxidase activity.

    Science.gov (United States)

    Du, Siqi; Wang, Yadi; Weatherly, Choyce A; Holden, Kylie; Armstrong, Daniel W

    2018-05-01

    D-amino acids are now recognized to be widely present in organisms and play essential roles in biological processes. Some D-amino acids are metabolized by D-amino acid oxidase (DAO), while D-Asp and D-Glu are metabolized by D-aspartate oxidase (DDO). In this study, levels of 22 amino acids and the enantiomeric compositions of the 19 chiral proteogenic entities have been determined in the whole brain of wild-type ddY mice (ddY/DAO +/+ ), mutant mice lacking DAO activity (ddY/DAO -/- ), and the heterozygous mice (ddY/DAO +/- ) using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). No significant differences were observed for L-amino acid levels among the three strains except for L-Trp which was markedly elevated in the DAO +/- and DAO -/- mice. The question arises as to whether this is an unknown effect of DAO inactivity. The three highest levels of L-amino acids were L-Glu, L-Asp, and L-Gln in all the three strains. The lowest L-amino acid level was L-Cys in ddY/DAO +/- and ddY/DAO -/- mice, while L-Trp showed the lowest level in ddY/DAO +/+ mice. The highest concentration of D-amino acid was found to be D-Ser, which also had the highest % D value (~ 25%). D-Glu had the lowest % D value (~ 0.01%) in all the three strains. Significant differences of D-Leu, D-Ala, D-Ser, D-Arg, and D-Ile were observed in ddY/DAO +/- and ddY/DAO -/- mice compared to ddY/DAO +/+ mice. This work provides the most complete baseline analysis of L- and D-amino acids in the brains of ddY/DAO +/+ , ddY/DAO +/- , and ddY/DAO -/- mice yet reported. It also provides the most effective and efficient analytical approach for measuring these analytes in biological samples. This study provides fundamental information on the role of DAO in the brain and may be relevant for future development involving novel drugs for DAO regulation.

  9. Selection of mutants of capsicum annuum induced by gamma ray

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Y. I.; Lee, Y. B. [Korea Atomic Energy Research Institute, Taejeon (Korea, Republic of); Lee, E. K. [Chungnam National Univ., Taejeon (Korea, Republic of)

    1998-06-01

    For induction and selection of mutations of Capsicum annuum L., dry seeds of pure lines No.1 and No.2 were irradiated with gamma ray of 150Gy, 200Gy and 250Gy. Various mutants were selected such as showing early maturity, short plant height, long fruit and chlorophyll mutations. Mutation frequency of No.1 line was 3.4% in the dose of 150Gy, while the frequency of No.2 line was 2.7% in the dose of 250Gy. For selection of resistant mutant to amino acid analog, the optimum concentration of 5-methyltryptophan (5-MT) and S-(2-aminoethyl)-L-cysteine were 25 ppm and 30 ppm, respectively. Four resistant mutant lines to 5-MT were selected among 400 mutant lines.

  10. Revealing differences in metabolic flux distributions between a mutant strain and its parent strain Gluconacetobacter xylinus CGMCC 2955.

    Directory of Open Access Journals (Sweden)

    Cheng Zhong

    Full Text Available A better understanding of metabolic fluxes is important for manipulating microbial metabolism toward desired end products, or away from undesirable by-products. A mutant strain, Gluconacetobacter xylinus AX2-16, was obtained by combined chemical mutation of the parent strain (G. xylinus CGMCC 2955 using DEC (diethyl sulfate and LiCl. The highest bacterial cellulose production for this mutant was obtained at about 11.75 g/L, which was an increase of 62% compared with that by the parent strain. In contrast, gluconic acid (the main byproduct concentration was only 5.71 g/L for mutant strain, which was 55.7% lower than that of parent strain. Metabolic flux analysis indicated that 40.1% of the carbon source was transformed to bacterial cellulose in mutant strain, compared with 24.2% for parent strain. Only 32.7% and 4.0% of the carbon source were converted into gluconic acid and acetic acid in mutant strain, compared with 58.5% and 9.5% of that in parent strain. In addition, a higher flux of tricarboxylic acid (TCA cycle was obtained in mutant strain (57.0% compared with parent strain (17.0%. It was also indicated from the flux analysis that more ATP was produced in mutant strain from pentose phosphate pathway (PPP and TCA cycle. The enzymatic activity of succinate dehydrogenase (SDH, which is one of the key enzymes in TCA cycle, was 1.65-fold higher in mutant strain than that in parent strain at the end of culture. It was further validated by the measurement of ATPase that 3.53-6.41 fold higher enzymatic activity was obtained from mutant strain compared with parent strain.

  11. Abscisic acid negatively regulates elicitor-induced synthesis of capsidiol in wild tobacco.

    Science.gov (United States)

    Mialoundama, Alexis Samba; Heintz, Dimitri; Debayle, Delphine; Rahier, Alain; Camara, Bilal; Bouvier, Florence

    2009-07-01

    In the Solanaceae, biotic and abiotic elicitors induce de novo synthesis of sesquiterpenoid stress metabolites known as phytoalexins. Because plant hormones play critical roles in the induction of defense-responsive genes, we have explored the effect of abscisic acid (ABA) on the synthesis of capsidiol, the major wild tobacco (Nicotiana plumbaginifolia) sesquiterpenoid phytoalexin, using wild-type plants versus nonallelic mutants Npaba2 and Npaba1 that are deficient in ABA synthesis. Npaba2 and Npaba1 mutants exhibited a 2-fold higher synthesis of capsidiol than wild-type plants when elicited with either cellulase or arachidonic acid or when infected by Botrytis cinerea. The same trend was observed for the expression of the capsidiol biosynthetic genes 5-epi-aristolochene synthase and 5-epi-aristolochene hydroxylase. Treatment of wild-type plants with fluridone, an inhibitor of the upstream ABA pathway, recapitulated the behavior of Npaba2 and Npaba1 mutants, while the application of exogenous ABA reversed the enhanced synthesis of capsidiol in Npaba2 and Npaba1 mutants. Concomitant with the production of capsidiol, we observed the induction of ABA 8'-hydroxylase in elicited plants. In wild-type plants, the induction of ABA 8'-hydroxylase coincided with a decrease in ABA content and with the accumulation of ABA catabolic products such as phaseic acid and dihydrophaseic acid, suggesting a negative regulation exerted by ABA on capsidiol synthesis. Collectively, our data indicate that ABA is not required per se for the induction of capsidiol synthesis but is essentially implicated in a stress-response checkpoint to fine-tune the amplification of capsidiol synthesis in challenged plants.

  12. Temperature sensitive riboflavin mutants of Penicillium vermiculatum Dangeard

    International Nuclear Information System (INIS)

    Mitra, J.; Chaudhari, K.L.

    1974-01-01

    Two temperature sensitive UV induced riboflavin mutants rib 1 and rib 6 have been physiologically and genetically characterized. The two mutants behave differently with regard to their temperature sensitivity. The rib 1 mutant exhibits a leaky growth in minimal medium between 15 0 C and 30 0 C but grows well when the medium is supplemented with riboflavin. At 35 0 C the growth response of the mutant is at its max. and at 40 0 C and below 15 0 C it ceases to grow. The rib 6 mutant which is red brown in colour shows wild type character at temp. below 25 0 C in minimal medium but requires riboflavin at 30 0 C and above. Heterokaryotic analysis revealed the nonallelic nature of the two temperature mutants. Genetic tests of allelic relationship between riboflavin markers by crossing were also done. (author)

  13. Probing Phosphorus Efficient Low Phytic Acid Content Soybean Genotypes with Phosphorus Starvation in Hydroponics Growth System.

    Science.gov (United States)

    Kumar, Varun; Singh, Tiratha Raj; Hada, Alkesh; Jolly, Monica; Ganapathi, Andy; Sachdev, Archana

    2015-10-01

    Phosphorus is an essential nutrient required for soybean growth but is bound in phytic acid which causes negative effects on both the environment as well as the animal nutrition. Lowering of phytic acid levels is associated with reduced agronomic characteristics, and relatively little information is available on the response of soybean plants to phosphorus (P) starvation. In this study, we evaluated the effects of different P starvation concentrations on the phytic acid content, growth, and yield of seven mutant genotypes along with the unirradiated control, JS-335, in a hydroponics growth system. The low phytic acid containing mutant genotypes, IR-JS-101, IR-DS-118, and IR-V-101, showed a relatively high growth rate in low P concentration containing nutrient solution (2 μM), whereas the high P concentration (50 μM) favored the growth of IR-DS-111 and IR-DS-115 mutant genotypes containing moderate phytate levels. The mutant genotypes with high phytic acid content, IR-DS-122, IR-DS-114, and JS-335, responded well under P starvation and did not have any significant effect on the growth and yield of plants. Moreover, the reduction of P concentration in nutrient solution from 50 to 2 μM also reduced the phytic acid content in the seeds of all the soybean genotypes under study. The desirable agronomic performance of low phytic acid containing mutant genotype IR-DS-118 reported in this study suggested it to be a P-efficient genotype which could be considered for agricultural practices under P limiting soils.

  14. Mutant human torsinA, responsible for early-onset dystonia, dominantly suppresses GTPCH expression, dopamine levels and locomotion in Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Noriko Wakabayashi-Ito

    2015-07-01

    Full Text Available Dystonia represents the third most common movement disorder in humans with over 20 genetic loci identified. TOR1A (DYT1, the gene responsible for the most common primary hereditary dystonia, encodes torsinA, an AAA ATPase family protein. Most cases of DYT1 dystonia are caused by a 3 bp (ΔGAG deletion that results in the loss of a glutamic acid residue (ΔE302/303 in the carboxyl terminal region of torsinA. This torsinAΔE mutant protein has been speculated to act in a dominant-negative manner to decrease activity of wild type torsinA. Drosophila melanogaster has a single torsin-related gene, dtorsin. Null mutants of dtorsin exhibited locomotion defects in third instar larvae. Levels of dopamine and GTP cyclohydrolase (GTPCH proteins were severely reduced in dtorsin-null brains. Further, the locomotion defect was rescued by the expression of human torsinA or feeding with dopamine. Here, we demonstrate that human torsinAΔE dominantly inhibited locomotion in larvae and adults when expressed in neurons using a pan-neuronal promoter Elav. Dopamine and tetrahydrobiopterin (BH4 levels were significantly reduced in larval brains and the expression level of GTPCH protein was severely impaired in adult and larval brains. When human torsinA and torsinAΔE were co-expressed in neurons in dtorsin-null larvae and adults, the locomotion rates and the expression levels of GTPCH protein were severely reduced. These results support the hypothesis that torsinAΔE inhibits wild type torsinA activity. Similarly, neuronal expression of a Drosophila DtorsinΔE equivalent mutation dominantly inhibited larval locomotion and GTPCH protein expression. These results indicate that both torsinAΔE and DtorsinΔE act in a dominant-negative manner. We also demonstrate that Dtorsin regulates GTPCH expression at the post-transcriptional level. This Drosophila model of DYT1 dystonia provides an important tool for studying the differences in the molecular function between the

  15. Isolation of new gravitropic mutants under hypergravity conditions

    Directory of Open Access Journals (Sweden)

    Akiko Mori

    2016-09-01

    Full Text Available Forward genetics is a powerful approach used to link genotypes and phenotypes, and mutant screening/analysis has provided deep insights into many aspects of plant physiology. Gravitropism is a tropistic response in plants, in which hypocotyls and stems sense the direction of gravity and grow upwards. Previous studies of gravitropic mutants have suggested that shoot endodermal cells in Arabidopsis stems and hypocotyls are capable of sensing gravity (i.e., statocytes. In the present study, we report a new screening system using hypergravity conditions to isolate enhancers of gravitropism mutants, and we also describe a rapid and efficient genome mapping method, using Next-Generation Sequencing (NGS and Single Nucleotide Polymorphism (SNP-based markers. Using the endodermal-amyloplast less 1 (eal1 mutant, which exhibits defective development of endodermal cells and gravitropism, we found that hypergravity (10 g restored the reduced gravity responsiveness in eal1 hypocotyls and could, therefore, be used to obtain mutants with further reduction in gravitropism in the eal1 background. Using the new screening system, we successfully isolated six ene (enhancer of eal1 mutants that exhibited little or no gravitropism under hypergravity conditions, and using NGS and map-based cloning with SNP markers, we narrowed down the potential causative genes, which revealed a new genetic network for shoot gravitropism in Arabidopsis.

  16. Sharing mutants and experimental information prepublication using FgMutantDb (https://scabusa.org/FgMutantDb).

    Science.gov (United States)

    Baldwin, Thomas T; Basenko, Evelina; Harb, Omar; Brown, Neil A; Urban, Martin; Hammond-Kosack, Kim E; Bregitzer, Phil P

    2018-06-01

    There is no comprehensive storage for generated mutants of Fusarium graminearum or data associated with these mutants. Instead, researchers relied on several independent and non-integrated databases. FgMutantDb was designed as a simple spreadsheet that is accessible globally on the web that will function as a centralized source of information on F. graminearum mutants. FgMutantDb aids in the maintenance and sharing of mutants within a research community. It will serve also as a platform for disseminating prepublication results as well as negative results that often go unreported. Additionally, the highly curated information on mutants in FgMutantDb will be shared with other databases (FungiDB, Ensembl, PhytoPath, and PHI-base) through updating reports. Here we describe the creation and potential usefulness of FgMutantDb to the F. graminearum research community, and provide a tutorial on its use. This type of database could be easily emulated for other fungal species. Published by Elsevier Inc.

  17. Generation of a glucose de-repressed mutant of Trichoderma reesei using disparity mutagenesis.

    Science.gov (United States)

    Iwakuma, Hidekazu; Koyama, Yoshiyuki; Miyachi, Ayako; Nasukawa, Masashi; Matsumoto, Hitoshi; Yano, Shuntaro; Ogihara, Jun; Kasumi, Takafumi

    2016-01-01

    We obtained a novel glucose de-repressed mutant of Trichoderma reesei using disparity mutagenesis. A plasmid containing DNA polymerase δ lacking proofreading activity, and AMAI, an autonomously replicating sequence was introduced into T. reesei ATCC66589. The rate of mutation evaluated with 5-fluoroorotic acid resistance was approximately 30-fold higher than that obtained by UV irradiation. The transformants harboring incompetent DNA polymerase δ were then selected on 2-deoxyglucose agar plates with hygromycin B. The pNP-lactoside hydrolyzing activities of mutants were 2 to 5-fold higher than the parent in liquid medium containing glucose. Notably, the amino acid sequence of cre1, a key gene involved in glucose repression, was identical in the mutant and parent strains, and further, the cre1 expression levels was not abolished in the mutant. Taken together, these results demonstrate that the strains of T. reesei generated by disparity mutagenesis are glucose de-repressed variants that contain mutations in yet-unidentified factors other than cre1.

  18. Data supporting mitochondrial morphological changes by SPG13-associated HSPD1 mutants

    Directory of Open Access Journals (Sweden)

    Yuki Miyamoto

    2016-03-01

    Full Text Available The data is related to the research article entitled “Hypomyelinating leukodystrophy-associated missense mutation in HSPD1 blunts mitochondrial dynamics” [1]. In addition to hypomyelinating leukodystrophy (HLD 4 (OMIM no. 612233, it is known that spastic paraplegia (SPG 13 (OMIM no. 605280 is caused by HSPD1’s amino acid mutation. Two amino acid mutations Val-98-to-Ile (V98I and Gln-461-to-Glu (Q461E are associated with SPG13 [2]. In order to investigate the effects of HSPD1’s V98I or Q461E mutant on mitochondrial morphological changes, we transfected each of the respective mutant-encoding genes into Cos-7 cells. Either of V98I or Q461E mutant exhibited increased number of mitochondria and short length mitochondrial morphologies. Using MitoTracker dye-incorporating assay, decreased mitochondrial membrane potential was also observed in both cases. The data described here supports that SPG13-associated HSPD1 mutant participates in causing aberrant mitochondrial morphological changes with decreased activities. Keywords: SPG13, HSPD1, Mitochondrion, Morphological change

  19. The aba mutant of Arabidopsis thaliana is impaired in epoxy-carotenoid biosynthesis

    Energy Technology Data Exchange (ETDEWEB)

    Rock, C.D.; Zeevaart, J.A.D. (Michigan State Univ., East Lansing (United States))

    1991-09-01

    The three mutant alleles of the ABA locus of Arabidopsis thaliana result in plants that are deficient in the plant growth regulator abscisic acid (ABA). The authors have used {sup 18}O{sub 2} to label ABA in water-stressed leaves of mutant and wild-type Arabidopsis. Analysis by selected ion monitoring and tandem mass spectrometry of ({sup 18}O)ABA and its catabolites, phaseic acid and ABA-glucose ester ({beta}-D-glucopyranosyl abscisate), indicates that the aba genotypes are impaired in ABA biosynthesis and have a small ABA precursor pool of compounds that contain oxygens on the rings, presumably oxygenated carotenoids (xanthophylls). Quantitation of the carotenoids form mutant and wild-type leaves establishes that the aba alleles cause a deficiency of the epoxy-carotenoids violaxanthin and neoxanthin and an accumulation of their biosynthetic precursor, zeaxanthin. These results provide evidence that ABA is synthesized by oxidative cleavage of epoxy-carotenoids (the indirect pathway). Furthermore the carotenoid mutant they describe undergoes normal greening. Thus the aba alleles provide an opportunity to study the physiological roles of epoxy-carotenoids in photosynthesis in a higher plants.

  20. The periplasmic transaminase PtaA of Pseudomonas fluorescens converts the glutamic acid residue at the pyoverdine fluorophore to α-ketoglutaric acid.

    Science.gov (United States)

    Ringel, Michael T; Dräger, Gerald; Brüser, Thomas

    2017-11-10

    The periplasmic conversion of ferribactin to pyoverdine is essential for siderophore biogenesis in fluorescent pseudomonads, such as pathogenic Pseudomonas aeruginosa or plant growth-promoting Pseudomonas fluorescens The non-ribosomal peptide ferribactin undergoes cyclizations and oxidations that result in the fluorophore, and a strictly conserved fluorophore-bound glutamic acid residue is converted to a range of variants, including succinamide, succinic acid, and α-ketoglutaric acid residues. We recently discovered that the pyridoxal phosphate-containing enzyme PvdN is responsible for the generation of the succinamide, which can be hydrolyzed to succinic acid. Based on this, a distinct unknown enzyme was postulated to be responsible for the conversion of the glutamic acid to α-ketoglutaric acid. Here we report the identification and characterization of this enzyme in P. fluorescens strain A506. In silico analyses indicated a periplasmic transaminase in fluorescent pseudomonads and other proteobacteria that we termed PtaA for " p eriplasmic t ransaminase A " An in-frame-deleted ptaA mutant selectively lacked the α-ketoglutaric acid form of pyoverdine, and recombinant PtaA complemented this phenotype. The ptaA / pvdN double mutant produced exclusively the glutamic acid form of pyoverdine. PtaA is homodimeric and contains a pyridoxal phosphate cofactor. Mutation of the active-site lysine abolished PtaA activity and affected folding as well as Tat-dependent transport of the enzyme. In pseudomonads, the occurrence of ptaA correlates with the occurrence of α-ketoglutaric acid forms of pyoverdines. As this enzyme is not restricted to pyoverdine-producing bacteria, its catalysis of periplasmic transaminations is most likely a general tool for specific biosynthetic pathways. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Relations between protein production, protein quality and environmental factors in Pisum mutants

    International Nuclear Information System (INIS)

    Gottschalk, W.; Mueller, H.P.; Wolff, G.

    1975-01-01

    The seed protein content of 138 radiation-induced Pisum mutants was determined. The variability of this genetically well-defined material agrees approximately with that of the world collection of Pisum sativum. Some environmental factors to a great extent influence the protein production of the mutants and the initial line. Therefore, it is necessary to consider the relations between the genetically controlled protein production and its dependence upon the environmental factors. This is especially evident if the protein situation of the same genotypes cultivated under the moderate climatic conditions of middle Europe is compared with the subtropical conditions of India. A generally firm correlation between seed size and protein content could not be found in material regarding 148 different mutants of our assortment. Therefore, the selection of small-grained mutants does not result in a selection of protein-rich genotypes in Pisum sativum. Considering all the criteria positively and negatively influencing the protein production, a positive situation could be found in some mutants, especially in the fasciated ones. Furthermore, an improvement of the protein quality could be reached by a genetically conditioned alteration of the globulin-albumin ratio leading to an increase of some essential amino acids such as methionine and lysine. The combined action of mutant genes results in unexpected changes of the protein quantity as well as the quality of the recombinants in relation to their parental mutants. The comparison of some essential amino acids of our useful mutants with those of the varieties of other genera of the Leguminosae shows certain trends of biochemical alterations realized during evolutionary development of the family. (author)

  2. Specific TP53 Mutants Overrepresented in Ovarian Cancer Impact CNV, TP53 Activity, Responses to Nutlin-3a, and Cell Survival

    Directory of Open Access Journals (Sweden)

    Lisa K. Mullany

    2015-10-01

    Full Text Available Evolutionary Action analyses of The Cancer Gene Atlas data sets show that many specific p53 missense and gain-of-function mutations are selectively overrepresented and functional in high-grade serous ovarian cancer (HGSC. As homozygous alleles, p53 mutants are differentially associated with specific loss of heterozygosity (R273; chromosome 17; copy number variation (R175H; chromosome 9; and up-stream, cancer-related regulatory pathways. The expression of immune-related cytokines was selectively related to p53 status, showing for the first time that specific p53 mutants impact, and are related to, the immune subtype of ovarian cancer. Although the majority (31% of HGSCs exhibit loss of heterozygosity, a significant number (24% maintain a wild-type (WT allele and represent another HGSC subtype that is not well defined. Using human and mouse cell lines, we show that specific p53 mutants differentially alter endogenous WT p53 activity; target gene expression; and responses to nutlin-3a, a small molecular that activates WT p53 leading to apoptosis, providing “proof of principle” that ovarian cancer cells expressing WT and mutant alleles represent a distinct ovarian cancer subtype. We also show that siRNA knock down of endogenous p53 in cells expressing homozygous mutant alleles causes apoptosis, whereas cells expressing WT p53 (or are heterozygous for WT and mutant p53 alleles are highly resistant. Therefore, despite different gene regulatory pathways associated with specific p53 mutants, silencing mutant p53 might be a suitable, powerful, global strategy for blocking ovarian cancer growth in those tumors that rely on mutant p53 functions for survival. Knowing p53 mutational status in HGSC should permit new strategies tailored to control this disease.

  3. Silverleaf whitefly induces salicylic acid defenses and suppresses effectual jasmonic acid defenses.

    Science.gov (United States)

    Zarate, Sonia I; Kempema, Louisa A; Walling, Linda L

    2007-02-01

    The basal defenses important in curtailing the development of the phloem-feeding silverleaf whitefly (Bemisia tabaci type B; SLWF) on Arabidopsis (Arabidopsis thaliana) were investigated. Sentinel defense gene RNAs were monitored in SLWF-infested and control plants. Salicylic acid (SA)-responsive gene transcripts accumulated locally (PR1, BGL2, PR5, SID2, EDS5, PAD4) and systemically (PR1, BGL2, PR5) during SLWF nymph feeding. In contrast, jasmonic acid (JA)- and ethylene-dependent RNAs (PDF1.2, VSP1, HEL, THI2.1, FAD3, ERS1, ERF1) were repressed or not modulated in SLWF-infested leaves. To test for a role of SA and JA pathways in basal defense, SLWF development on mutant and transgenic lines that constitutively activate or impair defense pathways was determined. By monitoring the percentage of SLWF nymphs in each instar, we show that mutants that activate SA defenses (cim10) or impair JA defenses (coi1) accelerated SLWF nymphal development. Reciprocally, mutants that activate JA defenses (cev1) or impair SA defenses (npr1, NahG) slowed SLWF nymphal development. Furthermore, when npr1 plants, which do not activate downstream SA defenses, were treated with methyl jasmonate, a dramatic delay in nymph development was observed. Collectively, these results showed that SLWF-repressed, JA-regulated defenses were associated with basal defense to the SLWF.

  4. Enhanced oxidative stress in the jasmonic acid-deficient tomato mutant def-1 exposed to NaCl stress.

    Science.gov (United States)

    Abouelsaad, Ibrahim; Renault, Sylvie

    2018-04-21

    Jasmonic acid (JA) has been mostly studied in responses to biotic stresses, such as herbivore attack and pathogenic infection. More recently, the involvement of JA in abiotic stresses including salinity was highlighted; yet, its role in salt stress remained unclear. In the current study, we compared the physiological and biochemical responses of wild-type (WT) tomato (Solanum lycopersicum) cv Castlemart and its JA-deficient mutant defenseless-1 (def-1) under salt stress to investigate the role of JA. Plant growth, photosynthetic pigment content, ion accumulation, oxidative stress-related parameters, proline accumulation and total phenolic compounds, in addition to both enzymatic and non-enzymatic antioxidant activities, were measured in both genotypes after 14 days of 100 mM NaCl treatment. Although we observed in both genotypes similar growth pattern and sodium, calcium and potassium levels in leaves under salt stress, def-1 plants exhibited a more pronounced decrease of nitrogen content in both leaves and roots and a slightly higher level of sodium in roots compared to WT plants. In addition, def-1 plants exposed to salt stress showed reactive oxygen species (ROS)-associated injury phenotypes. These oxidative stress symptoms in def-1 were associated with lower activity of both enzymatic antioxidants and non-enzymatic antioxidants. Furthermore, the levels of the non-enzymatic ROS scavengers proline and total phenolic compounds increased in both genotypes exposed to salt stress, with a higher amount of proline in the WT plants. Overall the results of this study suggest that endogenous JA mainly enhanced tomato salt tolerance by maintaining ROS homeostasis. Copyright © 2018 Elsevier GmbH. All rights reserved.

  5. Screening of allyl alcohol resistant mutant of Rhizopus oryzae and ...

    African Journals Online (AJOL)

    Ethanol is a main by-product in the fermentation broth of Rhizopus oryzae during the production of high-optical purity L-lactic acid. By screening the lower activity of alcohol dehydrogenase (ADH) mutant, thus decreasing the flux of pyruvic acid to ethanol may be a virtual method for increasing the conversion rate of glucose ...

  6. GPNMB ameliorates mutant TDP-43-induced motor neuron cell death.

    Science.gov (United States)

    Nagahara, Yuki; Shimazawa, Masamitsu; Ohuchi, Kazuki; Ito, Junko; Takahashi, Hitoshi; Tsuruma, Kazuhiro; Kakita, Akiyoshi; Hara, Hideaki

    2017-08-01

    Glycoprotein nonmetastatic melanoma protein B (GPNMB) aggregates are observed in the spinal cord of amyotrophic lateral sclerosis (ALS) patients, but the detailed localization is still unclear. Mutations of transactive response DNA binding protein 43kDa (TDP-43) are associated with neurodegenerative diseases including ALS. In this study, we evaluated the localization of GPNMB aggregates in the spinal cord of ALS patients and the effect of GPNMB against mutant TDP-43 induced motor neuron cell death. GPNMB aggregates were not localized in the glial fibrillary acidic protein (GFAP)-positive astrocyte and ionized calcium binding adaptor molecule-1 (Iba1)-positive microglia. GPNMB aggregates were localized in the microtubule-associated protein 2 (MAP-2)-positive neuron and neurofilament H non-phosphorylated (SMI-32)-positive neuron, and these were co-localized with TDP-43 aggregates in the spinal cord of ALS patients. Mock or TDP-43 (WT, M337V, and A315T) plasmids were transfected into mouse motor neuron cells (NSC34). The expression level of GPNMB was increased by transfection of mutant TDP-43 plasmids. Recombinant GPNMB ameliorated motor neuron cell death induced by transfection of mutant TDP-43 plasmids and serum-free stress. Furthermore, the expression of phosphorylated ERK1/2 and phosphorylated Akt were decreased by this stress, and these expressions were increased by recombinant GPNMB. These results indicate that GPNMB has protective effects against mutant TDP-43 stress via activating the ERK1/2 and Akt pathways, and GPNMB may be a therapeutic target for TDP-43 proteinopathy in familial and sporadic ALS. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  7. Isolation and analysis of lipase-overproducing mutants of Serratia marcescens.

    Science.gov (United States)

    Kawai, E; Akatsuka, H; Sakurai, N; Idei, A; Matsumae, H; Shibatani, T; Komatsubara, S; Omori, K

    2001-01-01

    We have isolated a lipase-overproducing mutant, GE14, from Serratia marcescens 8000 after three rounds of N-methyl-N'-nitro-N-nitrosoguanidine mutagenesis. The mutant GE14 produced 95 kU/ml of extracellular lipase in the lipase medium, which was about threefold higher than that of produced by the original strain 8000. Enzymatic characteristics including specific activity of purified lipases from culture supernatants of GE14 and 8000 were almost same. The lipase gene (lipA) of GE14 contained two base substitutions; one in the promoter region and another in the N-terminal region of the lipA gene without an amino acid substitution. Promoter analysis using lipA-lacZ fusion plasmids revealed that these substitutions were responsible for the increase in the lipA expression level, independently. In contrast, no base substitution was found in the genes encoding the lipase secretion device, the Lip system. In addition, the genes coding for metalloprotease and the cell surface layer protein which are both secreted through the Lip system and associated with extracellular lipase production, also contained no base substitution. The strain GE14 carrying a high-copy-number lipA plasmid produced a larger amount of the extracellular lipase than the recombinant strains of 8000 and other mutants also did, indicating that GE14 was not only a lipase-overproducing strain, but also an advantageous host strain for overproducing the lipase by a recombinant DNA technique. These results suggest that the lipase-overproducing mutant GE14 and its recombinant strains are promising candidates for the industrial production of the S. marcescens lipase.

  8. Bioethanol production using genetically modified and mutant wheat and barley straws

    Energy Technology Data Exchange (ETDEWEB)

    Li, Z. [Washington State Univ., Pullman, WA (US). Dept. of Biological Engineering; East China Univ. of Science and Technology, Shanghai (CN). State Key Laboratory of Bioreactor Engineering; Liu, Y. [Michigan State Univ., East Lansing, MI (US). Biosystems and Agricultural Engineering; Chen, S. [Washington State Univ., Pullman, WA (US). Dept. of Biological Systems Engineering; Zemetra, R.S. [Univ. of Idaho, Moscow, ID (US). Plant, Soil, and Entomological Sciences

    2011-01-15

    To improve the performance of wheat and barley straws as feedstocks for ethanol biorefining, the genetic modifications of down regulating Cinnamoyl-CoA reductase and low phytic acid mutation have been introduced into wheat and barley respectively. In this study, total 252 straw samples with different genetic background and location were collected from the field experiment based on a randomized complete block design. The fiber analysis (neutral detergent fiber, acid detergent fiber, and acid detergent lignin) indicated that there were no significant differences between modified and wild type straw lines in terms of straw compositions. However, the difference did exist among straw lines on fiber utilization. 16 straw samples were further selected to conduct diluted acid pretreatment, enzymatic hydrolysis and fermentation. The data indicated that the phytic acid mutant and transgenic straws have changed the fiber structure, which significantly influences their hydrolysibility. These results may lead to a possible solution of mutant or genetic modified plant species that is capable to increase the hydrolysibility of biomass without changing their compositions and sacrificing their agronomy performance. (author)

  9. Properties of adenyl cyclase and cyclic adenosine 3',5'-monophosphate receptor protein-deficient mutants of Escherichia coli

    International Nuclear Information System (INIS)

    Kumar, S.

    1976-01-01

    Several spontaneous cya and crp mutants of Escherichia coli have been selected as clones simultaneously resistant to phage lambda and nalidixic acid and characterized. Both cya and crp mutants have been found to grow as cocci with increased doubling times. They have increased resistance to some mutagens (methylmethanesulfonate, ultraviolet light, gamma rays), antibiotics (nalidixic acid, ampicillin), phages (lambda, T6), sublethal heat and hypotonic shock, and decreased resistance to neutral detergents (sodium dodecyl sulfate, sodium deoxycholate), a protein synthesis inhibitor (streptomycin), and a respiratory inhibitor (sodium azide). The nature of changes in cell parameters indicate fundamental alterations in the envelope structure of the cya and crp mutant cells. The new cya and crp mutants have been found to be multiply carbohydrate negative and nonmotile in conformity with similar previously isolated mutants. Studies of revertants and phi 80 cya + and phi 80 cya transductants indicated that the pleiotropic phenotype is related to a single mutational event at the cya or the crp locus in the mutants

  10. Natural modifiers of seed longevity in the Arabidopsis mutants abscisic acid insensitive3-5 (abi3-5) and leafy cotyledon1-3 (lec1-3)

    NARCIS (Netherlands)

    Sugliani, M.R.L.; Rajjou, L.; Clerkx, E.J.M.; Koornneef, M.; Soppe, W.J.J.

    2009-01-01

    • Seed longevity is an important trait in many crops and is essential for the success of most land plant species. Current knowledge of its molecular regulation is limited. The Arabidopsis mutants abscisic acid insensitive3-5 (abi3-5) and leafy cotyledon1-3 (lec1-3) have impaired seed maturation and

  11. Salicylic acid signaling inhibits apoplastic reactive oxygen species signaling.

    Science.gov (United States)

    Xu, Enjun; Brosché, Mikael

    2014-06-04

    Reactive oxygen species (ROS) are used by plants as signaling molecules during stress and development. Given the amount of possible challenges a plant face from their environment, plants need to activate and prioritize between potentially conflicting defense signaling pathways. Until recently, most studies on signal interactions have focused on phytohormone interaction, such as the antagonistic relationship between salicylic acid (SA)-jasmonic acid and cytokinin-auxin. In this study, we report an antagonistic interaction between SA signaling and apoplastic ROS signaling. Treatment with ozone (O3) leads to a ROS burst in the apoplast and induces extensive changes in gene expression and elevation of defense hormones. However, Arabidopsis thaliana dnd1 (defense no death1) exhibited an attenuated response to O3. In addition, the dnd1 mutant displayed constitutive expression of defense genes and spontaneous cell death. To determine the exact process which blocks the apoplastic ROS signaling, double and triple mutants involved in various signaling pathway were generated in dnd1 background. Simultaneous elimination of SA-dependent and SA-independent signaling components from dnd1 restored its responsiveness to O3. Conversely, pre-treatment of plants with SA or using mutants that constitutively activate SA signaling led to an attenuation of changes in gene expression elicited by O3. Based upon these findings, we conclude that plants are able to prioritize the response between ROS and SA via an antagonistic action of SA and SA signaling on apoplastic ROS signaling.

  12. Abscisic acid negatively regulates post-penetration resistance of Arabidopsis to the biotrophic powdery mildew fungus.

    Science.gov (United States)

    Xiao, Xiang; Cheng, Xi; Yin, Kangquan; Li, Huali; Qiu, Jin-Long

    2017-08-01

    Pytohormone abscisic acid (ABA) plays important roles in defense responses. Nonetheless, how ABA regulates plant resistance to biotrophic fungi remains largely unknown. Arabidopsis ABA-deficient mutants, aba2-1 and aba3-1, displayed enhanced resistance to the biotrophic powdery mildew fungus Golovinomyces cichoracearum. Moreover, exogenously administered ABA increased the susceptibility of Arabidopsis to G. cichoracearum. Arabidopsis ABA perception components mutants, abi1-1 and abi2-1, also displayed similar phenotypes to ABA-deficient mutants in resistance to G. cichoracearum. However, the resistance to G. cichoracearum is not changed in downstream ABA signaling transduction mutants, abi3-1, abi4-1, and abi5-1. Microscopic examination revealed that hyphal growth and conidiophore production of G. cichoracearum were compromised in the ABA deficient mutants, even though pre-penetration and penetration growth of the fungus were not affected. In addition, salicylic acid (SA) and MPK3 are found to be involved in ABA-regulated resistance to G. cichoracearum. Our work demonstrates that ABA negatively regulates post-penetration resistance of Arabidopsis to powdery mildew fungus G. cichoracearum, probably through antagonizing the function of SA.

  13. Epididymis response partly compensates for spermatozoa oxidative defects in snGPx4 and GPx5 double mutant mice.

    Directory of Open Access Journals (Sweden)

    Anaïs Noblanc

    Full Text Available We report here that spermatozoa of mice lacking both the sperm nucleus glutathione peroxidase 4 (snGPx4 and the epididymal glutathione peroxidase 5 (GPx5 activities display sperm nucleus structural abnormalities including delayed and defective nuclear compaction, nuclear instability and DNA damage. We show that to counteract the GPx activity losses, the epididymis of the double KO animals mounted an antioxydant response resulting in a strong increase in the global H(2O(2-scavenger activity especially in the cauda epididymis. Quantitative RT-PCR data show that together with the up-regulation of epididymal scavengers (of the thioredoxin/peroxiredoxin system as well as glutathione-S-transferases the epididymis of double mutant animals increased the expression of several disulfide isomerases in an attempt to recover normal disulfide-bridging activity. Despite these compensatory mechanisms cauda-stored spermatozoa of double mutant animals show high levels of DNA oxidation, increased fragmentation and greater susceptibility to nuclear decondensation. Nevertheless, the enzymatic epididymal salvage response is sufficient to maintain full fertility of double KO males whatever their age, crossed with young WT female mice.

  14. Amino acid substitutions in subunit 9 of the mitochondrial ATPase complex of Saccharomyces cerevisiae. Sequence analysis of a series of revertants of an oli1 mit- mutant carrying an amino acid substitution in the hydrophilic loop of subunit 9.

    Science.gov (United States)

    Willson, T A; Nagley, P

    1987-09-01

    This work concerns a biochemical genetic study of subunit 9 of the mitochondrial ATPase complex of Saccharomyces cerevisiae. Subunit 9, encoded by the mitochondrial oli1 gene, contains a hydrophilic loop connecting two transmembrane stems. In one particular oli1 mit- mutant 2422, the substitution of a positively charged amino acid in this loop (Arg39----Met) renders the ATPase complex non-functional. A series of 20 revertants, selected for their ability to grow on nonfermentable substrates, has been isolated from mutant 2422. The results of DNA sequence analysis of the oli1 gene in each revertant have led to the recognition of three groups of revertants. Class I revertants have undergone a same-site reversion event: the mutant Met39 is replaced either by arginine (as in wild-type) or lysine. Class II revertants maintain the mutant Met39 residue, but have undergone a second-site reversion event (Asn35----Lys). Two revertants showing an oligomycin-resistant phenotype carry this same second-site reversion in the loop region together with a further amino acid substitution in either of the two membrane-spanning segments of subunit 9 (either Gly23----Ser or Leu53----Phe). Class III revertants contain subunit 9 with the original mutant 2422 sequence, and additionally carry a recessive nuclear suppressor, demonstrated to represent a single gene. The results on the revertants in classes I and II indicate that there is a strict requirement for a positively charged residue in the hydrophilic loop close to the boundary of the lipid bilayer. The precise location of this positive charge is less stringent; in functional ATPase complexes it can be found at either residue 39 or 35. This charged residue is possibly required to interact with some other component of the mitochondrial ATPase complex. These findings, together with hydropathy plots of subunit 9 polypeptides from normal, mutant and revertant strains, led to the conclusion that the hydrophilic loop in normal subunit 9

  15. Interaction between sugar and abscisic acid signalling during early seedling development in Arabidopsis

    NARCIS (Netherlands)

    Dekkers, B.J.W.; Schuurmans, J.A.M.J.; Smeekens, J.C.M.

    2008-01-01

    Sugars regulate important processes and affect the expression of many genes in plants. Characterization of Arabidopsis (Arabidopsis thaliana) mutants with altered sugar sensitivity revealed the function of abscisic acid (ABA) signalling in sugar responses. However, the exact interaction between

  16. A chilling sensitive mutant of Arabidopsis with altered steryl-ester metabolism

    International Nuclear Information System (INIS)

    Hugly, S.; McCourt, P.; Somerville, C.; Browse, J.; Patterson, G.W.

    1990-01-01

    A chilling-sensitive mutant of Arabidopsis thaliana was isolated and subjected to genetic, physiological, and biochemical analysis. The chilling-sensitive nature of the mutant line is due to a single recessive nuclear mutation at a locus designated chs1. In contrast to wild-type plants, which are not adversely affected by low temperatures, the chs1 mutant is killed by several days of exposure to temperatures below 18 degree C. Following exposure to chilling temperatures, the mutant displays two common symptoms of chilling injury - leaf chlorosis and electrolyte leakage. In these respects, the physiological response of the mutant to low temperatures mimics the response observed in some naturally occurring chilling sensitive species. The biochemical basis of chilling sensitivity was explored by examining the pattern of incorporation of 14 CO 2 into soluble metabolites and lipids in wild-type and mutant plants. The only difference observed between the mutant and wild type was that following low temperature treatment, the mutant accumulated 10-fold more radioactivity in a specific class of neutral lipids which were identified by a variety of criteria to be steryl-esters. The accumulation of radioactivity in the steryl-ester fraction occurs 24 hours before there is any visible evidence of chilling injury

  17. Abscisic Acid Negatively Regulates Elicitor-Induced Synthesis of Capsidiol in Wild Tobacco1[W

    Science.gov (United States)

    Mialoundama, Alexis Samba; Heintz, Dimitri; Debayle, Delphine; Rahier, Alain; Camara, Bilal; Bouvier, Florence

    2009-01-01

    In the Solanaceae, biotic and abiotic elicitors induce de novo synthesis of sesquiterpenoid stress metabolites known as phytoalexins. Because plant hormones play critical roles in the induction of defense-responsive genes, we have explored the effect of abscisic acid (ABA) on the synthesis of capsidiol, the major wild tobacco (Nicotiana plumbaginifolia) sesquiterpenoid phytoalexin, using wild-type plants versus nonallelic mutants Npaba2 and Npaba1 that are deficient in ABA synthesis. Npaba2 and Npaba1 mutants exhibited a 2-fold higher synthesis of capsidiol than wild-type plants when elicited with either cellulase or arachidonic acid or when infected by Botrytis cinerea. The same trend was observed for the expression of the capsidiol biosynthetic genes 5-epi-aristolochene synthase and 5-epi-aristolochene hydroxylase. Treatment of wild-type plants with fluridone, an inhibitor of the upstream ABA pathway, recapitulated the behavior of Npaba2 and Npaba1 mutants, while the application of exogenous ABA reversed the enhanced synthesis of capsidiol in Npaba2 and Npaba1 mutants. Concomitant with the production of capsidiol, we observed the induction of ABA 8′-hydroxylase in elicited plants. In wild-type plants, the induction of ABA 8′-hydroxylase coincided with a decrease in ABA content and with the accumulation of ABA catabolic products such as phaseic acid and dihydrophaseic acid, suggesting a negative regulation exerted by ABA on capsidiol synthesis. Collectively, our data indicate that ABA is not required per se for the induction of capsidiol synthesis but is essentially implicated in a stress-response checkpoint to fine-tune the amplification of capsidiol synthesis in challenged plants. PMID:19420326

  18. A new method for the construction of a mutant library with a predictable occurrence rate using Poisson distribution.

    Science.gov (United States)

    Seong, Ki Moon; Park, Hweon; Kim, Seong Jung; Ha, Hyo Nam; Lee, Jae Yung; Kim, Joon

    2007-06-01

    A yeast transcriptional activator, Gcn4p, induces the expression of genes that are involved in amino acid and purine biosynthetic pathways under amino acid starvation. Gcn4p has an acidic activation domain in the central region and a bZIP domain in the C-terminus that is divided into the DNA-binding motif and dimerization leucine zipper motif. In order to identify amino acids in the DNA-binding motif of Gcn4p which are involved in transcriptional activation, we constructed mutant libraries in the DNA-binding motif through an innovative application of random mutagenesis. Mutant library made by oligonucleotides which were mutated randomly using the Poisson distribution showed that the actual mutation frequency was in good agreement with expected values. This method could save the time and effort to create a mutant library with a predictable mutation frequency. Based on the studies using the mutant libraries constructed by the new method, the specific residues of the DNA-binding domain in Gcn4p appear to be involved in the transcriptional activities on a conserved binding site.

  19. Mutants of Yarrowia lipolytica NCIM 3589 grown on waste cooking oil as a biofactory for biodiesel production.

    Science.gov (United States)

    Katre, Gouri; Ajmera, Namasvi; Zinjarde, Smita; RaviKumar, Ameeta

    2017-10-24

    Oleaginous yeasts are fast emerging as a possible feedstock for biodiesel production. Yarrowia lipolytica, a model oleaginous yeast is known to utilize a variety of hydrophobic substrates for lipid accumulation including waste cooking oil (WCO). Approaches to increase lipid content in this yeast include metabolic engineering which requires manipulation of multiple genes in the lipid biosynthesis pathway. A classical and cost-effective approach, namely, random chemical mutagenesis on the yeast can lead to increased production of biodiesel as is explored here. In this study, chemical mutagenesis using the alkylating agent, N- methyl-N'-nitro-N-nitrosoguanidine (MNNG) as well as an additional treatment with cerulenin, a fatty acid synthase inhibitor generated 800 mutants of Y. lipolytica NCIM 3589 (761 MNNG treated and 39 MNNG + cerulenin treated). A three-stage screening using Sudan Black B plate technique, Nile red fluorimetry and total lipid extraction using solvent was performed, which enabled selection of ten high lipid yielding mutants. Time course studies of all the ten mutants were further undertaken in terms of biomass, lipid yield and lipid content to select three stable mutants (YlB6, YlC7 and YlE1) capable of growing and accumulating lipid on WCO, with lipid contents of 55, 60 and 67% as compared to 45% for the wild type. The mutants demonstrated increased volumetric lipid productivities (0.062, 0.044 and 0.041 g L -1  h -1 ) as compared to the wild type (0.033 g L -1  h -1 ). The fatty acid profile of the three mutants consisted of a high content of C16 and C18 saturated and monounsaturated fatty acids and was found to be suitable for biodiesel production. The fuel properties, namely, density, kinematic viscosity, total acid number, iodine value of the three mutants were evaluated and found to lie within the limits specified by internationally accepted standards. Additionally, it was noted that the mutants demonstrated better cetane numbers and

  20. Reduction of FR900525 using an S-(2-aminoethyl) l-cysteine-resistant mutant.

    Science.gov (United States)

    Shimizu, Shiho; Futase, Ayako; Yokoyama, Tatsuya; Ueda, Satoshi; Honda, Hiroyuki

    2017-06-01

    FK506 (tacrolimus), a macrolide compound with immunosuppressant activity, has been proven to have clinical importance and has been manufactured industrially since 1993 by using mutants with high FK506-production ability; these mutants have been developed from the wild strain Streptomyces tsukubaensis No. 9993. FR900525 is one of the by-products of FK506 production. However, there was no effective industrial method to separate FR900525 from FK506 due to the structural similarity between the two compounds. Therefore, reducing the level of FR900525 was a serious problem in the industrial strain A. In this study, we aimed to reduce the FR900525 production. We first determined that pipecolic acid level was a critical parameter for controlling FR900525 production in strain A. S-(2-Aminoethyl) l-cysteine (AEC)-resistant mutants has been reported to increase lysine productivity successfully in a variety of lysine-producing microorganisms. Therefore, next, we applied a selection of AEC-resistant mutants to enhance pipecolic acid biosynthesis. Finally, four AEC-resistant mutants were obtained from strain A using ultraviolet irradiation, and three of them showed less FR900525 productivity compared to the parental strain A. Our findings indicated that AEC resistance was effective phenotype marker for increasing pipecolic acid productivity and for reducing FR900525 production in S. tsukubaensis. Thus, our study provides an efficient method for reducing FR90025 level during FK506 biosynthesis. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  1. Mutants induced in winter rye (Secale cereale L.): Short straw-mutant No. 2714 and late-senescence mutant

    Energy Technology Data Exchange (ETDEWEB)

    Muszynski, S; Darlewska, M [Department of Plant Breeding and Seed Science, Warsaw Agricultural University, Warsaw (Poland)

    1990-01-01

    Full text: Mutants were induced by treating dormant seeds with ionizing radiation (fast neutrons) or chemicals (N-nitroso-N-ethyl urea or sodium azide). Among several mutants obtained, of special value is the short-straw mutant No. 2714 and a late senescent mutant. (author)

  2. Global regulation of virulence and the stress response by CsrA in the highly adapted human gastric pathogen Helicobacter pylori

    DEFF Research Database (Denmark)

    Barnard, F.M.; Loughlin, M.F.; Fainberg, H.P.

    2004-01-01

    -induced transcriptional responses of napA and ahpC, the acid induction of napA, cagA, vacA, the urease operon, and fur, as well as the heat shock responses of napA, groESL and hspR. Although the level of napA transcript was higher in the csrA mutant, its stability was similar in the wild-type and mutant strains, and less...

  3. Feeding by whiteflies suppresses downstream jasmonic acid signaling by eliciting salicylic acid signaling.

    Science.gov (United States)

    Zhang, Peng-Jun; Li, Wei-Di; Huang, Fang; Zhang, Jin-Ming; Xu, Fang-Cheng; Lu, Yao-Bin

    2013-05-01

    Phloem-feeding whiteflies in the species complex Bemisia tabaci cause extensive crop damage worldwide. One of the reasons for their "success" is their ability to suppress the effectual jasmonic acid (JA) defenses of the host plant. However, little is understood about the mechanisms underlying whitefly suppression of JA-regulated defenses. Here, we showed that the expression of salicylic acid (SA)-responsive genes (EDS1 and PR1) in Arabidopsis thaliana was significantly enhanced during feeding by whitefly nymphs. Whereas upstream JA-responsive genes (LOX2 and OPR3) also were induced, the downstream JA-responsive gene (VSP1) was repressed, i.e., whiteflies only suppressed downstream JA signaling. Gene-expression analyses with various Arabidopsis mutants, including NahG, npr-1, ein2-1, and dde2-2, revealed that SA signaling plays a key role in the suppression of downstream JA defenses by whitefly feeding. Assays confirmed that SA activation enhanced whitefly performance by suppressing downstream JA defenses.

  4. Zoospore exudates from Phytophthora nicotianae affect immune responses in Arabidopsis.

    Science.gov (United States)

    Kong, Ping; McDowell, John M; Hong, Chuanxue

    2017-01-01

    Zoospore exudates play important roles in promoting zoospore communication, homing and germination during plant infection by Phytophthora. However, it is not clear whether exudates affect plant immunity. Zoospore-free fluid (ZFF) and zoospores of P. nicotianae were investigated comparatively for effects on resistance of Arabidopsis thaliana Col-0 and mutants that affect signaling mediated by salicylic acid (SA) and jasmonic acid (JA): eds16 (enhanced disease susceptibility16), pad4 (phytoalexin deficient4), and npr1 (nonexpressor of pathogenesis-related genes1). Col-0 attracted more zoospores and had severe tissue damage when flooded with a zoospore suspension in ZFF. Mutants treated with ZFF alone developed disease symptoms similar to those inoculated with zoospores and requirements of EDS16 and PAD4 for plant responses to zoospores and the exudates was apparent. Zoospore and ZFFs also induced expression of the PR1 and PDF1.2 marker genes for defense regulated by SA and JA, respectively. However, ZFF affected more JA defense signaling, down regulating PR1 when SA signaling or synthesis is deficient, which may be responsible for Arabidopsis mutant plants more susceptible to infection by high concentration of P. nicotianae zoospores. These results suggest that zoospore exudates can function as virulence factors and inducers of plant immune responses during plant infection by Phytophthora.

  5. Induced protein polymorphisms and nutritional quality of gamma irradiation mutants of sorghum

    Energy Technology Data Exchange (ETDEWEB)

    Mehlo, Luke, E-mail: LMehlo@csir.co.za [CSIR Biosciences, Meiring Naude Road, P.O. Box 395, Pretoria 0001 (South Africa); Mbambo, Zodwa [CSIR Biosciences, Meiring Naude Road, P.O. Box 395, Pretoria 0001 (South Africa); Microbiology Discipline, School of Life Sciences, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban 4000 (South Africa); Bado, Souleymane [Plant Breeding and Genetics Laboratory – Joint FAO/IAEA Agriculture and Biotechnology Laboratory, International Atomic Energy Agency Laboratories, A-2444 Seibersdorf (Austria); Lin, Johnson [Microbiology Discipline, School of Life Sciences, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban 4000 (South Africa); Moagi, Sydwell M.; Buthelezi, Sindisiwe; Stoychev, Stoyan; Chikwamba, Rachel [CSIR Biosciences, Meiring Naude Road, P.O. Box 395, Pretoria 0001 (South Africa)

    2013-09-15

    Highlights: • We analyse kafirin protein polymorphisms induced by gamma irradiation in sorghum. • One mutant with suppressed kafirins in the endosperm accumulated them in the germ. • Kafirin polymorphisms were associated with high levels of free amino acids. • Nutritional value of sorghum can be improved significantly by induced mutations. - Abstract: Physical and biochemical analysis of protein polymorphisms in seed storage proteins of a mutant population of sorghum revealed a mutant with redirected accumulation of kafirin proteins in the germ. The change in storage proteins was accompanied by an unusually high level accumulation of free lysine and other essential amino acids in the endosperm. This mutant further displayed a significant suppression in the synthesis and accumulation of the 27 kDa γ-, 24 kDa α-A1 and the 22 kDa α-A2 kafirins in the endosperm. The suppression of kafirins was counteracted by an upsurge in the synthesis and accumulation of albumins, globulins and other proteins. The data collectively suggest that sorghum has huge genetic potential for nutritional biofortification and that induced mutations can be used as an effective tool in achieving premium nutrition in staple cereals.

  6. Induced protein polymorphisms and nutritional quality of gamma irradiation mutants of sorghum

    International Nuclear Information System (INIS)

    Mehlo, Luke; Mbambo, Zodwa; Bado, Souleymane; Lin, Johnson; Moagi, Sydwell M.; Buthelezi, Sindisiwe; Stoychev, Stoyan; Chikwamba, Rachel

    2013-01-01

    Highlights: • We analyse kafirin protein polymorphisms induced by gamma irradiation in sorghum. • One mutant with suppressed kafirins in the endosperm accumulated them in the germ. • Kafirin polymorphisms were associated with high levels of free amino acids. • Nutritional value of sorghum can be improved significantly by induced mutations. - Abstract: Physical and biochemical analysis of protein polymorphisms in seed storage proteins of a mutant population of sorghum revealed a mutant with redirected accumulation of kafirin proteins in the germ. The change in storage proteins was accompanied by an unusually high level accumulation of free lysine and other essential amino acids in the endosperm. This mutant further displayed a significant suppression in the synthesis and accumulation of the 27 kDa γ-, 24 kDa α-A1 and the 22 kDa α-A2 kafirins in the endosperm. The suppression of kafirins was counteracted by an upsurge in the synthesis and accumulation of albumins, globulins and other proteins. The data collectively suggest that sorghum has huge genetic potential for nutritional biofortification and that induced mutations can be used as an effective tool in achieving premium nutrition in staple cereals

  7. Selection and agronomic evaluation of induced mutant lines of sesame

    International Nuclear Information System (INIS)

    Hoballah, A.A.

    2001-01-01

    Station yield trial: Three high yielding mutants (8, 48, and EFM92) with better and stable performance were developed in our breeding programme and submitted for registration to the Agricultural Research Center (ARC), Egyptian Ministry of Agriculture and Land Reclamation. Multi-location yield trials indicated that mutant line EFM92 ranked first in all locations; significant yield increases recorded for it ranged from 14.7 to 74.0% over the check variety. Moreover, it was 15-20 days earlier than the check and/or other mutants. Mutant lines 8 and 48 produced higher seed yields than the check at two different locations. These mutants can probably be grown and produce more yield than the check variety at the low yielding environments. Seed quality assay: During 1996 and 1997, 15 promising lines of sesame including mutants and hybrid populations as well as the local variety were evaluated for seed protein, oil content and fatty acid composition. The protein content varied from 20.6 to 26.7%; hybrid population EXM90 gave the highest value. About 85% of the total fatty acids in the oil are unsaturated (oleic and linoleic) and 15% saturated, mainly palmitic and stearic. Linoleic acid ranged from 41.8 to 47.9%. Mutant lines 6, 9, and EFM92, which gave high oil content (54-55.5%) together with high linoleic acid values (45.2-47.8%), are recommended for breeding for seed oil quality. Heterosis, combining ability and type of gene action in sesame: A half diallel set of crosses involving seven parents was used to study heterosis and combining ability in the F 1 generation as well as the nature of gene action controlling seed yield and its contributing traits in both F 1 and F 2 in order to identify the most efficient breeding methods leading to rapid genetic improvement. The expressions of heterosis varied with the crosses and characters investigated. The maximal significant positive useful heterosis was observed for branches/plant (52.9%) followed by seed yield/plant (38

  8. Zebrafish mutants in the von Hippel-Lindau tumor suppressor display a hypoxic response and recapitulate key aspects of Chuvash polycythemia

    NARCIS (Netherlands)

    van Rooijen, E.; Voest, E.E.; Logister, I.; Korving, J.; Schwerte, T.; Schulte-Merker, S.; Giles, R.H.; van Eeden, F.J.

    2009-01-01

    We have generated 2 zebrafish lines carrying inactivating germline mutations in the von Hippel-Lindau (VHL) tumor suppressor gene ortholog vhl. Mutant embryos display a general systemic hypoxic response, including the up-regulation of hypoxia-induced genes by 1 day after fertilization and a severe

  9. Inhibition of citric acid accumulation by manganese ions in Aspergillus niger mutants with reduced citrate control of phosphofructokinase

    Energy Technology Data Exchange (ETDEWEB)

    Schreferl, G.; Kubicek, C.P.; Roehr, M.

    1986-03-01

    Mutant strains of Aspergillus niger with reduced citrate control of carbohydrate catabolism (cic mutants) grow faster than the parent strain on media containing 5% (wt/vol) citrate. The mutants tolerated a higher intracellular citrate concentration than the parent strain. One mutant (cic-7/3) contained phosphofructokinase activity significantly less sensitive towards citrate than the enzyme from the parent strain. When this mutant was grown under citrate accumulating conditions, acidogenesis was far less sensitive to inhibition by Mn/sup 2 +/ than in the parent strain. Some of the cic mutants also showed altered citrate inhibition of NADP-specific isocitrate dehydrogenase.

  10. Induction of intestinal pro-inflammatory immune responses by lipoteichoic acid

    Directory of Open Access Journals (Sweden)

    Zadeh Mojgan

    2012-03-01

    Full Text Available Abstract Background The cellular and molecular mechanisms of inflammatory bowel disease are not fully understood; however, data indicate that uncontrolled chronic inflammation induced by bacterial gene products, including lipoteichoic acid (LTA, may trigger colonic inflammation resulting in disease pathogenesis. LTA is a constituent glycolipid of Gram-positive bacteria that shares many inflammatory properties with lipopolysaccharide and plays a critical role in the pathogenesis of severe inflammatory responses via Toll-like receptor 2. Accordingly, we elucidate the role of LTA in immune stimulation and induced colitis in vivo. Methods To better understand the molecular mechanisms utilized by the intestinal microbiota and their gene products to induce or subvert inflammation, specifically the effect(s of altered surface layer protein expression on the LTA-mediated pro-inflammatory response, the Lactobacillus acidophilus surface layer protein (Slp genes encoding SlpB and SlpX were deleted resulting in a SlpB- and SlpX- mutant that continued to express SlpA (assigned as NCK2031. Results Our data show profound activation of dendritic cells by NCK2031, wild-type L. acidophilus (NCK56, and purified Staphylococcus aureus-LTA. In contrary to the LTA-deficient strain NCK2025, the LTA-expressing strains NCK2031 and NCK56, as well as S. aureus-LTA, induce pro-inflammatory innate and T cell immune responses in vivo. Additionally, neither NCK2031 nor S. aureus-LTA supplemented in drinking water protected mice from DSS-colitis, but instead, induced significant intestinal inflammation resulting in severe colitis and tissue destruction. Conclusions These findings suggest that directed alteration of two of the L. acidophilus NCFM-Slps did not ameliorate LTA-induced pro-inflammatory signals and subsequent colitis.

  11. Ethylene signalling is involved in regulation of phosphate starvation-induced gene expression and production of acid phosphatases and anthocyanin in Arabidopsis

    KAUST Repository

    Lei, Mingguang

    2010-11-30

    With the exception of root hair development, the role of the phytohormone ethylene is not clear in other aspects of plant responses to inorganic phosphate (Pi) starvation. The induction of AtPT2 was used as a marker to find novel signalling components involved in plant responses to Pi starvation. Using genetic and chemical approaches, we examined the role of ethylene in the regulation of plant responses to Pi starvation. hps2, an Arabidopsis mutant with enhanced sensitivity to Pi starvation, was identified and found to be a new allele of CTR1 that is a key negative regulator of ethylene responses. 1-aminocyclopropane-1-carboxylic acid (ACC), the precursor of ethylene, increases plant sensitivity to Pi starvation, whereas the ethylene perception inhibitor Ag+ suppresses this response. The Pi starvation-induced gene expression and acid phosphatase activity are also enhanced in the hps2 mutant, but suppressed in the ethylene-insensitive mutant ein2-5. By contrast, we found that ethylene signalling plays a negative role in Pi starvation-induced anthocyanin production. These findings extend the roles of ethylene in the regulation of plant responses to Pi starvation and will help us to gain a better understanding of the molecular mechanism underlying these responses. © 2010 The Authors. New Phytologist © 2010 New Phytologist Trust.

  12. Abscisic acid and ethylene in mutants of Arabidopsis thaliana differing in their resistance to ultraviolet (UV-B) radiation stress

    International Nuclear Information System (INIS)

    Rakitina, T.Ya.; Vlasov, P.V.; Jalilova, F.Kh.; Kefeli, V.I.

    1994-01-01

    The effects of ultraviolet irradiation (between 280 and 320 nm) on plant survival, ethylene evolution, and abscisic acid (ABA) content were studied in Arabidopsis thaliana (L.) Heunh. plants. Three genetic lines of Arabidopsis differing in their resistance to ultraviolet (UV-B) radiation stress were used. UV-B irradiation had detrimental effects on plant survival, enhanced ethylene evolution, and increased ABA content in the plants of all three lines. The higher ultraviolet dose was absorbed, the less was the number of surviving plants and the higher were the levels of both phytohormones. The maximum ethylene evolution occurred during the initial two to four hours after irradiation, but the ABA content peaked only after 24 h. The most resistant line showed the highest ABA content and the fastest ethylene evolution, whereas, in the susceptible line, both indices were the lowest. After UV-B treatment, the ABA-deficient Arabidopsis mutant evolved four to six times more ethylene than the plants with normal ABA content. Stress ethylene production evidently did not depend on the level of endogenous ABA as the kinetics of ethylene evolution was similar in the ABA-deficient mutant and in other studied Arabidopsis lines

  13. The interaction of strigolactones with abscisic acid during the drought response in rice

    KAUST Repository

    Haider, Imran; Andreo-Jimenez, Beatriz; Bruno, Mark; Bimbo, Andrea; Floková , Kristý na; Abuauf, Haneen Waleed Hamza; Otang Ntui, Valentine; Guo, Xiujie; Charnikhova, Tatsiana; Al-Babili, Salim; Bouwmeester, Harro J; Ruyter-Spira, Carolien

    2018-01-01

    Both strigolactones (SLs) and abscisic acid (ABA) biosynthetically originate from carotenoids. Considering their common origin, the interaction of these two hormones at the biosynthetic and/or regulatory level may be anticipated. Here, we show in rice that drought simultaneously induces SL production in the root, and ABA production and the expression of SL biosynthetic genes in the shoot. Under control conditions, the ABA concentration was higher in shoots of the SL biosynthetic rice mutants dwarf10 (d10) and d17 than in wild-type plants, while a similar trend was observed for SL-perception mutant d3. These differences were enhanced under drought. However, drought did not result in an increase in leaf ABA content in rice mutant line d27, carrying a mutation in the gene encoding the first committed enzyme in SL biosynthesis, to the same extent as in the other SL mutants and the wild-type. Accordingly, d10, d17 and d3 lines were more drought tolerant than wild-type plants, whereas d27 displayed decreased tolerance. Finally, over-expression of OsD27 in rice resulted in increased levels of ABA when compared with wild-type plants. We conclude that the SL and ABA pathways are connected with each other through D27, which is playing a crucial role in determining ABA and SL content in rice.

  14. The interaction of strigolactones with abscisic acid during the drought response in rice

    KAUST Repository

    Haider, Imran

    2018-03-09

    Both strigolactones (SLs) and abscisic acid (ABA) biosynthetically originate from carotenoids. Considering their common origin, the interaction of these two hormones at the biosynthetic and/or regulatory level may be anticipated. Here, we show in rice that drought simultaneously induces SL production in the root, and ABA production and the expression of SL biosynthetic genes in the shoot. Under control conditions, the ABA concentration was higher in shoots of the SL biosynthetic rice mutants dwarf10 (d10) and d17 than in wild-type plants, while a similar trend was observed for SL-perception mutant d3. These differences were enhanced under drought. However, drought did not result in an increase in leaf ABA content in rice mutant line d27, carrying a mutation in the gene encoding the first committed enzyme in SL biosynthesis, to the same extent as in the other SL mutants and the wild-type. Accordingly, d10, d17 and d3 lines were more drought tolerant than wild-type plants, whereas d27 displayed decreased tolerance. Finally, over-expression of OsD27 in rice resulted in increased levels of ABA when compared with wild-type plants. We conclude that the SL and ABA pathways are connected with each other through D27, which is playing a crucial role in determining ABA and SL content in rice.

  15. The MEDIATOR genes MED12 and MED13 control Arabidopsis root system configuration influencing sugar and auxin responses.

    Science.gov (United States)

    Raya-González, Javier; López-Bucio, Jesús Salvador; Prado-Rodríguez, José Carlos; Ruiz-Herrera, León Francisco; Guevara-García, Ángel Arturo; López-Bucio, José

    2017-09-01

    Arabidopsis med12 and med13 mutants exhibit shoot and root phenotypes related to an altered auxin homeostasis. Sucrose supplementation reactivates both cell division and elongation in primary roots as well as auxin-responsive and stem cell niche gene expression in these mutants. An analysis of primary root growth of WT, med12, aux1-7 and med12 aux1 single and double mutants in response to sucrose and/or N-1-naphthylphthalamic acid (NPA) placed MED12 upstream of auxin transport for the sugar modulation of root growth. The MEDIATOR (MED) complex plays diverse functions in plant development, hormone signaling and biotic and abiotic stress tolerance through coordination of transcription. Here, we performed genetic, developmental, molecular and pharmacological analyses to characterize the role of MED12 and MED13 on the configuration of root architecture and its relationship with auxin and sugar responses. Arabidopsis med12 and med13 single mutants exhibit shoot and root phenotypes consistent with altered auxin homeostasis including altered primary root growth, lateral root development, and root hair elongation. MED12 and MED13 were required for activation of cell division and elongation in primary roots, as well as auxin-responsive and stem cell niche gene expression. Remarkably, most of these mutant phenotypes were rescued by supplying sucrose to the growth medium. The growth response of primary roots of WT, med12, aux1-7 and med12 aux1 single and double mutants to sucrose and application of auxin transport inhibitor N-1-naphthylphthalamic acid (NPA) revealed the correlation of med12 phenotype with the activity of the auxin intake permease and suggests that MED12 acts upstream of AUX1 in the root growth response to sugar. These data provide compelling evidence that MEDIATOR links sugar sensing to auxin transport and distribution during root morphogenesis.

  16. Mutations Enhancing Amino Acid Catabolism Confer a Growth Advantage in Stationary Phase

    Science.gov (United States)

    Zinser, Erik R.; Kolter, Roberto

    1999-01-01

    Starved cultures of Escherichia coli undergo successive rounds of population takeovers by mutants of increasing fitness. These mutants express the growth advantage in stationary phase (GASP) phenotype. Previous work identified the rpoS819 allele as a GASP mutation allowing cells to take over stationary-phase cultures after growth in rich media (M. M. Zambrano, D. A. Siegele, M. A. Almirón, A. Tormo, and R. Kolter, Science 259:1757–1760, 1993). Here we have identified three new GASP loci from an aged rpoS819 strain: sgaA, sgaB, and sgaC. Each locus is capable of conferring GASP on the rpoS819 parent, and they can provide successively higher fitnesses for the bacteria in the starved cultures. All four GASP mutations isolated thus far allow for faster growth on both individual and mixtures of amino acids. Each mutation confers a growth advantage on a different subset of amino acids, and these mutations act in concert to increase the overall catabolic capacity of the cell. We present a model whereby this enhanced ability to catabolize amino acids is responsible for the fitness gain during carbon starvation, as it may allow GASP mutants to outcompete the parental cells when growing on the amino acids released by dying cells. PMID:10482523

  17. Functional improvement of Saccharomyces cerevisiae to reduce volatile acidity in wine.

    Science.gov (United States)

    Luo, Zongli; Walkey, Christopher J; Madilao, Lufiani L; Measday, Vivien; Van Vuuren, Hennie J J

    2013-08-01

    Control of volatile acidity (VA) is a major issue for wine quality. In this study, we investigated the production of VA by a deletion mutant of the fermentation stress response gene AAF1 in the budding yeast Saccharomyces cerevisiae. Fermentations were carried out in commercial Chardonnay grape must to mimic industrial wine-making conditions. We demonstrated that a wine yeast strain deleted for AAF1 reduced acetic acid levels in wine by up to 39.2% without increasing the acetaldehyde levels, revealing a potential for industrial application. Deletion of the cytosolic aldehyde dehydrogenase gene ALD6 also reduced acetic acid levels dramatically, but increased the acetaldehyde levels by 41.4%, which is not desired by the wine industry. By comparison, ALD4 and the AAF1 paralog RSF2 had no effects on acetic acid production in wine. Deletion of AAF1 was detrimental to the growth of ald6Δ and ald4Δald6Δ mutants, but had no effect on acetic acid production. Overexpression of AAF1 dramatically increased acetic acid levels in wine in an Ald6p-dependent manner, indicating that Aaf1p regulates acetic acid production mainly via Ald6p. Overexpression of AAF1 in an ald4Δald6Δ strain produced significantly more acetic acid in wine than the ald4Δald6Δ mutant, suggesting that Aaf1p may also regulate acetic acid synthesis independently of Ald4p and Ald6p. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

  18. Variation of several agronomic and biochemical traits in γ-ray induced mutant soybeans

    International Nuclear Information System (INIS)

    Shim, Kyo Moon; Kim, Sun Hyung; Kim, Nam Soo; Son, Hi Sup; Rhee, Hae Ik

    1996-01-01

    Two soybean cultivars(Paldalkong and Bangsakong) were irradiated with gamma-ray to reduce seed size, which is an important trait for soybean sprout and the derived mutant soybeans were analyzed in several agronomic and biochemical traits. There were high levels of variations in quantitative traits among the mutants. Several mutant lines showed higher yield and smaller seed than their parents. Qualitative traits such as seed coat color or pubescent color were also changed in a few lines. Biochemical variations were also observed among the mutants. In seed storage protein analysis, many mutant lines showed reduced intensities in β-subunits in 7S globulin than their parents and an additional band in the acidic subunit at 31KDa. Two mutant lines derived from Paldalkong showed an additional band and a shifted band of protease inhibitor by electrophoretic analysis. Variations in isozymes and RAPD were also observed among the mutants. Six isozymes(Adh, Est, Gdh, Idh, Mdh and Pgm) among eleven isozymes showed some variations and six out of ten primers showed polymorphic amplified DNA fragments among the mutants. (author)

  19. Optimization of Medium Components for Cell Biomass and Polyhydroxy Butyric Acid Production by Azotobacter vinelandii Mutant Using Response Surface Methodology

    International Nuclear Information System (INIS)

    Safiyyah Zainuddin; Nur Izzah Mohd Razak; Ying, P.L.W.; Chyan, J.B.; Elly Ellyna Rashid

    2016-01-01

    Polyhydroxy butyric acid (PHB) is a biodegradable and food-safe alternative to petroleum-based polymers. Using RSM approach, the interaction of sucrose, urea and K_2HPO_4 were investigated to determine the optimum medium compositions for cell biomass and PHB production by Azotobacter vinelandii mutant. Fifteen medium types were prepared and each contained different amount of sucrose, urea and K_2HPO_4. Analyses of cell biomass and PHB concentration were performed from day-2 until day-4 (3 days). Based on the biomass analysis, Medium 13 achieved the highest cell dry weight of 15.4 mg/ mL on day-3. Medium 13 contained 0.5 g/ L of urea, 0.1 g/ L of K_2HPO_4 and 10 g/ L of sucrose. For PHB production, Medium 11 achieved the highest PHB production on day-3 (3.7 mg/ mL) and dropped to 1.3 mg/ mL on day-4. Sample 11 contained 0.5 g/ L of urea, 0 g/ L of K_2HPO_4 and 20 g/ L of sucrose. Sample 2 (1.0 g/ L urea, 0.05 g/ L K_2HPO_4 and 15 g/ L sucrose) and 6 (1.0 g/ L urea, 0.05 g/ L K_2HPO_4 and 25 g/ L sucrose) showed PHB production of >2.0 mg/ mL on day-3 and persisted to day-4. Sample 3 (0.25 g/ L urea, 0.2 g/ L K_2HPO_4 and 15 g/L sucrose) achieved PHB production of >2.0 mg/mL only on day-4. All the other medium types showed PHB production of lower than 1.5 mg/ mL throughout the experiment. (author)

  20. Breeding of a sake yeast mutant with enhanced ethyl caproate productivity in sake brewing using rice milled at a high polishing ratio.

    Science.gov (United States)

    Takahashi, Toshinari; Ohara, Yusuke; Sueno, Kazuo

    2017-06-01

    Sake yeast produces a fruity flavor known as ginjo-ko-which is mainly attributable to ethyl caproate and isoamyl acetate-during fermentation in sake brewing. The production of these flavor components is inhibited by unsaturated fatty acids derived from the outer layer of rice as raw material. We isolated three mutants (hec2, hec3, and hec6) with enhanced ethyl caproate productivity in sake brewing using rice milled at a high polishing ratio from a cerulenin-resistant mutant derived from the hia1 strain, which shows enhanced isoamyl acetate productivity. The hec2 mutant had the homozygous FAS2 mutation Gly1250Ser, which is known to confer high ethyl caproate productivity. When the homozygous FAS2 mutation Gly1250Ser was introduced into strain hia1, ethyl caproate productivity was increased but neither this nor intracellular caproic acid content approached the levels observed in the hec2 mutant, indicating that a novel mutation was responsible for the high ethyl caproate productivity. We also found that the expression of EEB1 encoding acyl-coenzyme A:ethanol O-acyltransferase (AEATase) and enzymatic activity were increased in the hec2 mutant. These results suggest that the upregulation of EEB1 expression and AEATase activity may also have contributed to the enhancement of ethyl caproate synthesis from ethanol and caproyl-CoA. Our findings are useful for the brewing of sake with improved flavor due to high levels of isoamyl acetate and ethyl caproate. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  1. Acidophilus Milk Shelf-life Prolongation by the Use of Cold Sensitive Mutants of Lactobacillus acidophilus MDC 9626

    Directory of Open Access Journals (Sweden)

    Alireza Goodarzi

    2017-09-01

    Full Text Available  Background and Objective: The shelf-life of Acidophilus milk fermented by probiotic culture Lactobacillus acidophilus is limited due to acidification caused by continued organic acid formation at low temperatures. Increasing of titrable acidity in turn causes reducing of the total viable count of probiotic bacteria. To overcome acidification we suggested to use coldsensitive mutants of Lactobacillus acidophilus, with limited metabolism at low temperatures. In order to facilitate the selection of cold sensitive mutants, it was decided to use Rifampicin and Streotomycin mutations affecting thermostability of the key molecules of cell metabolism the RNA polymerase and ribosome, respectively.Material and Methods: Ultra violet mutagenesis was used to enhance the yield and diversity of rifampicin and streptomycin resistant mutants of Lactobacillus acidophilus. To perform negative selection of cold sensitive mutants, antibiotic resistant colonies replica plated and incubated at 23ºC. The growth rate, milk fermenting rate, titratable acidity were measured.Results and Conclusion: Among tested resistant to either rifampicin or streptomycin clones with frequency mean of 1.0 %, ten mutants were isolated which have lost the ability to grow at minimal temperature. Fermented with cold-sensitive mutants of Lactobacillus acidophilus milks, during storage in the refrigerator, almost twice as long retained high amount of probiotic bacteria and low titratable acidity as compared to the parent strain. Thus, direct relationship between temperature sensitivity of the starter and shelf life of acidophilic milk was confirmed. Rifampicin and Streptomycin resistant mutations are powerful tools for selection of cold-sensitive dairy starters for preparing dairy fermented products with long shelf-life.Conflict of interest: The authors declare no conflict of interest.

  2. Biochemical responses and ultrastructural changes in ethylene insensitive mutants of Arabidopsis thialiana subjected to bisphenol A exposure.

    Science.gov (United States)

    Ali, Imran; Jan, Mehmood; Wakeel, Abdul; Azizullah, Azizullah; Liu, Bohan; Islam, Faisal; Ali, Abid; Daud, M K; Liu, Yihua; Gan, Yinbo

    2017-10-01

    Bisphenol A (BPA), an important raw material in plastic industry, has become a serious environmental contaminant due to its wide spread use in different products and increasing release into the environment. BPA is known to cause adverse effects in living organisms including plants. Several studies reported that BPA affects growth and development in plants, mainly through oxidative stress. Plants are known to generally cope with stress mainly through hormonal regulation and adaptation, but little is known about the role of plant hormones in plants under BPA stress. The present study was conducted to investigate the role of ethylene in BPA induced oxidative stress in plants using Arabidopsis thaliana as a test plant. The response of ethylene insensitive mutants of Arabidopsis (ein2-1 and etr1-3) to BPA exposure was studied in comparison to the wild type Arabidopsis (WT). In all three genotypes, exposure to BPA adversely affected cellular structures, stomata and light-harvesting pigments. An increase in reactive oxygen species (ROS) lipid peroxidation and other oxidative stress markers indicated that BPA induced toxicity through oxidative stress. However, the overall results revealed that WT Arabidopsis had more pronounced BPA induced damages while ein2-1 and etr1-3 mutants withstood the BPA induced stress more efficiently. The activity of antioxidant enzymes and expression of antioxidants related genes revealed that the antioxidant defense system in both mutants was more efficiently activated than in WT against BPA induced oxidative stress, which further evidenced the involvement of ethylene in regulating BPA induced oxidative stress. It is concluded that ethylene perception and signaling may be involved in BPA induced oxidative stress responses in plants. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Genetics of Ustilago violacea. I. Carotenoid mutants and carotenogenesis

    International Nuclear Information System (INIS)

    Garber, E.D.; Baird, M.L.; Chapman, D.J.

    1975-01-01

    Wild-type strains of Ustilago violacea produce pink colonies on laboratory medium and yield white, orange, pumpkin, and yellow colonies after uv mutagenesis. The wild-type strains contain neurosporene and lycopene; one orange mutant, γ-carotene; and one yellow mutant, β-carotene. One white mutant had no detectable carotenoids. Diploid colonies heterozygous for wild type and orange, pumpkin, yellow, or white are phenotypically wild type. Diploid colonies heterozygous for yellow and orange are also phenotypically wild type. Diploid colonies heterozygous for white and orange; white and yellow; and white, yellow, and orange are phenotypically light orange, light yellow, and orange-yellow, respectively. The white mutants give a circular complementation map; the color mutants fit a linear complementation map. We propose a multienzyme of four identical dehydrogenases and one or two identical cyclases for carotenogenesis in this species. The white and color mutants represent structural mutations altering the conformation of the dehydrogenase or cyclase, respectively. Furthermore, cyclases may or may not aggregate in association with the dehydrogenase aggregate to form the multienzyme aggregate responsible for the color mutants

  4. Identification of genes required for secretion of the Francisella oxidative burst-inhibiting acid phosphatase AcpA

    Directory of Open Access Journals (Sweden)

    John S Gunn

    2016-04-01

    Full Text Available Francisella tularensis is a Tier 1 bioterror threat and the intracellular pathogen responsible for tularemia in humans and animals. Upon entry into the host, Francisella uses multiple mechanisms to evade killing. Our previous studies have shown that after entering its primary cellular host, the macrophage, Francisella immediately suppresses the oxidative burst by secreting a series of acid phosphatases including AcpA-B-C and HapA, thereby evading the innate immune response of the macrophage and enhancing survival and further infection. However, the mechanism of acid phosphatase secretion by Francisella is still unknown. In this study, we screened for genes required for AcpA secretion in Francisella. We initially demonstrated that the known secretion systems, the putative Francisella-pathogenicity island (FPI-encoded Type VI secretion system and the Type IV pili, do not secrete AcpA. Using random transposon mutagenesis in conjunction with ELISA, Western blotting and acid phosphatase enzymatic assays, a transposon library of 5450 mutants was screened for strains with a minimum 1.5-fold decrease in secreted (culture supernatant AcpA, but no defect in cytosolic AcpA. Three mutants with decreased supernatant AcpA were identified. The transposon insertion sites of these mutants were revealed by direct genomic sequencing or inverse-PCR and sequencing. One of these mutants has a severe defect in AcpA secretion (at least 85% decrease and is a predicted hypothetical inner membrane protein. Interestingly, this mutant also affected the secretion of the FPI-encoded protein, VgrG. Thus, this screen identified novel protein secretion factors involved in the subversion of host defenses.

  5. Silverleaf Whitefly Induces Salicylic Acid Defenses and Suppresses Effectual Jasmonic Acid Defenses1[W][OA

    Science.gov (United States)

    Zarate, Sonia I.; Kempema, Louisa A.; Walling, Linda L.

    2007-01-01

    The basal defenses important in curtailing the development of the phloem-feeding silverleaf whitefly (Bemisia tabaci type B; SLWF) on Arabidopsis (Arabidopsis thaliana) were investigated. Sentinel defense gene RNAs were monitored in SLWF-infested and control plants. Salicylic acid (SA)-responsive gene transcripts accumulated locally (PR1, BGL2, PR5, SID2, EDS5, PAD4) and systemically (PR1, BGL2, PR5) during SLWF nymph feeding. In contrast, jasmonic acid (JA)- and ethylene-dependent RNAs (PDF1.2, VSP1, HEL, THI2.1, FAD3, ERS1, ERF1) were repressed or not modulated in SLWF-infested leaves. To test for a role of SA and JA pathways in basal defense, SLWF development on mutant and transgenic lines that constitutively activate or impair defense pathways was determined. By monitoring the percentage of SLWF nymphs in each instar, we show that mutants that activate SA defenses (cim10) or impair JA defenses (coi1) accelerated SLWF nymphal development. Reciprocally, mutants that activate JA defenses (cev1) or impair SA defenses (npr1, NahG) slowed SLWF nymphal development. Furthermore, when npr1 plants, which do not activate downstream SA defenses, were treated with methyl jasmonate, a dramatic delay in nymph development was observed. Collectively, these results showed that SLWF-repressed, JA-regulated defenses were associated with basal defense to the SLWF. PMID:17189328

  6. Distinct Fiber Type Signature in Mouse Muscles Expressing a Mutant Lamin A Responsible for Congenital Muscular Dystrophy in a Patient

    Directory of Open Access Journals (Sweden)

    Alice Barateau

    2017-04-01

    Full Text Available Specific mutations in LMNA, which encodes nuclear intermediate filament proteins lamins A/C, affect skeletal muscle tissues. Early-onset LMNA myopathies reveal different alterations of muscle fibers, including fiber type disproportion or prominent dystrophic and/or inflammatory changes. Recently, we identified the p.R388P LMNA mutation as responsible for congenital muscular dystrophy (L-CMD and lipodystrophy. Here, we asked whether viral-mediated expression of mutant lamin A in murine skeletal muscles would be a pertinent model to reveal specific muscle alterations. We found that the total amount and size of muscle fibers as well as the extent of either inflammation or muscle regeneration were similar to wildtype or mutant lamin A. In contrast, the amount of fast oxidative muscle fibers containing myosin heavy chain IIA was lower upon expression of mutant lamin A, in correlation with lower expression of genes encoding transcription factors MEF2C and MyoD. These data validate this in vivo model for highlighting distinct muscle phenotypes associated with different lamin contexts. Additionally, the data suggest that alteration of muscle fiber type identity may contribute to the mechanisms underlying physiopathology of L-CMD related to R388P mutant lamin A.

  7. Isolation and characterization of acyclovir-resistant mutants of herpes simplex virus.

    Science.gov (United States)

    Field, H J; Darby, G; Wildy, P

    1980-07-01

    Mutants of HSV which are resistant to acyclovir (acycloguanosine) have been isolated following serial passages of several herpes simplex virus (HSV) strains in the presence of the drug. The majority of the mutants isolated are defective in induction of thymidine kinase (TK) and this is consistent with the observation that independently isolated TK- viruses are naturally resistant to ACV. One mutant is described (SC16 R9C2) which is resistant in biochemically transformed cells which express HSV TK. This suggests that its resistance resides at a level other than TK. It is also resistant to phosphonoacetic acid, suggesting that the DNA polymerase locus may be involved. A further mutant is described [Cl (101) P2C5] which induces normal levels of TK, although the nature of resistance of this virus is not yet elucidated.

  8. Disparate peroxisome-related defects in Arabidopsis pex6 and pex26 mutants link peroxisomal retrotranslocation and oil body utilization.

    Science.gov (United States)

    Gonzalez, Kim L; Fleming, Wendell A; Kao, Yun-Ting; Wright, Zachary J; Venkova, Savina V; Ventura, Meredith J; Bartel, Bonnie

    2017-10-01

    Catabolism of fatty acids stored in oil bodies is essential for seed germination and seedling development in Arabidopsis. This fatty acid breakdown occurs in peroxisomes, organelles that sequester oxidative reactions. Import of peroxisomal enzymes is facilitated by peroxins including PEX5, a receptor that delivers cargo proteins from the cytosol to the peroxisomal matrix. After cargo delivery, a complex of the PEX1 and PEX6 ATPases and the PEX26 tail-anchored membrane protein removes ubiquitinated PEX5 from the peroxisomal membrane. We identified Arabidopsis pex6 and pex26 mutants by screening for inefficient seedling β-oxidation phenotypes. The mutants displayed distinct defects in growth, response to a peroxisomally metabolized auxin precursor, and peroxisomal protein import. The low PEX5 levels in these mutants were increased by treatment with a proteasome inhibitor or by combining pex26 with peroxisome-associated ubiquitination machinery mutants, suggesting that ubiquitinated PEX5 is degraded by the proteasome when the function of PEX6 or PEX26 is reduced. Combining pex26 with mutations that increase PEX5 levels either worsened or improved pex26 physiological and molecular defects, depending on the introduced lesion. Moreover, elevating PEX5 levels via a 35S:PEX5 transgene exacerbated pex26 defects and ameliorated the defects of only a subset of pex6 alleles, implying that decreased PEX5 is not the sole molecular deficiency in these mutants. We found peroxisomes clustered around persisting oil bodies in pex6 and pex26 seedlings, suggesting a role for peroxisomal retrotranslocation machinery in oil body utilization. The disparate phenotypes of these pex alleles may reflect unanticipated functions of the peroxisomal ATPase complex. © 2017 The Authors The Plant Journal © 2017 John Wiley & Sons Ltd.

  9. Enterocin A mutants identified by saturation mutagenesis enhance potency towards vancomycin-resistant Enterococci.

    Science.gov (United States)

    McClintock, Maria K; Kaznessis, Yiannis N; Hackel, Benjamin J

    2016-02-01

    Vancomycin-resistant Enterococci infections are a significant clinical problem. One proposed solution is to use probiotics, such as lactic acid bacteria, to produce antimicrobial peptides at the site of infection. Enterocin A, a class 2a bacteriocin, exhibits inhibitory activity against E. faecium and E. faecalis, which account for 86% of vancomycin-resistant Enterococci infections. In this study, we aimed to engineer enterocin A mutants with enhanced potency within a lactic acid bacterial production system. Peptide mutants resulting from saturation mutagenesis at sites A24 and T27 were efficiently screened in a 96-well plate assay for inhibition of pathogen growth. Several mutants exhibit increased potency relative to wild-type enterocin A in both liquid- and solid-medium growth assays. In particular, A24P and T27G exhibit enhanced inhibition of multiple strains of E. faecium and E. faecalis, including clinically isolated vancomycin-resistant strains. A24P and T27G enhance killing of E. faecium 8 by 13 ± 3- and 18 ± 4-fold, respectively. The engineered enterocin A/lactic acid bacteria systems offer significant potential to combat antibiotic-resistant infections. © 2015 Wiley Periodicals, Inc.

  10. Characterization of Helicobacter pylori dapE and construction of a conditionally lethal dapE mutant.

    Science.gov (United States)

    Karita, M; Etterbeek, M L; Forsyth, M H; Tummuru, M K; Blaser, M J

    1997-10-01

    Helicobacter pylori colonizes the human gastric mucosa and causes gastritis, ulceration, or gastric cancer. A previously uncharacterized region of the H. pylori genome was identified and sequenced. This region includes a putative operon containing three open reading frames termed gidA (1,866 bp), dapE (1,167 bp), and orf2 (753 bp); the gidA and dapE products are highly homologous to other bacterial proteins. In E. coli, dapE encodes N-succinyl-L-diaminopimelic acid desuccinylase, which catalyzes the hydrolysis of N-succinyl-L-diaminopimelic acid to L-diaminopimelic acid (L-DAP) and succinate. When wild-type H. pylori strains were transformed to select for dapE mutagenesis, mutants were present when plates were supplemented with DAP but not with lysine; orf2 mutants were selected without DAP supplementation. Consistent with the finding that GidA is essential in Escherichia coli, we were unable to obtain a gidA mutant in H. pylori despite evidence that insertional mutagenesis had occurred. The positions of gidA, dapE, and orf2 suggest that they form an operon, which was supported by slot blot RNA hybridization and reverse transcriptase PCR studies. The data imply that the H. pylori dapE mutant may be useful as a conditionally lethal vaccine.

  11. The omega-3 fatty acid eicosapentaenoic acid is required for normal alcohol response behaviors in C. elegans.

    Directory of Open Access Journals (Sweden)

    Richard C Raabe

    Full Text Available Alcohol addiction is a widespread societal problem, for which there are few treatments. There are significant genetic and environmental influences on abuse liability, and understanding these factors will be important for the identification of susceptible individuals and the development of effective pharmacotherapies. In humans, the level of response to alcohol is strongly predictive of subsequent alcohol abuse. Level of response is a combination of counteracting responses to alcohol, the level of sensitivity to the drug and the degree to which tolerance develops during the drug exposure, called acute functional tolerance. We use the simple and well-characterized nervous system of Caenorhabditis elegans to model the acute behavioral effects of ethanol to identify genetic and environmental factors that influence level of response to ethanol. Given the strong molecular conservation between the neurobiological machinery of worms and humans, cellular-level effects of ethanol are likely to be conserved. Increasingly, variation in long-chain polyunsaturated fatty acid levels has been implicated in complex neurobiological phenotypes in humans, and we recently found that fatty acid levels modify ethanol responses in worms. Here, we report that 1 eicosapentaenoic acid, an omega-3 polyunsaturated fatty acid, is required for the development of acute functional tolerance, 2 dietary supplementation of eicosapentaenoic acid is sufficient for acute tolerance, and 3 dietary eicosapentaenoic acid can alter the wild-type response to ethanol. These results suggest that genetic variation influencing long-chain polyunsaturated fatty acid levels may be important abuse liability loci, and that dietary polyunsaturated fatty acids may be an important environmental modulator of the behavioral response to ethanol.

  12. Isolation and characterization of a Ds-tagged rice (Oryza sativa L.) GA-responsive dwarf mutant defective in an early step of the gibberellin biosynthesis pathway.

    Science.gov (United States)

    Margis-Pinheiro, Marcia; Zhou, Xue-Rong; Zhu, Qian-Hao; Dennis, Elizabeth S; Upadhyaya, Narayana M

    2005-03-01

    We have isolated a severe dwarf transposon (Ds) insertion mutant in rice (Oryza sativa L.), which could be differentiated early in the seedling stage by reduced shoot growth and dark green leaves, and later by severe dwarfism and failure to initiate flowering. These mutants, however, showed normal seed germination and root growth. One of the sequences flanking Ds, rescued from the mutant, was of a chromosome 4-located putative ent-kaurene synthase (KS) gene, encoding the enzyme catalyzing the second step of the gibberellin (GA) biosynthesis pathway. Dwarf mutants were always homozygous for this Ds insertion and no normal plants homozygous for this mutation were recovered in the segregating progeny, indicating that the Ds insertion mutation is recessive. As mutations in three recently reported rice GA-responsive dwarf mutant alleles and the dwarf mutation identified in this study mapped to the same locus, we designate the corresponding gene OsKS1. The osks1 mutant seedlings were responsive to exogenous gibberellin (GA3). OsKS1 transcripts of about 2.3 kb were detected in leaves and stem of wild-type plants, but not in germinating seeds or roots, suggesting that OsKS1 is not involved in germination or root growth. There are at least five OsKS1-like genes in the rice genome, four of which are also represented in rice expressed sequence tag (EST) databases. All OsKS1-like genes are transcribed with different expression patterns. ESTs corresponding to all six OsKS genes are represented in other cereal databases including barley, wheat and maize, suggesting that they are biologically active.

  13. Symbiotic N fixation of several soybean varieties and mutants

    International Nuclear Information System (INIS)

    Soertini, G.; Hendratno

    1988-01-01

    Symbiotic N fixation of several soybean varieties and mutants. Research activities comprising of three experiments were carried out to screen several soybean varieties and mutants for symbiotic N fixation potential. The first two experiments involved screening of seven rhizobium strains/isolate for effective N fixation. Depending on the medium used, plant response to strains was different. In sterile medium, rhizobium strain USDA 136, 142 and TAL 102 showed a high nitrogen fixation potential. In soil only rhizobium strain USDA 110 had better performance and proved to be competitive to the native strains. Nitrogen-15 dilution method was used to screen nitrogen fixing ability of several soybean varieties and mutants. Guntur variety showed a better response to high dose of N fertilizer without disturbance in its fixing ability. This variety then was considered good to be introduced in the cropping system. (author). 8 refs

  14. Mutants of Streptomyces coeruleorubidus impaired in the biosynthesis of daunomycinone glycosides and related metabolites

    International Nuclear Information System (INIS)

    Blumauerova, M.; Stajner, K.; Pokorny, V.; Hostalek, Z.; Vanek, Z.

    1978-01-01

    Mutants of Streptomyces coeruleorubidus, blocked in the biosynthesis of anthracycline antibiotics of the daunomycine complex, were isolated from the production strains after treatment with UV light, γ-radiation, nitrous acid, and after natural selection; according to their different biosynthetic activity the mutants were divided into five phenotypic groups. Mutants of two of these groups produced compounds that had not yet been described in Streptomyces coeruleorubidus (aklavinone, 7-deoxyaklavinone, zeta-rhodomycinone and glycosides of epsilon-rhodomycinone). The mutants differed from the parent strains and also mutually in morphological characteristics but no direct correlation between these changes and the biosynthetic activity could be observed in most cases. (author)

  15. Genome-wide Escherichia coli stress response and improved tolerance towards industrially relevant chemicals

    DEFF Research Database (Denmark)

    Rau, Martin Holm; Calero Valdayo, Patricia; Lennen, Rebecca

    2016-01-01

    Economically viable biobased production of bulk chemicals and biofuels typically requires high product titers. During microbial bioconversion this often leads to product toxicity, and tolerance is therefore a critical element in the engineering of production strains. Here, a systems biology...... approach was employed to understand the chemical stress response of Escherichia coli, including a genome-wide screen for mutants with increased fitness during chemical stress. Twelve chemicals with significant production potential were selected, consisting of organic solvent-like chemicals (butanol......, hydroxy-γ-butyrolactone, 1,4-butanediol, furfural), organic acids (acetate, itaconic acid, levulinic acid, succinic acid), amino acids (serine, threonine) and membrane-intercalating chemicals (decanoic acid, geraniol). The transcriptional response towards these chemicals revealed large overlaps...

  16. Some biochemistry characteristics of soja (Glycine Max.L.) Tshibashi and its two mutants

    International Nuclear Information System (INIS)

    Botula, Manyala; Kasende, Mundeke; Kembola, Kejuni.

    1978-01-01

    Two mutants of a variety of soybeans (Thibashi) obtained by neutron irradiation in a reactor have been compared to their parents for their proteins and lipids contents. Analysis have been made by a modified micro-kjedhal method for proteins and by gaz chromatography for lipids converted into methylic esters. A slight lysine and hyptidine increase and a slight linolenic acid decrease are observed in the two mutants

  17. Covalent modification of mutant rat P2X2 receptors with a thiol-reactive fluorophore allows channel activation by zinc or acidic pH without ATP.

    Directory of Open Access Journals (Sweden)

    Shlomo S Dellal

    Full Text Available Rat P2X2 receptors open at an undetectably low rate in the absence of ATP. Furthermore, two allosteric modulators, zinc and acidic pH, cannot by themselves open these channels. We describe here the properties of a mutant receptor, K69C, before and after treatment with the thiol-reactive fluorophore Alexa Fluor 546 C(5-maleimide (AM546. Xenopus oocytes expressing unmodified K69C were not activated under basal conditions nor by 1,000 µM ATP. AM546 treatment caused a small increase in the inward holding current which persisted on washout and control experiments demonstrated this current was due to ATP independent opening of the channels. Following AM546 treatment, zinc (100 µM or acidic external solution (pH 6.5 elicited inward currents when applied without any exogenous ATP. In the double mutant K69C/H319K, zinc elicited much larger inward currents, while acidic pH generated outward currents. Suramin, which is an antagonist of wild type receptors, behaved as an agonist at AM546-treated K69C receptors. Several other cysteine-reactive fluorophores tested on K69C did not cause these changes. These modified receptors show promise as a tool for studying the mechanisms of P2X receptor activation.

  18. Zoospore exudates from Phytophthora nicotianae affect immune responses in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Ping Kong

    Full Text Available Zoospore exudates play important roles in promoting zoospore communication, homing and germination during plant infection by Phytophthora. However, it is not clear whether exudates affect plant immunity. Zoospore-free fluid (ZFF and zoospores of P. nicotianae were investigated comparatively for effects on resistance of Arabidopsis thaliana Col-0 and mutants that affect signaling mediated by salicylic acid (SA and jasmonic acid (JA: eds16 (enhanced disease susceptibility16, pad4 (phytoalexin deficient4, and npr1 (nonexpressor of pathogenesis-related genes1. Col-0 attracted more zoospores and had severe tissue damage when flooded with a zoospore suspension in ZFF. Mutants treated with ZFF alone developed disease symptoms similar to those inoculated with zoospores and requirements of EDS16 and PAD4 for plant responses to zoospores and the exudates was apparent. Zoospore and ZFFs also induced expression of the PR1 and PDF1.2 marker genes for defense regulated by SA and JA, respectively. However, ZFF affected more JA defense signaling, down regulating PR1 when SA signaling or synthesis is deficient, which may be responsible for Arabidopsis mutant plants more susceptible to infection by high concentration of P. nicotianae zoospores. These results suggest that zoospore exudates can function as virulence factors and inducers of plant immune responses during plant infection by Phytophthora.

  19. A Sordaria macrospora mutant lacking the leu1 gene shows a developmental arrest during fruiting body formation.

    Science.gov (United States)

    Kück, Ulrich

    2005-10-01

    Developmental mutants with defects in fruiting body formation are excellent resources for the identification of genetic components that control cellular differentiation processes in filamentous fungi. The mutant pro4 of the ascomycete Sordaria macrospora is characterized by a developmental arrest during the sexual life cycle. This mutant generates only pre-fruiting bodies (protoperithecia), and is unable to form ascospores. Besides being sterile, pro4 is auxotrophic for leucine. Ascospore analysis revealed that the two phenotypes are genetically linked. After isolation of the wild-type leu1 gene from S. macrospora, complementation experiments demonstrated that the gene was able to restore both prototrophy and fertility in pro4. To investigate the control of leu1 expression, other genes involved in leucine biosynthesis specifically and in the general control of amino acid biosynthesis ("cross-pathway control") have been analysed using Northern hybridization and quantitative RT-PCR. These analyses demonstrated that genes of leucine biosynthesis are transcribed at higher levels under conditions of amino acid starvation. In addition, the expression data for the cpc1 and cpc2 genes indicate that cross-pathway control is superimposed on leucine-specific regulation of fruiting body development in the leu1 mutant. This was further substantiated by growth experiments in which the wild-type strain was found to show a sterile phenotype when grown on a medium containing the amino acid analogue 5-methyl-tryptophan. Taken together, these data show that pro4 represents a novel mutant type in S. macrospora, in which amino acid starvation acts as a signal that interrupts the development of the fruiting body.

  20. Phosphorus Partitioning of Soybean Lines Containing Different Mutant Alleles of Two Soybean Seed-Specific Adenosine Triphosphate-Binding Cassette Phytic Acid Transporter Paralogs

    Directory of Open Access Journals (Sweden)

    Jason D. Gillman

    2013-03-01

    Full Text Available Seed phytate is a repository of P and minerals in soybean [ (L. Merr.] seeds that limits P and mineral bioavailability for monogastric animals (e.g., humans, swine [], and poultry [especially chicken, ] due to insufficient digestive tract phytase activity. We previously identified epistatic recessive mutations affecting two paralogous adenosine triphosphate-binding cassette phytic acid transporter genes (one a nonsense mutation in and the other a missense mutation in as the molecular genetic basis in the ethyl methanesulfonate (EMS-induced mutant low phytate soybean line M153. An additional mutant low phytate line, M766, contained one single nucleotide polymorphism within the ninth intron of the locus as well as a nonsense mutation in . The objectives of this research were to clarify the genetics underlying the low phytate phenotype in line M766 and to determine P partitioning in new combinations of mutant alleles from M766 and M153. Inheritance of nonsense alleles affecting both ( genes (one from M153 and one from M766 led to the production of viable seeds that contained transgressive reductions in total seed phytate and significantly higher levels of inorganic phosphate than has been reported for nontransgenic soybean material and will allow efficient molecular selection of soybeans with even greater reductions of phytate for improved quality soybean meal.

  1. Isolation of new gravitropic mutants under hypergravity conditions

    OpenAIRE

    Akiko Mori; Masatsugu Toyota; Masatsugu Toyota; Masayoshi Shimada; Mika Mekata; Tetsuya Kurata; Masao Tasaka; Miyo Terao Morita

    2016-01-01

    Forward genetics is a powerful approach used to link genotypes and phenotypes, and mutant screening/analysis has provided deep insights into many aspects of plant physiology. Gravitropism is a tropistic response in plants, in which hypocotyls and stems sense the direction of gravity and grow upwards. Previous studies of gravitropic mutants have suggested that shoot endodermal cells in Arabidopsis stems and hypocotyls are capable of sensing gravity (i.e., statocytes). In the present study, we ...

  2. Evaluation Of Yield And Chemical Properties Of Some Faba Bean (Vicia FABA L.) Mutants Induced By GAMMA Radiation And Ethyl Methane Sulphonate

    International Nuclear Information System (INIS)

    Sallam, E.M.; Nasr, E.H.; Attia, Z.M.; Shawki, H.A.

    2013-01-01

    This investigation aims to study the effect of physical and chemical mutagens on the yield and chemical properties as well as amino acids composition of defatted faba bean (Vicia faba L.) seeds meal as a control (Giza 2) compared with mutants produced by gamma radiation and ethyl methane sulphonate (EMS). Also, the functional properties of these samples were determined. The results indicated high differences between mutation for seed yield and its components than the untreated samples. In addition, radiation mutation of faba bean seeds showed slight increase in protein content as the main constituent of faba bean seeds as well as total oil percentages in some mutant of these seeds in return of decreasing in total carbohydrate. Furthermore, radiation mutation had detectable effects on the total amino acids contents of faba bean seeds meal which had a higher percentages on essential amino acids (EAA) and non-essential amino acids (NEAA) and mutant 3 was the highest values of EAA and NEAA as compared to the control. On the other hand, radiation mutation improved the protein functional properties of some mutant of faba bean meal flour than the other mutant samples as compared to local commercial variety

  3. Growth of catalase A and catalase T deficient mutant strains of Saccharomyces cerevisiae on ethanol and oleic acid : Growth profiles and catalase activities in relation to microbody proliferation

    NARCIS (Netherlands)

    Klei, Ida J. van der; Rytka, Joanna; Kunau, Wolf H.; Veenhuis, Marten

    The parental strain (A+T+) of Saccharomyces cerevisiae and mutants, deficient in catalase T (A+T-), catalase A (A-T+) or both catalases (A-T-), grew on ethanol and oleic acid with comparable doubling times. Specific activities of catalase were low in glucose- and ethanol-grown cells. In the two

  4. Officially released mutant varieties in China

    International Nuclear Information System (INIS)

    Liu, L.; Van Zanten, L.; Shu, Q.Y.; Maluszynski, M.

    2004-01-01

    The use of mutation techniques for crop improvement in China has a long and well-established tradition of more than 50 years. As the result of intensive research in many institutes dealing with application of nuclear technologies more than 620 cultivars of 44 crop species have been released. Numerous mutant varieties have been grown on a large scale bringing significant economic impact, sustaining crop production and greatly contributing to increase of food production also in stress prone areas of the country. However, there is still missing information not only on the number of mutant varieties released in particular crop species but also on mutagens applied, selection approaches and on the use of mutants in cross breeding. Numerous Chinese scientists collected and systematized this information. Results of their work were often published in local scientific journals in the Chinese language and as such were unavailable to breeders from other countries. Having this in mind, we requested Dr. Liu Luxiang, the Director of the Department of Plant Mutation Breeding and Genetics, Institute for Application of Atomic Energy, Chinese Academy of Agricultural Sciences in Beijing to help us in finding as much information as possible on mutant varieties officially released in China. The data has been collected in close collaboration with his colleagues from various institutions all over the country and then evaluated, edited and prepared for publication by our team responsible for the FAO/IAEA Database of Officially Released Mutant Varieties. We would like to thank all Chinese colleagues who contributed to this list of Chinese mutant varieties. We hope that this publication will stimulate plant breeders in China to collect more information on released mutant varieties and especially on the use of mutated genes in cross breeding. (author)

  5. Prion propagation in cells expressing PrP glycosylation mutants.

    Science.gov (United States)

    Salamat, Muhammad K; Dron, Michel; Chapuis, Jérôme; Langevin, Christelle; Laude, Hubert

    2011-04-01

    Infection by prions involves conversion of a host-encoded cell surface protein (PrP(C)) to a disease-related isoform (PrP(Sc)). PrP(C) carries two glycosylation sites variably occupied by complex N-glycans, which have been suggested by previous studies to influence the susceptibility to these diseases and to determine characteristics of prion strains. We used the Rov cell system, which is susceptible to sheep prions, to generate a series of PrP(C) glycosylation mutants with mutations at one or both attachment sites. We examined their subcellular trafficking and ability to convert into PrP(Sc) and to sustain stable prion propagation in the absence of wild-type PrP. The susceptibility to infection of mutants monoglycosylated at either site differed dramatically depending on the amino acid substitution. Aglycosylated double mutants showed overaccumulation in the Golgi compartment and failed to be infected. Introduction of an ectopic glycosylation site near the N terminus fully restored cell surface expression of PrP but not convertibility into PrP(Sc), while PrP(C) with three glycosylation sites conferred cell permissiveness to infection similarly to the wild type. In contrast, predominantly aglycosylated molecules with nonmutated N-glycosylation sequons, produced in cells expressing glycosylphosphatidylinositol-anchorless PrP(C), were able to form infectious PrP(Sc). Together our findings suggest that glycosylation is important for efficient trafficking of anchored PrP to the cell surface and sustained prion propagation. However, properly trafficked glycosylation mutants were not necessarily prone to conversion, thus making it difficult in such studies to discern whether the amino acid changes or glycan chain removal most influences the permissiveness to prion infection.

  6. Stomatal responses to carbon dioxide of isolated epidermis from a C/sub 3/ plant, the Argenteum mutant of Pisum sativum L. , and a crassulacean-acid-metabolism plant Kalanchoe daigremontiana Hamet et Perr

    Energy Technology Data Exchange (ETDEWEB)

    Jewer, P.C.; Neales, T.F.; Incoll, L.D.

    1985-01-01

    The response of stomata in isolated epidermis to the concentration of CO/sub 2/ in the gaseous phase was examined in a C/sub 3/ species, the Argenteum mutant of Pisum sativum, and a crassulacean-acid-metabolism (CAM) species, Kalanchoe daigremontiana. Epidermis from leaves of both species was incubated on buffer solutions in the presence of air containing various volume fractions of CO/sub 2/ (0 to 10,000 x 10/sup -6/). In both species and in the light and in darkness, the effect of CO/sub 2/ was to inhibit stomatal opening, the maximum inhibition of opening occurring in the range 0 to 360 x 10/sup -6/. The inhibition of opening per unit change in concentration was greatest between volume fractions of 0 and 240 x 10/sup -6/. There was little further closure above the volume fraction of 360 x 10/sup -6/, i.e. approximately ambient concentration of CO/sub 2/. Thus, although leaves of CAM species may experience much higher internal concentrations of CO/sub 2/ in the light than those of C/sub 3/ plants, this does not affect the sensitivity of their stomata to CO/sub 2/ concentration or the range over which they respond. Stomatal responses to CO/sub 2/ were similar in both the light and the dark, indicating that effects of CO/sub 2/ on stomata occur via mechanisms which are independent of light. The responses of stomata to CO/sub 2/ in the gaseous phase took place without the treatments changing the pH of the buffered solutions. Thus, it is unlikely that CO/sub 2/ elicited stomatal movement by changing either the pH or the HCO/sub 3//sup -//CO/sub 3//sup 2 -/ equilibria. It is suggested that the concentration of dissolved unhydrated CO/sub 2/ may be the effector of stomatal movement and that its activity is related to its reactivity with amines.

  7. The Response Regulator YycF Inhibits Expression of the Fatty Acid Biosynthesis Repressor FabT in Streptococcus pneumoniae

    Science.gov (United States)

    Mohedano, Maria L.; Amblar, Mónica; de la Fuente, Alicia; Wells, Jerry M.; López, Paloma

    2016-01-01

    The YycFG (also known as WalRK, VicRK, MicAB, or TCS02) two-component system (TCS) is highly conserved among Gram-positive bacteria with a low G+C content. In Streptococcus pneumoniae the YycF response regulator has been reported to be essential due to its control of pcsB gene expression. Previously we showed that overexpression of yycF in S. pneumoniae TIGR4 altered the transcription of genes involved in cell wall metabolism and fatty acid biosynthesis, giving rise to anomalous cell division and increased chain length of membrane fatty acids. Here, we have overexpressed the yycFG system in TIGR4 wild-type strain and yycF in a TIGR4 mutant depleted of YycG, and analyzed their effects on expression of proteins involved in fatty acid biosynthesis during activation of the TCS. We demonstrate that transcription of the fab genes and levels of their products were only altered in the YycF overexpressing strain, indicating that the unphosphorylated form of YycF is involved in the regulation of fatty acid biosynthesis. In addition, DNA-binding assays and in vitro transcription experiments with purified YycF and the promoter region of the FabTH-acp operon support a direct inhibition of transcription of the FabT repressor by YycF, thus confirming the role of the unphosphorylated form in transcriptional regulation. PMID:27610104

  8. Sterol-mediated regulation of mevalonic acid synthesis. Accumulation of 4-carboxysterols as the predominant sterols synthesized in a Chinese hamster ovary cell cholesterol auxotroph (mutant 215)

    International Nuclear Information System (INIS)

    Plemenitas, A.; Havel, C.M.; Watson, J.A.

    1990-01-01

    Chinese hamster ovary-215 (CHO-215) mutant cells are auxotrophic for cholesterol. Berry and Chang (Berry, D. J., and Chang, T. Y. (1982) Biochemistry 21, 573-580) suggested that the metabolic lesion was at the level of 4-methyl sterol oxidation. However, the observed cellular accumulation of lanosterol was not consistent with a defect at this metabolic site. With the use of a novel Silica Sep Pak sterol separation procedure, we demonstrated that 60-80% of the acetonesoluble lipid radioactivity in [5-3H]mevalonate-labeled CHO-215 cells was incorporated into acidic sterols. 7(8),Cholesten-4 beta-methyl,4 alpha-carboxy,3 beta-ol was the dominant end product. In addition to this acidic sterol, 7(8),24-cholestadien,4 beta-methyl,4 alpha-carboxy,3 beta-ol and 7(8),24-cholestadien,4 alpha-carboxy,3 beta-ol were also isolated. Incubation of cell-free extracts with [3H]7(8)-cholesten-4 beta-methyl, 4 alpha-carboxy,3 beta-ol and pyridine nucleotides confirmed that CHO-215 4-carboxysterol decarboxylase activity was less than 1% of that for wild type cells. Thus, a correspondence between decreased 4-carboxysterol decarboxylase activity and the spectrum of accumulated sterol products by intact CHO-215 cells was demonstrated. No detectable cholesterol was synthesized by CHO-215 cells. 3H-Product accumulation studies demonstrated that 7(8),24-cholestadien, 4 beta-methyl,4 alpha-carboxy,3 beta-ol increased prior to its subsequent saturation at the delta 24 carbon. Furthermore, the steady state ratio for delta 24-saturated acidic sterols/unsaturated acidic sterols was dependent on media cholesterol source and amount. Finally, the accumulated acidic sterol(s) were not regulatory signal molecules for the modulation of 3-hydroxy-3-methyl-glutaryl coenzyme. A reductase activity in response to cholesterol availability

  9. Membrane composition and physiological activity of plastids from an oenothera plastome mutator-induced chloroplast mutant.

    Science.gov (United States)

    Johnson, E M; Sears, B B

    1990-01-01

    Plastids were isolated from a plastome mutator-induced mutant (pm7) of Oenothera hookeri and were analyzed for various physiological and biochemical attributes. No photosynthetic electron transport activity was detected in the mutant plastids. This is consistent with previous ultrastructural analysis showing the absence of thylakoid membranes in the pm7 plastids and with the observation of aberrant processing and accumulation of chloroplast proteins in the mutant. In comparison to wild type, the mutant tissue lacks chlorophyll, and has significant differences in levels of four fatty acids. The analyses did not reveal any differences in carotenoid levels nor in the synthesis of several chloroplast lipids. The consequences of the altered composition of the chloroplast membrane are discussed in terms of their relation to the aberrant protein processing of the pm7 plastids. The pigment, fatty acid, and lipid measurements were also performed on two distinct nuclear genotypes (A/A and A/C) which differ in their compatibility with the plastid genome (type I) contained in these lines. In these cases, only chlorophyll concentrations differed significantly.

  10. Homocysteine regulates fatty acid and lipid metabolism in yeast.

    Science.gov (United States)

    Visram, Myriam; Radulovic, Maja; Steiner, Sabine; Malanovic, Nermina; Eichmann, Thomas O; Wolinski, Heimo; Rechberger, Gerald N; Tehlivets, Oksana

    2018-04-13

    S -Adenosyl-l-homocysteine hydrolase (AdoHcy hydrolase; Sah1 in yeast/AHCY in mammals) degrades AdoHcy, a by-product and strong product inhibitor of S -adenosyl-l-methionine (AdoMet)-dependent methylation reactions, to adenosine and homocysteine (Hcy). This reaction is reversible, so any elevation of Hcy levels, such as in hyperhomocysteinemia (HHcy), drives the formation of AdoHcy, with detrimental consequences for cellular methylation reactions. HHcy, a pathological condition linked to cardiovascular and neurological disorders, as well as fatty liver among others, is associated with a deregulation of lipid metabolism. Here, we developed a yeast model of HHcy to identify mechanisms that dysregulate lipid metabolism. Hcy supplementation to wildtype cells up-regulated cellular fatty acid and triacylglycerol content and induced a shift in fatty acid composition, similar to changes observed in mutants lacking Sah1. Expression of the irreversible bacterial pathway for AdoHcy degradation in yeast allowed us to dissect the impact of AdoHcy accumulation on lipid metabolism from the impact of elevated Hcy. Expression of this pathway fully suppressed the growth deficit of sah1 mutants as well as the deregulation of lipid metabolism in both the sah1 mutant and Hcy-exposed wildtype, showing that AdoHcy accumulation mediates the deregulation of lipid metabolism in response to elevated Hcy in yeast. Furthermore, Hcy supplementation in yeast led to increased resistance to cerulenin, an inhibitor of fatty acid synthase, as well as to a concomitant decline of condensing enzymes involved in very long-chain fatty acid synthesis, in line with the observed shift in fatty acid content and composition. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. Isolation of New Gravitropic Mutants under Hypergravity Conditions

    OpenAIRE

    Mori, Akiko; Toyota, Masatsugu; Shimada, Masayoshi; Mekata, Mika; Kurata, Tetsuya; Tasaka, Masao; Morita, Miyo T.

    2016-01-01

    Forward genetics is a powerful approach used to link genotypes and phenotypes, and mutant screening/analysis has provided deep insights into many aspects of plant physiology. Gravitropism is a tropistic response in plants, in which hypocotyls and stems sense the direction of gravity and grow upward. Previous studies of gravitropic mutants have suggested that shoot endodermal cells in Arabidopsis stems and hypocotyls are capable of sensing gravity (i.e., statocytes). In the present study, we r...

  12. Investigating the Role of the Arabidopsis Homologue of the Human G3BP in RNA Metabolism, Cellular Stress Responses and Innate Immunity

    KAUST Repository

    Abulfaraj, Aala A.

    2018-01-01

    immunity and defense immune responses. Atg3bp1 mutant lines show constitutive stomata closure, expression of a number of key defense marker genes, and accumulate salicylic acid but not jasmonic acid. Furthermore, Atg3bp1 plants exhibit enhanced resistance

  13. Wild-type catalase peroxidase vs G279D mutant type: Molecular basis of Isoniazid drug resistance in Mycobacterium tuberculosis.

    Science.gov (United States)

    Singh, Aishwarya; Singh, Aditi; Grover, Sonam; Pandey, Bharati; Kumari, Anchala; Grover, Abhinav

    2018-01-30

    Mycobacterium tuberculosis katG gene is responsible for production of an enzyme catalase peroxidase that peroxidises and activates the prodrug Isoniazid (INH), a first-line antitubercular agent. INH interacts with catalase peroxidase enzyme within its heme pocket and gets converted to an active form. Mutations occurring in katG gene are often linked to reduced conversion rates for INH. This study is focussed on one such mutation occurring at residue 279, where glycine often mutates to aspartic acid (G279D). In the present study, several structural analyses were performed to study the effect of this mutation on functionality of KatG protein. On comparison, mutant protein exhibited a lower docking score, smaller binding cavity and reduced affinity towards INH. Molecular dynamics analysis revealed the mutant to be more rigid and less compact than the native protein. Essential dynamics analysis determined correlated motions of residues within the protein structure. G279D mutant was found to have many residues that showed related motions and an undesirable effect on the functionality of protein. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Functional analysis of mutant human carnitine acylcarnitine translocases in yeast

    NARCIS (Netherlands)

    IJlst, L.; van Roermund, C. W.; Iacobazzi, V.; Oostheim, W.; Ruiter, J. P.; Williams, J. C.; Palmieri, F.; Wanders, R. J.

    2001-01-01

    Long chain fatty acids are translocated as carnitine esters across the mitochondrial inner membrane by carnitine acylcarnitine translocase (CACT). We report functional studies on the mutant CACT proteins from a severe and a mild patient with CACT deficiency. CACT activities in fibroblasts of both

  15. Analysis of AtCry1 and Mutants

    Science.gov (United States)

    Burdick, Derek; Purvis, Adam; Ahmad, Margaret; Link, Justin J.; Engle, Dorothy

    Cryptochrome is an incredibly versatile protein that influences numerous biological processes such as plant growth, bird migration, and sleep cycles. Due to the versatility of this protein, understanding the mechanism would allow for advances in numerous fields such as crop growth, animal behavior, and sleep disorders. It is known that cryptochrome requires blue light to function, but the exact processes in the regulation of biological activity are still not fully understood. It is believed that the c-terminal domain of the protein undergoes a conformational change when exposed to blue light which allows for biological function. Three different non-functioning mutants were tested during this study to gain insight on the mechanism of cryptochrome. Absorbance spectra showed a difference between two of the mutants and the wild type with one mutant showing little difference. Immunoprecipitation experiments were also conducted to identify the different c-terminal responses of the mutants. By studying non functioning mutants of this protein, the mechanism of the protein can be further characterized. This two-month research experience in Paris allowed us to experience international and interdisciplinary collaborations in science and immerse in a different culture. The Borcer Fund for Student Research, Xavier University, Cincinnati, OH, and John Hauck Foundation.

  16. Construction of a mutant of Actinoplanes sp. N902-109 that produces a new rapamycin analog.

    Science.gov (United States)

    Huang, He; Gao, Ping; Zhao, Qi; Hu, Hai-Feng

    2018-03-01

    In the present study, we introduced point mutations into Ac_rapA which encodes a polyketide synthase responsible for rapamycin biosynthesis in Actinoplanes sp. N902-109, in order to construct a mutant with an inactivated enoylreductase (ER) domain, which was able to synthesize a new rapamycin analog. Based on the homologous recombination induced by double-strand breaks in chromosome mediated by endonuclease I-SceI, the site-directed mutation in the first ER domain of Ac_rapA was introduced using non-replicating plasmid pLYERIA combined with an I-SceI expression plasmid. Three amino acid residues of the active center, Ala-Gly-Gly, were converted to Ala-Ser-Pro. The broth of the mutant strain SIPI-027 was analyzed by HPLC and a new peak with the similar UV spectrum to that of rapamycin was found. The sample of the new peak was prepared by solvent extraction, column chromatography, and crystallization methods. The structure of new compound, named as SIPI-rapxin, was elucidated by determining and analyzing its MS and NMR spectra and its biological activity was assessed using mixed lymphocyte reaction (MLR). An ER domain-deficient mutant of Actinoplanes sp. N902-109, named as SIPI-027, was constructed, which produced a novel rapamycin analog SIPI-rapxin and its structure was elucidated to be 35, 36-didehydro-27-O-demethylrapamycin. The biological activity of SIPI-rapxin was better than that of rapamycin. In conclusion, inactivation of the first ER domain of rapA, one of the modular polyketide synthase responsible for macro-lactone synthesis of rapamycin, gave rise to a mutant capable of producing a novel rapamycin analog, 35, 36-didehydro-27-O-demethylrapamycin, demonstrating that the enoylreductase domain was responsible for the reduction of the double bond between C-35 and C-36 during rapamycin synthesis. Copyright © 2018 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

  17. Plants Regeneration Derived From Various on Peanut on Mutant Lines

    International Nuclear Information System (INIS)

    Dewi, Kumala; Masrizal; Mugiono

    1998-01-01

    The study of calli, greenspot formation and shoot regeneration on peanut mutant lines has ben conducted by MS media. Three explants derived from shoot tips, embryo and seeding root of two mutant lines a/20/3 and D/25/3/2 were used in this experiment. the explants were cultured on modified MS media enriched by vitamins, growth regulation, amino acids for fourth teen calli were transferred on regeneration media. The ability of calli formation and plant regeneration of each explant and genotypes of plants was varied. Greenspot and shoot formation were observed seventh days after the calli transferred on regeneration media. It is shown that the ability of calli, greenspot and shoot formation of each explants and genotypes was varied. the high ability of calli, greenspot and shoot formation were found in explant derived from shoot tip and embryo. Seedling root explant has lower ability in calli formation, while greenspot and shoot was formatted. The ability of calli, greenspot and shoot formation on A/20/3 mutant line was better than D/25/3/2 mutant line. (author)

  18. Single Amino Acid Modification of Adeno-Associated Virus Capsid Changes Transduction and Humoral Immune Profiles

    Science.gov (United States)

    Diprimio, Nina; Bowles, Dawn E.; Hirsch, Matthew L.; Monahan, Paul E.; Asokan, Aravind; Rabinowitz, Joseph; Agbandje-McKenna, Mavis

    2012-01-01

    Adeno-associated virus (AAV) vectors have the potential to promote long-term gene expression. Unfortunately, humoral immunity restricts patient treatment and in addition provides an obstacle to the potential option of vector readministration. In this study, we describe a comprehensive characterization of the neutralizing antibody (NAb) response to AAV type 1 (AAV1) through AAV5 both in vitro and in vivo. These results demonstrated that NAbs generated from one AAV type are unable to neutralize the transduction of other types. We extended this observation by demonstrating that a rationally engineered, muscle-tropic AAV2 mutant containing 5 amino acid substitutions from AAV1 displayed a NAb profile different from those of parental AAV2 and AAV1. Here we found that a single insertion of Thr from AAV1 into AAV2 capsid at residue 265 preserved high muscle transduction, while also changing the immune profile. To better understand the role of Thr insertion at position 265, we replaced all 20 amino acids and evaluated both muscle transduction and the NAb response. Of these variants, 8 mutants induced higher muscle transduction than AAV2. Additionally, three classes of capsid NAb immune profile were defined based on the ability to inhibit transduction from AAV2 or mutants. While no relationship was found between transduction, amino acid properties, and NAb titer or its cross-reactivity, these studies map a critical capsid motif involved in all steps of AAV infectivity. Our results suggest that AAV types can be utilized not only as templates to generate mutants with enhanced transduction efficiency but also as substrates for repeat administration. PMID:22593151

  19. Biochemical Analysis of Two Single Mutants that Give Rise to a Polymorphic G6PD A-Double Mutant

    Directory of Open Access Journals (Sweden)

    Edson Jiovany Ramírez-Nava

    2017-10-01

    Full Text Available Glucose-6-phosphate dehydrogenase (G6PD is a key regulatory enzyme that plays a crucial role in the regulation of cellular energy and redox balance. Mutations in the gene encoding G6PD cause the most common enzymopathy that drives hereditary nonspherocytic hemolytic anemia. To gain insights into the effects of mutations in G6PD enzyme efficiency, we have investigated the biochemical, kinetic, and structural changes of three clinical G6PD variants, the single mutations G6PD A+ (Asn126AspD and G6PD Nefza (Leu323Pro, and the double mutant G6PD A− (Asn126Asp + Leu323Pro. The mutants showed lower residual activity (≤50% of WT G6PD and displayed important kinetic changes. Although all Class III mutants were located in different regions of the three-dimensional structure of the enzyme and were not close to the active site, these mutants had a deleterious effect over catalytic activity and structural stability. The results indicated that the G6PD Nefza mutation was mainly responsible for the functional and structural alterations observed in the double mutant G6PD A−. Moreover, our study suggests that the G6PD Nefza and G6PD A− mutations affect enzyme functions in a similar fashion to those reported for Class I mutations.

  20. Growth of catalase A and catalase T deficient mutant strains of Saccharomyces cerevisiae on ethanol and oleic acid: Growth profiles and catalase activities in relation to microbody proliferation

    OpenAIRE

    Klei, Ida J. van der; Rytka, Joanna; Kunau, Wolf H.; Veenhuis, Marten

    1990-01-01

    The parental strain (A+T+) of Saccharomyces cerevisiae and mutants, deficient in catalase T (A+T-), catalase A (A-T+) or both catalases (A-T-), grew on ethanol and oleic acid with comparable doubling times. Specific activities of catalase were low in glucose- and ethanol-grown cells. In the two oleic acid-grown A+-strains (A+T+ and A+T-) high catalase activities were found; catalase activity invariably remained low in the A-T+ strain and was never detected in the A-T- strain. The levels of β-...

  1. The coat protein of Alfalfa mosaic virus interacts and interferes with the transcriptional activity of the bHLH transcription factor ILR3 promoting salicylic acid-dependent defence signalling response.

    Science.gov (United States)

    Aparicio, Frederic; Pallás, Vicente

    2017-02-01

    During virus infection, specific viral component-host factor interaction elicits the transcriptional reprogramming of diverse cellular pathways. Alfalfa mosaic virus (AMV) can establish a compatible interaction in tobacco and Arabidopsis hosts. We show that the coat protein (CP) of AMV interacts directly with transcription factor (TF) ILR3 of both species. ILR3 is a basic helix-loop-helix (bHLH) family member of TFs, previously proposed to participate in diverse metabolic pathways. ILR3 has been shown to regulate NEET in Arabidopsis, a critical protein in plant development, senescence, iron metabolism and reactive oxygen species (ROS) homeostasis. We show that the AMV CP-ILR3 interaction causes a fraction of this TF to relocate from the nucleus to the nucleolus. ROS, pathogenesis-related protein 1 (PR1) mRNAs, salicylic acid (SA) and jasmonic acid (JA) contents are increased in healthy Arabidopsis loss-of-function ILR3 mutant (ilr3.2) plants, which implicates ILR3 in the regulation of plant defence responses. In AMV-infected wild-type (wt) plants, NEET expression is reduced slightly, but is induced significantly in ilr3.2 mutant plants. Furthermore, the accumulation of SA and JA is induced in Arabidopsis wt-infected plants. AMV infection in ilr3.2 plants increases JA by over 10-fold, and SA is reduced significantly, indicating an antagonist crosstalk effect. The accumulation levels of viral RNAs are decreased significantly in ilr3.2 mutants, but the virus can still systemically invade the plant. The AMV CP-ILR3 interaction may down-regulate a host factor, NEET, leading to the activation of plant hormone responses to obtain a hormonal equilibrium state, where infection remains at a level that does not affect plant viability. © 2016 BSPP AND JOHN WILEY & SONS LTD.

  2. Acidic deposition: State of science and technology. Report 11. Historical changes in surface-water acid-base chemistry in response to acidic deposition. Final report

    International Nuclear Information System (INIS)

    Sullivan, T.J.; Small, M.J.; Kingston, J.C.; Bernert, J.A.; Thomas, D.R.

    1990-09-01

    The objectives of the analyses reported in the State of Science report are to: identify the lake and stream populations in the United States that have experienced chronic changes in biologically significant constituents of surface water chemistry (e.g. pH, Al) in response to acidic deposition; quantify biologically meaningful historical changes in chronic surface water chemistry associated with acidic deposition, with emphasis on ANC, pH, and Al; estimate the proportion of lakes nor acidic that were not acidic in pre-industrial times; estimate the proportional response of each of the major chemical constituents that have changed in response to acidic deposition using a subset of statistically selected Adirondack lakes for which paleolimnological reconstructions of pre-industrial surface water chemistry have been performed; evaluate and improve, where appropriate and feasible, empirical models of predicting changes in ANC; and evaluate the response of seepage lakes to acidic deposition

  3. Breeding cultivars of barley and mustard containing biochemical mutants

    Energy Technology Data Exchange (ETDEWEB)

    Oram, R N [Division of Plant industry, CSIRO, Canberra (Australia)

    1990-01-01

    Full text: The inactivation of dominant and co-dominant alleles is becoming increasingly important in changing the composition of seed carbohydrates, protein, oil, fibre and secondary products to suit modern food and feed technologies. In barley, breeding lines adapted to south-eastern Australian conditions have been developed containing a waxy endosperm from the Japanese variety 'Sumire Mochi', the high lysine gene lys from cv. 'Hiproly' of Ethiopia, and the induced high lysine mutant gene lys 3a from 'Risoe 1508'. The improved mutant lines yield 12-34% less than the highest yielding feed barley. The lys and lys 3a alleles suppress the formation of prolamins, the waxy allele inhibits the formation of amylose. It seems difficult to modify the background genotype to fully compensate for the reduction of major storage carbohydrate or protein compounds. However, waxy barleys have uses in some human foods and a premium can be paid to producers. The grain of the provisionally-patented waxy cultivar Wasiro is suitable for pearling. It contains 5% {beta}-glucan (soluble fibre) and therefore should be as effective as oat bran for reducing blood cholesterol. In Indian mustard (Brassica juncea), three cultivars differing in date of maturity, each containing the spontaneous mutant alleles for low erucic acid levels in the seed oil, have been developed to produce a high quality, mildly flavoured cooking/salad oil. The concentration of glucosinolates in the seed meal must be reduced to make it palatable and non-toxic to pigs and poultry. Three B. juncea lines were treated in up to four successive generations with gamma rays or EMS. 60,000 seed samples were analysed in subsequent generations. Two induced mutants with reduced glucosinolate concentrations are now available besides 4 naturally-occurring sources with only little reduced yields. Recombination may give a high-yielding low erucic acid and low glucosinolate variety of B. juncea. (author)

  4. Breeding cultivars of barley and mustard containing biochemical mutants

    International Nuclear Information System (INIS)

    Oram, R.N.

    1990-01-01

    Full text: The inactivation of dominant and co-dominant alleles is becoming increasingly important in changing the composition of seed carbohydrates, protein, oil, fibre and secondary products to suit modern food and feed technologies. In barley, breeding lines adapted to south-eastern Australian conditions have been developed containing a waxy endosperm from the Japanese variety 'Sumire Mochi', the high lysine gene lys from cv. 'Hiproly' of Ethiopia, and the induced high lysine mutant gene lys 3a from 'Risoe 1508'. The improved mutant lines yield 12-34% less than the highest yielding feed barley. The lys and lys 3a alleles suppress the formation of prolamins, the waxy allele inhibits the formation of amylose. It seems difficult to modify the background genotype to fully compensate for the reduction of major storage carbohydrate or protein compounds. However, waxy barleys have uses in some human foods and a premium can be paid to producers. The grain of the provisionally-patented waxy cultivar Wasiro is suitable for pearling. It contains 5% β-glucan (soluble fibre) and therefore should be as effective as oat bran for reducing blood cholesterol. In Indian mustard (Brassica juncea), three cultivars differing in date of maturity, each containing the spontaneous mutant alleles for low erucic acid levels in the seed oil, have been developed to produce a high quality, mildly flavoured cooking/salad oil. The concentration of glucosinolates in the seed meal must be reduced to make it palatable and non-toxic to pigs and poultry. Three B. juncea lines were treated in up to four successive generations with gamma rays or EMS. 60,000 seed samples were analysed in subsequent generations. Two induced mutants with reduced glucosinolate concentrations are now available besides 4 naturally-occurring sources with only little reduced yields. Recombination may give a high-yielding low erucic acid and low glucosinolate variety of B. juncea. (author)

  5. Both the Jasmonic Acid and the Salicylic Acid Pathways Contribute to Resistance to the Biotrophic Clubroot Agent Plasmodiophora brassicae in Arabidopsis.

    Science.gov (United States)

    Lemarié, Séverine; Robert-Seilaniantz, Alexandre; Lariagon, Christine; Lemoine, Jocelyne; Marnet, Nathalie; Jubault, Mélanie; Manzanares-Dauleux, Maria J; Gravot, Antoine

    2015-11-01

    The role of salicylic acid (SA) and jasmonic acid (JA) signaling in resistance to root pathogens has been poorly documented. We assessed the contribution of SA and JA to basal and partial resistance of Arabidopsis to the biotrophic clubroot agent Plasmodiophora brassicae. SA and JA levels as well as the expression of the SA-responsive genes PR2 and PR5 and the JA-responsive genes ARGAH2 and THI2.1 were monitored in infected roots of the accessions Col-0 (susceptible) and Bur-0 (partially resistant). SA signaling was activated in Bur-0 but not in Col-0. The JA pathway was weakly activated in Bur-0 but was strongly induced in Col-0. The contribution of both pathways to clubroot resistance was then assessed using exogenous phytohormone application and mutants affected in SA or JA signaling. Exogenous SA treatment decreased clubroot symptoms in the two Arabidopsis accessions, whereas JA treatment reduced clubroot symptoms only in Col-0. The cpr5-2 mutant, in which SA responses are constitutively induced, was more resistant to clubroot than the corresponding wild type, and the JA signaling-deficient mutant jar1 was more susceptible. Finally, we showed that the JA-mediated induction of NATA1 drove N(δ)-acetylornithine biosynthesis in infected Col-0 roots. The 35S::NATA1 and nata1 lines displayed reduced or enhanced clubroot symptoms, respectively, thus suggesting that in Col-0 this pathway was involved in the JA-mediated basal clubroot resistance. Overall, our data support the idea that, depending on the Arabidopsis accession, both SA and JA signaling can play a role in partial inhibition of clubroot development in compatible interactions with P. brassicae. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  6. Selective response of dopamine in the presence of ascorbic acid ...

    African Journals Online (AJOL)

    Selective response of dopamine in the presence of ascorbic acid and uric acid at gold nanoparticles and multi-walled carbon nanotubes grafted with ethylene diamine tetraacetic acid modified electrode.

  7. Rhodosporidium toruloides BANNO: Dose-response relationship, mutagenic efficiency and spectrum of mutants of auxotrophy-producing mutations induced by ultraviolet light and N-methyl-N'-nitro-N-nitrosoguanidine

    International Nuclear Information System (INIS)

    Boettcher, F.; Samsonova, I.A.

    1978-01-01

    The kinetics, efficiency, and specificity of induction of forward mutations to auxotrophy by ultraviolet light (UV) and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) was examined in stationary phase cells of Rhodosporidium (Rhodotorula) wild strain Rg1. In comparison to the spontaneous level the frequency of auxotrophic mutants was increased more than 1000 times by both mutagens, however, the mutagenic efficiency of MNNG was higher than that of UV. We found that the forward mutation rate is a linear function of the applicated UV and MNNG doses in the range to 600 J m -2 or 25 mM x min, respectively. The 35 studied biosynthetic pathways to amino acids, purines, pyrimidines, and vitamins are genetically blocked at different frequencies, but there is not any significant difference between UV and MNNG induced frequencies of mutants with a specific requirement. However, in difference to the approximately equal distribution of the MNNG-induced nic mutants among the genetic blocks of the tryptophan-nicotinamide pathway, UV-induced nic mutants occurred with a higher frequency in the genes of the tryptophan pyrrolase and the 3-hydroxykynureninase than in the genes of the other enzymes of the pathway. (author)

  8. X-ray-sensitive mutants of Chinese hamster ovary cell line

    International Nuclear Information System (INIS)

    Jeggo, P.A.; Kemp, L.M.

    1983-01-01

    A standard technique of microbial genetics, which involves the transfer of cells from single colonies by means of sterile toothpicks, has been adapted to somatic cell genetics. Its use has been demonstrated in the isolation of X-ray-sensitive mutants of CHO cells. 9000 colonies have been tested and 6 appreciably X-ray-sensitive mutants were isolated. (D 10 values 5-10-fold of wild-type D 10 value.) A further 6 mutants were obtained which showed a slight level of sensitivity (D 10 values less than 2-fold of wild-type D 10 value). The 6 more sensitive mutants were also sensitive to bleomycin, a chemotherapeutic agent inducing X-ray-like damage. Cross-sensitivity to UV-irradiation and treatment with the alkylating agents, MMS, EMS and MNNG, was investigated for these mutants. Some sensitivity to these other agents was observed, but in all cases it was less severe than the level of sensitivity to X-irradiation. Each mutant showed a different overall response to the spectrum of agents examined and these appear to represent new mutant phenotypes derived from cultured mammalian cell lines. One mutant strain, xrs-7, was cross-sensitive to all the DNA-damaging agents, but was proficient in the repair of single-strand breaks. (Auth.)

  9. Response Surface Optimized Extraction of Total Triterpene Acids ...

    African Journals Online (AJOL)

    Purpose: To optimize extraction of total triterpene acids from loquat leaf and evaluate their in vitro antioxidant activities. Methods: The independent variables were ethanol concentration, extraction time, and solvent ratio, while the dependent variable was content of total triterpene acids. Composite design and response ...

  10. Chenodeoxycholic acid stimulated fibroblast growth factor 19 response

    DEFF Research Database (Denmark)

    Borup, C; Wildt, S; Rumessen, J J

    2017-01-01

    BACKGROUND: Bile acid diarrhoea is underdiagnosed and better diagnostic tests are needed. Fasting serum fibroblast growth factor-19 (FGF19) has insufficient diagnostic value, but this may be improved by stimulation. AIM: To explore if an impaired FGF19 response identifies primary bile acid...

  11. Membrane Composition and Physiological Activity of Plastids from an Oenothera Plastome Mutator-Induced Chloroplast Mutant 1

    Science.gov (United States)

    Johnson, Ellen M.; Sears, Barbara B.

    1990-01-01

    Plastids were isolated from a plastome mutator-induced mutant (pm7) of Oenothera hookeri and were analyzed for various physiological and biochemical attributes. No photosynthetic electron transport activity was detected in the mutant plastids. This is consistent with previous ultrastructural analysis showing the absence of thylakoid membranes in the pm7 plastids and with the observation of aberrant processing and accumulation of chloroplast proteins in the mutant. In comparison to wild type, the mutant tissue lacks chlorophyll, and has significant differences in levels of four fatty acids. The analyses did not reveal any differences in carotenoid levels nor in the synthesis of several chloroplast lipids. The consequences of the altered composition of the chloroplast membrane are discussed in terms of their relation to the aberrant protein processing of the pm7 plastids. The pigment, fatty acid, and lipid measurements were also performed on two distinct nuclear genotypes (A/A and A/C) which differ in their compatibility with the plastid genome (type I) contained in these lines. In these cases, only chlorophyll concentrations differed significantly. PMID:16667256

  12. Data on quantification of signaling pathways activated by KIT and PDGFRA mutants

    Directory of Open Access Journals (Sweden)

    Christelle Bahlawane

    2016-12-01

    Full Text Available The present data are related to the article entitled “Insights into ligand stimulation effects on gastro-intestinal stromal tumors signaling” (C. Bahlawane, M. Schmitz, E. Letellier, K. Arumugam, N. Nicot, P.V. Nazarov, S. Haan, 2016 [1]. Constitutive and ligand-derived signaling pathways mediated by KIT and PDGFRA mutated proteins found in gastrointestinal stromal tumors (GIST were compared. Expression of mutant proteins was induced by doxycycline in an isogenic background (Hek293 cells. Kit was identified by FACS at the cell surface and found to be quickly degraded or internalized upon SCF stimulation for both Kit Wild type and Kit mutant counterparts. Investigation of the main activated pathways in GIST unraveled a new feature specific for oncogenic KIT mutants, namely their ability to be further activated by Kit ligand, the stem cell factor (scf. We were also able to identify the MAPK pathway as the most prominent target for a common inhibition of PDGFRA and KIT oncogenic signaling. Western blotting and micro-array analysis were applied to analyze the capacities of the mutant to induce an effective STATs response. Among all Kit mutants, only Kit Ex11 deletion mutant was able to elicit an effective STATs response whereas all PDGFRA were able to do so.

  13. Promising semi-dwarf mutant in wheat variety K68

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, D [Banaras Hindu Univ. (India). Dept. of Genetics and Plant Breeding

    1977-04-01

    A semi-dwarf mutant (HUW-SDf 1) was induced from common wheat Var. K68 through the exposure of /sup 60/Co ..gamma..-rays at 15 kR. This mutant along with other induced mutants and control was assessed for yield components, yield and grain quality (M/sub 4/ generation); internode length reduction pattern and the yielding ability at three levels of nitrogen (M/sub 5/ generation). The mutant was significantly shorter in height and almost equal in tillers per plant and grains per spike to K68. However, it showed marked reduction in spike length and spikelets per spike. On the other hand, it possessed significantly higher (50.04 g) 1000-grain weight against control (41.15 g). The mutant gave 56.0% higher yield than the control. Grain quality studies indicated that the mutant possessed significantly higher (14.15%) total protein than K68. It was equally as good as K68 in lysine content. Pelshenke value (62.5 min) of the mutant indicated medium hard nature of gluten as compared to hard nature (198.0) of the control. The mutant showed 24.0% reduction in total culm length compared to K68. Reduction occurred due to maximum and almost equal reduction in 5th and 4th internodes (ca 34.0%) followed by 3rd, 2nd and 1st. The mutant showed similar yield and yield response to increasing nitrogen levels (80 to 160 kg per ha.) as for current commercial semi-dwarf varieties.

  14. Properties of the simian virus 40 (SV40) large T antigens encoded by SV40 mutants with deletions in gene A.

    Science.gov (United States)

    Cole, C N; Tornow, J; Clark, R; Tjian, R

    1986-01-01

    The biochemical properties of the large T antigens encoded by simian virus 40 (SV40) mutants with deletions at DdeI sites in the SV40 A gene were determined. Mutant large T antigens containing only the first 138 to 140 amino acids were unable to bind to the SV40 origin of DNA replication as were large T antigens containing at their COOH termini 96 or 97 amino acids encoded by the long open reading frame located between 0.22 and 0.165 map units (m.u.). All other mutant large T antigens were able to bind to the SV40 origin of replication. Mutants with in-phase deletions at 0.288 and 0.243 m.u. lacked ATPase activity, but ATPase activity was normal in mutants lacking origin-binding activity. The 627-amino acid large T antigen encoded by dlA2465, with a deletion at 0.219 m.u., was the smallest large T antigen displaying ATPase activity. Mutant large T antigens with the alternate 96- or 97-amino acid COOH terminus also lacked ATPase activity. All mutant large T antigens were found in the nuclei of infected cells; a small amount of large T with the alternate COOH terminus was also located in the cytoplasm. Mutant dlA2465 belonged to the same class of mutants as dlA2459. It was unable to form plaques on CV-1p cells at 37 or 32 degrees C but could form plaques on BSC-1 monolayers at 37 degrees C but not at 32 degrees C. It was positive for viral DNA replication and showed intracistronic complementation with any group A mutant whose large T antigen contained a normal carboxyl terminus. These findings and those of others suggest that both DNA binding and ATPase activity are required for the viral DNA replication function of large T antigen, that these two activities must be located on the same T antigen monomer, and that these two activities are performed by distinct domains of the polypeptide. These domains are distinct and separable from the domain affected by the mutation of dlA2465 and indicate that SV40 large T antigen is made up of at least three separate functional

  15. A critical role for very long-chain fatty acid elongases in oleic acid-mediated Saccharomyces cerevisiae cytotoxicity.

    Science.gov (United States)

    Wang, Qiao; Du, Xiuxiu; Ma, Ke; Shi, Ping; Liu, Wenbin; Sun, Jing; Peng, Min; Huang, Zhiwei

    2018-03-01

    Elongases FEN1/ELO2 and SUR4/ELO3 are important enzymes involved in the elongation of long-chain fatty acids (LCFAs) to very long-chain fatty acids (VLCFAs) in Saccharomyces cerevisiae. The molecular mechanism of the involvement of these elongases in lipotoxicity is unclear. In the present study, we investigated the role of VLCFA elongases in oleic acid-mediated yeast cytotoxicity. The spot test showed that yeast strains with the deletion of ELO2 or ELO3 were strikingly sensitive to oleic acid, while there was no change on the growth of strain with deleted ELO1 which was involved in the elongation of C 14 fatty acid (FA) to C 16 FA. By using GC-MS, the unsaturation index was increased in elo2△ and elo3△ mutants after treatment with oleic acid (OLA). However, the proportion of VLCFAs was increased in response to OLA in the wild-type strain. The growth inhibition of elo2△ and elo3△ could be partially rescued by two commonly used antioxidant agents N-acetyl cysteine (NAC) and Ascorbic acid (VC). The further study showed that exposure to excess OLA led to an increase in the levels of reactive oxygen species (ROS) and thiobarbituric acid reactive substances (TBARS), and a decline in the quantity of reduced glutathione (GSH) in both the wild type and mutant strains. However, the antioxidant enzyme activities of superoxide dismutase (SOD) and catalase (CAT) were increased in the wild type and elo1△ strains, while they were significantly decreased in the mutants of elo2△ and elo3△ after treated with excess OLA. Thus, oxidative damage mainly contributed to the cell death induced by OLA in ole2△ and ole3△. Taken together, although disruption of ELO2 or ELO3 did not affect the cellular lipid unsaturation, they altered the distribution and propotion of cellular VLCFAs, leading to the cell membrane impairment, which augmented the ability of OLA to permeabilize the plasma membrane. The data suggest that the very long-chain fatty acids elongases ELO2 and ELO3

  16. Prion Propagation in Cells Expressing PrP Glycosylation Mutants

    Science.gov (United States)

    Salamat, Muhammad K.; Dron, Michel; Chapuis, Jérôme; Langevin, Christelle; Laude, Hubert

    2011-01-01

    Infection by prions involves conversion of a host-encoded cell surface protein (PrPC) to a disease-related isoform (PrPSc). PrPC carries two glycosylation sites variably occupied by complex N-glycans, which have been suggested by previous studies to influence the susceptibility to these diseases and to determine characteristics of prion strains. We used the Rov cell system, which is susceptible to sheep prions, to generate a series of PrPC glycosylation mutants with mutations at one or both attachment sites. We examined their subcellular trafficking and ability to convert into PrPSc and to sustain stable prion propagation in the absence of wild-type PrP. The susceptibility to infection of mutants monoglycosylated at either site differed dramatically depending on the amino acid substitution. Aglycosylated double mutants showed overaccumulation in the Golgi compartment and failed to be infected. Introduction of an ectopic glycosylation site near the N terminus fully restored cell surface expression of PrP but not convertibility into PrPSc, while PrPC with three glycosylation sites conferred cell permissiveness to infection similarly to the wild type. In contrast, predominantly aglycosylated molecules with nonmutated N-glycosylation sequons, produced in cells expressing glycosylphosphatidylinositol-anchorless PrPC, were able to form infectious PrPSc. Together our findings suggest that glycosylation is important for efficient trafficking of anchored PrP to the cell surface and sustained prion propagation. However, properly trafficked glycosylation mutants were not necessarily prone to conversion, thus making it difficult in such studies to discern whether the amino acid changes or glycan chain removal most influences the permissiveness to prion infection. PMID:21248032

  17. Gamma-aminobutyric acid depletion affects stomata closure and drought tolerance of Arabidopsis thaliana.

    Science.gov (United States)

    Mekonnen, Dereje Worku; Flügge, Ulf-Ingo; Ludewig, Frank

    2016-04-01

    A rapid accumulation of γ-aminobutyric acid (GABA) during biotic and abiotic stresses is well documented. However, the specificity of the response and the primary role of GABA under such stress conditions are hardly understood. To address these questions, we investigated the response of the GABA-depleted gad1/2 mutant to drought stress. GABA is primarily synthesized from the decarboxylation of glutamate by glutamate decarboxylase (GAD) which exists in five copies in the genome of Arabidopsis thaliana. However, only GAD1 and GAD2 are abundantly expressed, and knockout of these two copies dramatically reduced the GABA content. Phenotypic analysis revealed a reduced shoot growth of the gad1/2 mutant. Furthermore, the gad1/2 mutant was wilted earlier than the wild type following a prolonged drought stress treatment. The early-wilting phenotype was due to an increase in stomata aperture and a defect in stomata closure. The increase in stomata aperture contributed to higher stomatal conductance. The drought oversensitive phenotype of the gad1/2 mutant was reversed by functional complementation that increases GABA level in leaves. The functionally complemented gad1/2 x pop2 triple mutant contained more GABA than the wild type. Our findings suggest that GABA accumulation during drought is a stress-specific response and its accumulation induces the regulation of stomatal opening thereby prevents loss of water. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  18. Isolation of reovirus T3D mutants capable of infecting human tumor cells independent of junction adhesion molecule-A.

    Directory of Open Access Journals (Sweden)

    Diana J M van den Wollenberg

    Full Text Available Mammalian Reovirus is a double-stranded RNA virus with a distinctive preference to replicate in and lyse transformed cells. On that account, Reovirus type 3 Dearing (T3D is clinically evaluated as oncolytic agent. The therapeutic efficacy of this approach depends in part on the accessibility of the reovirus receptor Junction Adhesion Molecule-A (JAM-A on the target cells. Here, we describe the isolation and characterization of reovirus T3D mutants that can infect human tumor cells independent of JAM-A. The JAM-A-independent (jin mutants were isolated on human U118MG glioblastoma cells, which do not express JAM-A. All jin mutants harbour mutations in the S1 segments close to the region that encodes the sialic acid-binding pocket in the shaft of the spike protein. In addition, two of the jin mutants encode spike proteins with a Q336R substitution in their head domain. The jin mutants can productively infect a wide range of cell lines that resist wt reovirus T3D infection, including chicken LMH cells, hamster CHO cells, murine endothelioma cells, human U2OS and STA-ET2.1 cells, but not primary human fibroblasts. The jin-mutants rely on the presence of sialic-acid residues on the cell surface for productive infection, as is evident from wheat germ agglutinin (WGA inhibition experiments, and from the jin-reovirus resistance of CHO-Lec2 cells, which have a deficiency of sialic-acids on their glycoproteins. The jin mutants may be useful as oncolytic agents for use in tumors in which JAM-A is absent or inaccessible.

  19. Characterisation of SalRAB a salicylic acid inducible positively regulated efflux system of Rhizobium leguminosarum bv viciae 3841.

    Directory of Open Access Journals (Sweden)

    Adrian J Tett

    Full Text Available Salicylic acid is an important signalling molecule in plant-microbe defence and symbiosis. We analysed the transcriptional responses of the nitrogen fixing plant symbiont, Rhizobium leguminosarum bv viciae 3841 to salicylic acid. Two MFS-type multicomponent efflux systems were induced in response to salicylic acid, rmrAB and the hitherto undescribed system salRAB. Based on sequence similarity salA and salB encode a membrane fusion and inner membrane protein respectively. salAB are positively regulated by the LysR regulator SalR. Disruption of salA significantly increased the sensitivity of the mutant to salicylic acid, while disruption of rmrA did not. A salA/rmrA double mutation did not have increased sensitivity relative to the salA mutant. Pea plants nodulated by salA or rmrA strains did not have altered nodule number or nitrogen fixation rates, consistent with weak expression of salA in the rhizosphere and in nodule bacteria. However, BLAST analysis revealed seventeen putative efflux systems in Rlv3841 and several of these were highly differentially expressed during rhizosphere colonisation, host infection and bacteroid differentiation. This suggests they have an integral role in symbiosis with host plants.

  20. A mutant of a mutant of a mutant of a ...: Irradiation of progressive radiation-induced mutants in a mutation-breeding programme with Chrysanthenum morifolium RAM

    International Nuclear Information System (INIS)

    Broertjes, C.; Koene, P.; Veen, J.W.H. van.

    1980-01-01

    Radiation-induced sports in Chrysanthemum morifolium RAM. have been reported for several years. It has become an everyday practice to produce flower-colour mutants from outstanding cross-breeding products, even before they are distributed for the commercial production of cut flowers. One of the most successful and recent examples is that of cv. Horim, of which hundreds of mutants were produced by successive use of radiation-induced mutants in the mutation-breeding programme. Over about 4 years a variety of flower-colour mutants was obtained, not only largely including the outstanding characteristics of the original cultivar but sometimes even with an appreciable improvement in quality and yield. It is expected that the latter types, the Miros group, will soon completely supersede the spontaneous or raditation-induced Horim sports and mutants and take over the leading position of the Horim group in the production of all-year-round (AYR) cut-flowers. (orig.)

  1. p21-ras effector domain mutants constructed by "cassette" mutagenesis

    DEFF Research Database (Denmark)

    Stone, J C; Vass, W C; Willumsen, B M

    1988-01-01

    A series of mutations encoding single-amino-acid substitutions within the v-rasH effector domain were constructed, and the ability of the mutants to induce focal transformation of NIH 3T3 cells was studied. The mutations, which spanned codons 32 to 40, were made by a "cassette" mutagenesis...

  2. Cauliflower mosaic virus protein P6 inhibits signaling responses to salicylic acid and regulates innate immunity.

    Directory of Open Access Journals (Sweden)

    Andrew J Love

    Full Text Available Cauliflower mosaic virus (CaMV encodes a multifunctional protein P6 that is required for translation of the 35S RNA and also acts as a suppressor of RNA silencing. Here we demonstrate that P6 additionally acts as a pathogenicity effector of an unique and novel type, modifying NPR1 (a key regulator of salicylic acid (SA- and jasmonic acid (JA-dependent signaling and inhibiting SA-dependent defence responses We find that that transgene-mediated expression of P6 in Arabidopsis and transient expression in Nicotiana benthamiana has profound effects on defence signaling, suppressing expression of representative SA-responsive genes and increasing expression of representative JA-responsive genes. Relative to wild-type Arabidopsis P6-expressing transgenics had greatly reduced expression of PR-1 following SA-treatment, infection by CaMV or inoculation with an avirulent bacterial pathogen Pseudomonas syringae pv tomato (Pst. Similarly transient expression in Nicotiana benthamiana of P6 (including a mutant form defective in translational transactivation activity suppressed PR-1a transcript accumulation in response to Agrobacterium infiltration and following SA-treatment. As well as suppressing the expression of representative SA-regulated genes, P6-transgenic Arabidopsis showed greatly enhanced susceptibility to both virulent and avirulent Pst (titres elevated 10 to 30-fold compared to non-transgenic controls but reduced susceptibility to the necrotrophic fungus Botrytis cinerea. Necrosis following SA-treatment or inoculation with avirulent Pst was reduced and delayed in P6-transgenics. NPR1 an important regulator of SA/JA crosstalk, was more highly expressed in the presence of P6 and introduction of the P6 transgene into a transgenic line expressing an NPR1:GFP fusion resulted in greatly increased fluorescence in nuclei even in the absence of SA. Thus in the presence of P6 an inactive form of NPR1 is mislocalized in the nucleus even in uninduced plants

  3. Optimization of flask culture medium and conditions for hyaluronic acid production by a streptococcus equisimilis mutant nc2168

    Directory of Open Access Journals (Sweden)

    Yong-Hao Chen

    2012-12-01

    Full Text Available A mutant designated NC2168, which was selected from wild-type Streptococcus equisimilis CVCC55116by ultraviolet ray combined with60Co-γ ray treatment and does not produce streptolysin, was employed to produce hyaluronic acid (HA. In order to increase the output of HA in a flask, the culture medium and conditions for NC2168 were optimized in this study. The influence of culture medium ingredients including carbon sources, nitrogen sources and metal ions on HA production was evaluated using factional factorial design. The mathematical model, which represented the effect of each medium component and their interaction on the yield of HA, was established by the quadratic rotary combination design and response surface method. The model estimated that, a maximal yield of HA could be obtained when the concentrations of yeast extract, peptone, glucose, and MgSO4 were set at 3 g/100 mL, 2 g/100 mL, 0.5 g/100 mL and 0.15 g/100 mL, respectively. Compared with the values obtained by other runs in the experimental design, the optimized medium resulted in a remarkable increase in the output of HA and the maximum of the predicted HA production was 174.76 mg/L. The model developed was accurate and reliable for predicting the production of HA by NC2168.Cultivation conditions were optimized by an orthogonal experimental design and the optimal conditions were as follows: temperature 33ºC, pH 7.8, agitation speed 200 rpm, medium volume 20 mL.

  4. Isolation of a novel mutant gene for soil-surface rooting in rice (Oryza sativa L.).

    Science.gov (United States)

    Hanzawa, Eiko; Sasaki, Kazuhiro; Nagai, Shinsei; Obara, Mitsuhiro; Fukuta, Yoshimichi; Uga, Yusaku; Miyao, Akio; Hirochika, Hirohiko; Higashitani, Atsushi; Maekawa, Masahiko; Sato, Tadashi

    2013-11-20

    Root system architecture is an important trait affecting the uptake of nutrients and water by crops. Shallower root systems preferentially take up nutrients from the topsoil and help avoid unfavorable environments in deeper soil layers. We have found a soil-surface rooting mutant from an M2 population that was regenerated from seed calli of a japonica rice cultivar, Nipponbare. In this study, we examined the genetic and physiological characteristics of this mutant. The primary roots of the mutant showed no gravitropic response from the seedling stage on, whereas the gravitropic response of the shoots was normal. Segregation analyses by using an F2 population derived from a cross between the soil-surface rooting mutant and wild-type Nipponbare indicated that the trait was controlled by a single recessive gene, designated as sor1. Fine mapping by using an F2 population derived from a cross between the mutant and an indica rice cultivar, Kasalath, revealed that sor1 was located within a 136-kb region between the simple sequence repeat markers RM16254 and 2935-6 on the terminal region of the short arm of chromosome 4, where 13 putative open reading frames (ORFs) were found. We sequenced these ORFs and detected a 33-bp deletion in one of them, Os04g0101800. Transgenic plants of the mutant transformed with the genomic fragment carrying the Os04g0101800 sequence from Nipponbare showed normal gravitropic responses and no soil-surface rooting. These results suggest that sor1, a rice mutant causing soil-surface rooting and altered root gravitropic response, is allelic to Os04g0101800, and that a 33-bp deletion in the coding region of this gene causes the mutant phenotypes.

  5. Glycoprotein cytoplasmic domain sequences required for rescue of a vesicular stomatitis virus glycoprotein mutant

    International Nuclear Information System (INIS)

    Whitt, M.A.; Chong, L.; Rose, J.K.

    1989-01-01

    The authors have used transient expression of the wild-type vesicular stomatitis virus (VSV) glycoprotein (G protein) from cloned cDNA to rescue a temperature-sensitive G protein mutant of VSV in cells at the nonpermissive temperature. Using cDNAs encoding G proteins with deletions in the normal 29-amino-acid cytoplasmic domain, they determined that the presence of either the membrane-proximal 9 amino acids or the membrane-distal 12 amino acids was sufficient for rescue of the temperature-sensitive mutant. G proteins with cytoplasmic domains derived from other cellular or viral G proteins did not rescue the mutant, nor did G proteins with one or three amino acids of the normal cytoplasmic domain. Rescue correlated directly with the ability of the G proteins to be incorporated into virus particles. This was shown by analysis of radiolabeled particles separated on sucrose gradients as well as by electron microscopy of rescued virus after immunogold labeling. Quantitation of surface expression showed that all of the mutated G proteins were expressed less efficiently on the cell surface than was wild-type G protein. However, they were able to correct for differences in rescue efficiency resulting from differences in the level of surface expression by reducing wild-type G protein expression to levels equivalent to those observed for the mutated G proteins. The results provide evidence that at least a portion of the cytoplasmic domain is required for efficient assembly of the VSV G protein into virions during virus budding

  6. Physiological and fermentation properties of Bacillus coagulans and a mutant lacking fermentative lactate dehydrogenase activity.

    Science.gov (United States)

    Su, Yue; Rhee, Mun Su; Ingram, Lonnie O; Shanmugam, K T

    2011-03-01

    Bacillus coagulans, a sporogenic lactic acid bacterium, grows optimally at 50-55 °C and produces lactic acid as the primary fermentation product from both hexoses and pentoses. The amount of fungal cellulases required for simultaneous saccharification and fermentation (SSF) at 55 °C was previously reported to be three to four times lower than for SSF at the optimum growth temperature for Saccharomyces cerevisiae of 35 °C. An ethanologenic B. coagulans is expected to lower the cellulase loading and production cost of cellulosic ethanol due to SSF at 55 °C. As a first step towards developing B. coagulans as an ethanologenic microbial biocatalyst, activity of the primary fermentation enzyme L-lactate dehydrogenase was removed by mutation (strain Suy27). Strain Suy27 produced ethanol as the main fermentation product from glucose during growth at pH 7.0 (0.33 g ethanol per g glucose fermented). Pyruvate dehydrogenase (PDH) and alcohol dehydrogenase (ADH) acting in series contributed to about 55% of the ethanol produced by this mutant while pyruvate formate lyase and ADH were responsible for the remainder. Due to the absence of PDH activity in B. coagulans during fermentative growth at pH 5.0, the l-ldh mutant failed to grow anaerobically at pH 5.0. Strain Suy27-13, a derivative of the l-ldh mutant strain Suy27, that produced PDH activity during anaerobic growth at pH 5.0 grew at this pH and also produced ethanol as the fermentation product (0.39 g per g glucose). These results show that construction of an ethanologenic B. coagulans requires optimal expression of PDH activity in addition to the removal of the LDH activity to support growth and ethanol production.

  7. Characterization of D-myo-inositol 3-phosphate synthase gene expression in two soybean low phytate mutants

    International Nuclear Information System (INIS)

    Yuan Fengjie; Dong Dekun; Li Baiquan; Yu Xiaomin; Fu Xujun; Zhu Danhua; Zhu Shenlong; Yang Qinghua

    2013-01-01

    1D-myo-inositol 3-phosphate synthase (MIPS) gene plays a significant role in phytic acid biosynthesis. In this study, we used two low phytic acid mutants Gm-lpa-TW-1, Gm-lpa-ZC-2 and their respective wild type parents Taiwan75 and Zhechun No.3 to analyze the expression pattern and characterization of MIPS1 gene. The results showed that there was a common expression pattern of MIPS1 in soybean developing seeds. Expression was weak as detected by RT-PCR in initial stage, increased in the following stages, and the peak expression was appeared in 22 day after flowering (DAF). The expression of MIPS1 gene of non-seed tissues in mutant Gm-lpa-TW-1 and its wildtype Taiwan75 was very weak. In the developing seeds, the MIPS1 expression by qRT-PCR revealed a significant reduction in 22 DAF in mutant Gm-lpa-TW-1 as compared with the wildtype. Similarly, the expression of MIPS1 gene in non-seed tissue of Zhenchun No.3 and Gm-lpa-ZC-2 was very weak. However, stronger expression in developing seeds of the mutant Gm-lpa-ZC-2 than Zhechun No.3 was found. We concluded that the MIPS1 gene expression in the developing seed exhibited an up-regulation pattern in mutant Gm-lpa-ZC-2, but a down-regulation pattern in the mutant Gm-lpa-TW-1. (authors)

  8. EDR2 negatively regulates salicylic acid-based defenses and cell death during powdery mildew infections of Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Nishimura Marc

    2007-07-01

    Full Text Available Abstract Background The hypersensitive necrosis response (HR of resistant plants to avirulent pathogens is a form of programmed cell death in which the plant sacrifices a few cells under attack, restricting pathogen growth into adjacent healthy tissues. In spite of the importance of this defense response, relatively little is known about the plant components that execute the cell death program or about its regulation in response to pathogen attack. Results We isolated the edr2-6 mutant, an allele of the previously described edr2 mutants. We found that edr2-6 exhibited an exaggerated chlorosis and necrosis response to attack by three pathogens, two powdery mildew and one downy mildew species, but not in response to abiotic stresses or attack by the bacterial leaf speck pathogen. The chlorosis and necrosis did not spread beyond inoculated sites suggesting that EDR2 limits the initiation of cell death rather than its spread. The pathogen-induced chlorosis and necrosis of edr2-6 was correlated with a stimulation of the salicylic acid defense pathway and was suppressed in mutants deficient in salicylic acid signaling. EDR2 encodes a novel protein with a pleckstrin homology and a StAR transfer (START domain as well as a plant-specific domain of unknown function, DUF1336. The pleckstrin homology domain binds to phosphatidylinositol-4-phosphate in vitro and an EDR2:HA:GFP protein localizes to endoplasmic reticulum, plasma membrane and endosomes. Conclusion EDR2 acts as a negative regulator of cell death, specifically the cell death elicited by pathogen attack and mediated by the salicylic acid defense pathway. Phosphatidylinositol-4-phosphate may have a role in limiting cell death via its effect on EDR2. This role in cell death may be indirect, by helping to target EDR2 to the appropriate membrane, or it may play a more direct role.

  9. Structural characterization of V57D and V57P mutants of human cystatin C, an amyloidogenic protein

    Energy Technology Data Exchange (ETDEWEB)

    Orlikowska, Marta; Szymańska, Aneta [University of Gdansk, Sobieskiego 18/19, 80-952 Gdansk (Poland); Borek, Dominika; Otwinowski, Zbyszek [University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, TX 75390-8816 (United States); Skowron, Piotr; Jankowska, Elżbieta, E-mail: elaj@chem.univ.gda.pl [University of Gdansk, Sobieskiego 18/19, 80-952 Gdansk (Poland)

    2013-04-01

    Val57 point mutants of human cystatin C, which were designed to assess the influence of changes in the properties of the L1 loop on the dimerization propensity, were structurally characterized. Wild-type human cystatin C (hCC wt) is a low-molecular-mass protein (120 amino-acid residues, 13 343 Da) that is found in all nucleated cells. Physiologically, it functions as a potent regulator of cysteine protease activity. While the biologically active hCC wt is a monomeric protein, all crystallization efforts to date have resulted in a three-dimensional domain-swapped dimeric structure. In the recently published structure of a mutated hCC, the monomeric fold was preserved by a stabilization of the conformationally constrained loop L1 caused by a single amino-acid substitution: Val57Asn. Additional hCC mutants were obtained in order to elucidate the relationship between the stability of the L1 loop and the propensity of human cystatin C to dimerize. In one mutant Val57 was substituted by an aspartic acid residue, which is favoured in β-turns, and in the second mutant proline, a residue known for broadening turns, was substituted for the same Val57. Here, 2.26 and 3.0 Å resolution crystal structures of the V57D andV57P mutants of hCC are reported and their dimeric architecture is discussed in terms of the stabilization and destabilization effects of the introduced mutations.

  10. Genetic analysis and molecular detection of the corn endosperm mutants induced by space flight

    International Nuclear Information System (INIS)

    Zhang Caibo; Zhou Yuanyuan; Wang Hanyu; Wang Hongwei; Wang Shengqing; Rong Tingzhao; Cao Moju

    2013-01-01

    In this study, two maize inbred lines 08-641 and 18-599 were carried into cosmic space by recoverable satellite 'Shijian 8', grain shrunken transparently and opaquely mutants were selected as experimental materials and their soluble sugar content in kernel were measured by annthrone colorimetry. The content of soluble sugar in mutant st1 kernels began to rise in 10 days after pollination, to reach the peak in 25 days and significantly higher than the contrast 08-641, while in mutant sol kernels it began to rise in 10 days after pollination, to reach the peak in 20 days and significantly higher than the contrast 18-599. The results of genetic analysis and allelism test showed that the trait in both mutants was all controlled by a single recessive gene, the mutant st1 was allelic to the su1 and the mutant sol was allelic to the sh2. DNA sequence alignment found 2 single-base mutations in 2 and 13 exon of su1 gene in the mutant st1 and 3 single-base mutations in 2, 5 and 16 exon of sh2 gene in mutant so1 leading to the change in amino acid sequences. So it is inferred that starch biosynthesis in the mutants may be blocked by these mutations, which lead to the increase of soluble sugar content in kernel. (authors)

  11. [Genetic control of the sensitivity of Aspergillus nidulans to mutagenic factors. VII. Inheritance of cross-sensitivity to different mutagenic factors by uvs-mutants].

    Science.gov (United States)

    Evseeva, G V; Kameneva, S V

    1977-01-01

    To study the inheritance of the sensitivity to UV, X-rays, methylmethanesulphonate (MMS), nitrosoguanidine (NG) and nitrous acid (NA) in five uvs mutants of Aspergillus nidulans, having multiple sensitivity to these factors, the sensitivity of recombinants obtained from crossing uvs mutants with uvs+ strain, resistant to all the factors analysed, and uvs leads to uvs+ revertants is investigated. Four uvs mutants (15, 17, 19 and 26) are found to have a nomogenic control of sensitivity to different mutagens. In one mutant (uvs11) the sensitivity to five factors is controlled by two non-linked mutations, one of them determining the sensitivity to UV, NG, NA, and the other--to X-rays and MMC. Phenotypic manifestations of uvs mutations is modified by cell genotype, both chromosomal and cytoplasmic factors being responsible for the modification. Phenotypic modification of uvs mutation results in the change to some (but not to all) mutagenic factors. It suggests, that not the product of uvs gene, but some other components of the reparation complex are modified. Otherwise, reparation of different DNA damages can be carried out by a single enzyme acting in different reparation complexes.

  12. Preliminary Study of the Characteristics of Several Glossy Cabbage (Brassica oleracea var. capitata L. Mutants

    Directory of Open Access Journals (Sweden)

    Tang Jun

    2015-09-01

    Full Text Available To determine the characteristics and potential practical applications of glossy cabbage (Brassica oleracea var. capitata L. mutants, five different glossy mutants were studied. The amount of epicuticular wax covering the mutant leaves was only approximately 30% that of the wild-type (WT leaves. The wax crystals of WT plants were columnar and linear, while they were granular and rod-shaped in the mutants. Additionally, in WT cabbage, the primary wax components were alkanes, alcohols, fatty acids, ketones, and aldehydes. There was a significant decrease in the abundance of alkanes and ketones in the wax of the mutants. The glossy-green trait of the mutants may be the result of an inhibited alkane-forming pathway. Higher rates of chlorophyll leaching and water loss demonstrate that the mutant leaves were more permeable and sensitive to drought stress than the WT leaves. Growth curve results indicated that the growth rate of mutant-1 and mutant-3 was slower than that of the corresponding WT cabbage, resulting in shorter plants. However, the growth rate of mutant-2 was not influenced by the lack of coating wax. An investigation of the agronomic traits and heterosis of the glossy cabbage mutants indicated that all five mutants had glossy-green leaves, which was a favorable characteristic. The F1 plants derived from crosses involving mutant-2 exhibited obvious heterosis, suggesting the observed glossy-green trait is controlled by a dominant gene. Therefore, mutant-2 may be useful as a source of genetic material for future cabbage breeding experiments.

  13. Lipidomic Profiling of Lung Pleural Effusion Identifies Unique Metabotype for EGFR Mutants in Non-Small Cell Lung Cancer.

    Science.gov (United States)

    Ho, Ying Swan; Yip, Lian Yee; Basri, Nurhidayah; Chong, Vivian Su Hui; Teo, Chin Chye; Tan, Eddy; Lim, Kah Ling; Tan, Gek San; Yang, Xulei; Yeo, Si Yong; Koh, Mariko Si Yue; Devanand, Anantham; Takano, Angela; Tan, Eng Huat; Tan, Daniel Shao Weng; Lim, Tony Kiat Hon

    2016-10-14

    Cytology and histology forms the cornerstone for the diagnosis of non-small cell lung cancer (NSCLC) but obtaining sufficient tumour cells or tissue biopsies for these tests remains a challenge. We investigate the lipidome of lung pleural effusion (PE) for unique metabolic signatures to discriminate benign versus malignant PE and EGFR versus non-EGFR malignant subgroups to identify novel diagnostic markers that is independent of tumour cell availability. Using liquid chromatography mass spectrometry, we profiled the lipidomes of the PE of 30 benign and 41 malignant cases with or without EGFR mutation. Unsupervised principal component analysis revealed distinctive differences between the lipidomes of benign and malignant PE as well as between EGFR mutants and non-EGFR mutants. Docosapentaenoic acid and Docosahexaenoic acid gave superior sensitivity and specificity for detecting NSCLC when used singly. Additionally, several 20- and 22- carbon polyunsaturated fatty acids and phospholipid species were significantly elevated in the EGFR mutants compared to non-EGFR mutants. A 7-lipid panel showed great promise in the stratification of EGFR from non-EGFR malignant PE. Our data revealed novel lipid candidate markers in the non-cellular fraction of PE that holds potential to aid the diagnosis of benign, EGFR mutation positive and negative NSCLC.

  14. Frictional response of fatty acids on steel.

    Science.gov (United States)

    Sahoo, Rashmi R; Biswas, S K

    2009-05-15

    Self-assembled monolayers of fatty acids were formed on stainless steel by room-temperature solution deposition. The acids are covalently bound to the surface as carboxylate in a bidentate manner. To explore the effect of saturation in the carbon backbone on friction in sliding tribology, we study the response of saturated stearic acid (SA) and unsaturated linoleic acid (LA) as self-assembled monolayers using lateral force microscopy and nanotribometry and when the molecules are dispersed in hexadecane, using pin-on-disc tribometry. Over a very wide range (10 MPa-2.5 GPa) of contact pressures it is consistently demonstrated that the unsaturated linoleic acid molecules yield friction which is significantly lower than that of the saturated stearic acid. It is argued, using density functional theory predictions and XPS of slid track, that when the molecular backbone of unsaturated fatty acids are tilted and pressed strongly by a probe, in tribological contact, the high charge density of the double bond region of the backbone allows coupling with the steel substrate. The interaction yields a low friction carboxylate soap film on the substrate. The saturated fatty acid does not show this effect.

  15. Chemotaxis to cyclic AMP and folic acid is mediated by different G proteins in Dictyostelium discoideum

    NARCIS (Netherlands)

    Kesbeke, Fanja; Haastert, Peter J.M. van; Wit, René J.W. de; Snaar-Jagalska, B. Ewa

    1990-01-01

    Mutant Frigid A (fgdA) of Dictyostelium discoideum is defective in a functional Gα2 subunit of a G protein and is characterized by a complete blockade of the cyclic AMP-mediated sensory transduction steps, including cyclic AMP relay, chemotaxis and the cyclic GMP response. Folic acid-mediated

  16. The Barley Magnesium Chelatase 150-kD Subunit Is Not an Abscisic Acid Receptor1[OA

    Science.gov (United States)

    Müller, André H.; Hansson, Mats

    2009-01-01

    Magnesium chelatase is the first unique enzyme of the chlorophyll biosynthetic pathway. It is composed of three gene products of which the largest is 150 kD. This protein was recently identified as an abscisic acid receptor in Arabidopsis (Arabidopsis thaliana). We have evaluated whether the barley (Hordeum vulgare) magnesium chelatase large subunit, XanF, could be a receptor for the phytohormone. The study involved analysis of recombinant magnesium chelatase protein as well as several induced chlorophyll-deficient magnesium chelatase mutants with defects identified at the gene and protein levels. Abscisic acid had no effect on magnesium chelatase activity and binding to the barley 150-kD protein could not be shown. Magnesium chelatase mutants showed a wild-type response in respect to postgermination growth and stomatal aperture. Our results question the function of the large magnesium chelatase subunit as an abscisic acid receptor. PMID:19176716

  17. Characterization and fine mapping of a light-dependent leaf lesion mimic mutant 1 in rice.

    Science.gov (United States)

    Wang, Jing; Ye, Bangquan; Yin, Junjie; Yuan, Can; Zhou, Xiaogang; Li, Weitao; He, Min; Wang, Jichun; Chen, Weilan; Qin, Peng; Ma, Bintian; Wang, Yuping; Li, Shigui; Chen, Xuewei

    2015-12-01

    Plants that spontaneously produce lesion mimics or spots, without any signs of obvious adversity, such as pesticide and mechanical damage, or pathogen infection, are so-called lesion mimic mutants (lmms). In rice, many lmms exhibit enhanced resistance to pathogens, which provides a unique opportunity to uncover the molecular mechanism underlying lmms. We isolated a rice light-dependent leaf lesion mimic mutant 1 (llm1). Lesion spots appeared in the leaves of the llm1 mutant at the tillering stage. Furthermore, the mutant llm1 had similar agronomic traits to wild type rice. Trypan blue and diamiobenzidine staining analyses revealed that the lesion spot formation on the llm1 mutant was due to programmed cell death and reactive oxygen species. The chloroplasts were severely damaged in the llm1 mutant, suggesting that chloroplast damage was associated with the formation of lesion spots in llm1. More importantly, llm1 exhibited enhanced resistance to bacterial blight pathogens within increased expression of pathogenesis related genes (PRs). Using a map-based cloning approach, we delimited the LLM1 locus to a 121-kb interval between two simple sequence repeat markers, RM17470 and RM17473, on chromosome 4. We sequenced the candidate genes on the interval and found that a base mutation had substituted adenine phosphate for thymine in the last exon of LOC_Os04g52130, which led to an amino acid change (Asp(388) to Val) in the llm1 mutant. Our investigation showed that the putative coproporphyrinogen III oxidase (CPOX) encoded by LOC_Os04g52130 was produced by LLM1 and that amino acid Asp(388) was essential for CPOX function. Our study provides the basis for further investigations into the mechanism underlying lesion mimic initiation associated with LLM1. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  18. Screening of a lactobacillus plantarum mutant with high cla productivity induced by n+ implantation

    International Nuclear Information System (INIS)

    Li Shurong; Meng Xianjun; Zhang Tao; Zhao Hongwei; Lu Jiaping; Zhao Yun; Gao Yanhong; Li Qingpeng

    2009-01-01

    The initial lactic acid bacteria strain A6-1 was treated by N + ions implantation of 50 keV with doses of 1 x 10 13 , 3 x 10 13 , 5 x 10 13 , 8 x 10 13 , 10 x 10 13 , 30 x 10 13 , 50 x 10 13 , 80 x 10 13 , and 100 x 10 13 ions/cm 2 . The survival curve showed a saddle model, and the high survival rate was 20% ∼ 35% from the treatments of 30 x 10 13 ions/cm 2 and 50 x 10 13 ions/cm 2 implantation. Considering the survival rate, positive mutation and range of mutation rate, N + ions implantation of 30 x 10 13 ions/cm 2 was recommended for mutation breeding of lactic acid bacteria. Selected mutants with high ability of producing CLA after fermentation. Generic stable was observed until 8 generations of F mutant, and average yield of CLA was 162.5 μg/ml, which was 69.87% higher than the original stain. F mutant was named A6-1F. (authors)

  19. Jasmonic acid is a key regulator of spider mite-induced volatile terpenoid and methyl salicylate emission in tomato

    NARCIS (Netherlands)

    Ament, K.; Kant, M.R.; Sabelis, M.W.; Haring, M.A.; Schuurink, R.C.

    2004-01-01

    The tomato (Lycopersicon esculentum) mutant def-1, which is deficient in induced jasmonic acid (JA) accumulation upon wounding or herbivory, was used to study the role of JA in the direct and indirect defense responses to phytophagous mites (Tetranychus urticae). In contrast to earlier reports,

  20. Purification and characterization of mutant miniPlasmin for thrombolytic therapy

    Directory of Open Access Journals (Sweden)

    Lin Xiaotao

    2013-01-01

    Full Text Available Abstract Background Previous animal studies by us and others have indicated that catheter-administered plasmin or its des-kringle derivatives may be more appropriate alternatives to plasminogen activators for treating thrombolytic diseases, since it has a very short serum half-life and therefore does not result in hemorrhaging. We have previously produced recombinant miniPlasmin (mPlasmin that was proven suitable for treating peripheral arterial occlusion in animal models. However, our previous results showed that non-specific cleavage at position K698 of mPlasmin during activation hindered the further development of this promising therapeutic candidate. In order to minimize or eliminate the non-specific cleavage problem, we performed saturation mutagenesis at the K698 position to develop a mutant form of mPlasmin for thrombolytic therapy. Methods We changed K698 to 16 other amino acids, with preferred E. coli codons. Each of these mutants were expressed in E. coli as inclusion bodies and then refolded, purified, and subsequently characterized by detailed kinetic assays/experiments/studies which identified highly active mutants devoid of non-specific cleavage. Results Activation studies indicated that at those conditions in which the wild type enzyme is cut at the non-specific position K698, the active mutants can be activated without being cleaved at this position. Conclusions From the above results, we selected two mutants, K698Q and K698N, as our lead candidates for further thrombolytic drug developments. The selected mutants are potentially better therapeutic candidates for thrombolytic therapy.

  1. Inflammatory response of TLR4 deficient spleen macrophages (CRL 2471) to Brucella abortus S19 and an isogenic ΔmglA deletion mutant.

    Science.gov (United States)

    Jacob, Jens; Makou, Patricia; Finke, Antje; Mielke, Martin

    2016-05-01

    Brucellosis is a worldwide distributed zoonosis caused by members of the genus Brucella. One of them, Brucella abortus, is the etiological agent of bovine brucellosis. With the attenuated strain B. abortus S19 a vaccine is available. However, both, virulence (safety) and the ability to induce a protective B and T cell response (efficacy) have to be tested in suitable assays before successful use in the field. For this purpose, several macrophage cell lines of various origins have been used while splenic macrophages are the preferred host cells in vivo. We here characterized the in vitro response of the murine splenic macrophage cell line CRL 2471(I-13.35) to B. abortus. This cell line still depends on the presence of colony-stimulating factor 1 (CSF1) and is derived from LPS resistant (TLR4 deficient) C3H/HeJ mice. For infection the vaccine strain B. abortus S19A as well as the formerly described isogenic deletion mutant B. abortus S19A ΔmglA 3.14 were used. While numbers of viable bacteria did not differ significantly between the vaccine strain and the deletion mutant at 6h post infection, a higher bacterial load was measured in case of the mutant at 24h and 48h after infection. This was also true, when IFNγ was used for macrophage activation. A comprehensive gene expression profile of macrophages was analysed 6 and 24h after infection by means of an RT-PCR based gene expression array. The mutant strain B. abortus S19A ΔmglA 3.14 elicited a stronger cellular response of the splenic macrophages as compared to the parental vaccine strain. This was most prominent for the pro-inflammatory cytokines IL-1α, IL-1β, TNF-α and IL6 as well as for the chemokine ligands CXCL1, CXCL2, CXCL10, CCL2, CCL5, CCL7, CCL17 and the co-stimulatory molecules CD40 and ICAM1. While these differences were also present in IFNγ-stimulated macrophages, an addition of IFNγ after infection not only resulted in a dramatic increase of the translation of the afore mentioned genes but also

  2. Conjugated fatty acid synthesis: residues 111 and 115 influence product partitioning of Momordica charantia conjugase.

    Science.gov (United States)

    Rawat, Richa; Yu, Xiao-Hong; Sweet, Marie; Shanklin, John

    2012-05-11

    Conjugated linolenic acids (CLNs), 18:3 Δ(9,11,13), lack the methylene groups found between the double bonds of linolenic acid (18:3 Δ(9,12,15)). CLNs are produced by conjugase enzymes that are homologs of the oleate desaturases FAD2. The goal of this study was to map the domain(s) within the Momordica charantia conjugase (FADX) responsible for CLN formation. To achieve this, a series of Momordica FADX-Arabidopsis FAD2 chimeras were expressed in the Arabidopsis fad3fae1 mutant, and the transformed seeds were analyzed for the accumulation of CLN. These experiments identified helix 2 and the first histidine box as a determinant of conjugase product partitioning into punicic acid (18:3 Δ(9cis,11trans,13cis)) or α-eleostearic acid (18:3 Δ(9cis,11trans,13trans)). This was confirmed by analysis of a FADX mutant containing six substitutions in which the sequence of helix 2 and first histidine box was converted to that of FAD2. Each of the six FAD2 substitutions was individually converted back to the FADX equivalent identifying residues 111 and 115, adjacent to the first histidine box, as key determinants of conjugase product partitioning. Additionally, expression of FADX G111V and FADX G111V/D115E resulted in an approximate doubling of eleostearic acid accumulation to 20.4% and 21.2%, respectively, compared with 9.9% upon expression of the native Momordica FADX. Like the Momordica conjugase, FADX G111V and FADX D115E produced predominantly α-eleostearic acid and little punicic acid, but the FADX G111V/D115E double mutant produced approximately equal amounts of α-eleostearic acid and its isomer, punicic acid, implicating an interactive effect of residues 111 and 115 in punicic acid formation.

  3. Responsive Boronic Acid-Decorated (Copolymers: From Glucose Sensors to Autonomous Drug Delivery

    Directory of Open Access Journals (Sweden)

    Gertjan Vancoillie

    2016-10-01

    Full Text Available Boronic acid-containing (copolymers have fascinated researchers for decades, garnering attention for their unique responsiveness toward 1,2- and 1,3-diols, including saccharides and nucleotides. The applications of materials that exert this property are manifold including sensing, but also self-regulated drug delivery systems through responsive membranes or micelles. In this review, some of the main applications of boronic acid containing (copolymers are discussed focusing on the role of the boronic acid group in the response mechanism. We hope that this summary, which highlights the importance and potential of boronic acid-decorated polymeric materials, will inspire further research within this interesting field of responsive polymers and polymeric materials.

  4. Isolation and molecular characterization of a urease-negative Actinobacillus pleuropneumoniae mutant.

    Science.gov (United States)

    Ito, Hiroya; Takahashi, Sayaka; Asai, Tetsuo; Tamura, Yutaka; Yamamoto, Koshi

    2018-01-01

    An atypical urease-negative mutant of Actinobacillus pleuropneumoniae serovar 2 was isolated in Japan. Nucleotide sequence analysis of the urease gene cluster revealed that the insertion of a short DNA sequence into the cbiM gene was responsible for the urease-negative activity of the mutant. Veterinary diagnostic laboratories should be watchful for the presence of aberrant urease-negative A. pleuropneumoniae isolates.

  5. Regulation of phenylacetic acid uptake is sigma54 dependent in Pseudomonas putida CA-3.

    LENUS (Irish Health Repository)

    O' Leary, Niall D

    2011-10-13

    Abstract Background Styrene is a toxic and potentially carcinogenic alkenylbenzene used extensively in the polymer processing industry. Significant quantities of contaminated liquid waste are generated annually as a consequence. However, styrene is not a true xenobiotic and microbial pathways for its aerobic assimilation, via an intermediate, phenylacetic acid, have been identified in a diverse range of environmental isolates. The potential for microbial bioremediation of styrene waste has received considerable research attention over the last number of years. As a result the structure, organisation and encoded function of the genes responsible for styrene and phenylacetic acid sensing, uptake and catabolism have been elucidated. However, a limited understanding persists in relation to host specific regulatory molecules which may impart additional control over these pathways. In this study the styrene degrader Pseudomonas putida CA-3 was subjected to random mini-Tn5 mutagenesis and mutants screened for altered styrene\\/phenylacetic acid utilisation profiles potentially linked to non-catabolon encoded regulatory influences. Results One mutant, D7, capable of growth on styrene, but not on phenylacetic acid, harboured a Tn5 insertion in the rpoN gene encoding σ54. Complementation of the D7 mutant with the wild type rpoN gene restored the ability of this strain to utilise phenylacetic acid as a sole carbon source. Subsequent RT-PCR analyses revealed that a phenylacetate permease, PaaL, was expressed in wild type P. putida CA-3 cells utilising styrene or phenylacetic acid, but could not be detected in the disrupted D7 mutant. Expression of plasmid borne paaL in mutant D7 was found to fully restore the phenylacetic acid utilisation capacity of the strain to wild type levels. Bioinformatic analysis of the paaL promoter from P. putida CA-3 revealed two σ54 consensus binding sites in a non-archetypal configuration, with the transcriptional start site being resolved by

  6. Dose-response functions for effects of acidic precipitation on vegetation

    Energy Technology Data Exchange (ETDEWEB)

    Jacobson, J S; Troiano, J J

    1983-01-01

    Research on the effect of sulfuric and nitric acids, as well as other substances, in rain on plant growth has focused on quantifying the relationship between doses of acids in precipitation and plant response. After eight years, there has been no direct demonstration of harmful effects to plants by ambient acidic rain in North America, and there remains considerable uncertainty about the potential risk to cultivated and native plants. Current efforts to describe the relationships between dose of acidity and effects on plants need better experimental approaches if the results are to be more relevant to actual field situations. Mechanistic models that describe the physiological and biochemical basis for effects of acidic rain on plants will be needed to provide confidence in the predictions of plant response. 34 references, 1 figure.

  7. Bioactive compounds of fourth generation gamma-irradiated Typhoniumflagelliforme Lodd. mutants based on gas chromatography-mass spectrometry

    Science.gov (United States)

    Sianipar, N. F.; Purnamaningsih, R.; Rosaria

    2016-08-01

    Rodent tuber (Typhonium flagelliforme Lodd.) is an Indonesian anticancer medicinal plant. The natural genetic diversity of rodent tuber is low due to vegetative propagation. Plant's genetic diversity has to be increased for obtaining clones which contain a high amount of anticancer compounds. In vitro calli were irradiated with 6 Gy of gamma ray to produce in vitro mutant plantlets. Mutant plantlets were acclimated and propagated in a greenhouse. This research was aimed to identify the chemical compounds in the leaves and tubers ofthe fourth generation of rodent tuber's vegetative mutant clones (MV4) and control plantsby using GC- MS method. Leaves and tubers of MV4 each contained 2 and 5 anticancer compounds which quantities were higher compared to control plants. MV4 leaves contained 5 new anticancer compounds while its tubers contained 3 new anticancer compounds which were not found in control. The new anticancer compounds in leaves were hexadecanoic acid, stigmast-5-en-3-ol, ergost-5-en-3-ol, farnesol isomer a, and oleic acid while the new anticancer compounds in tubers were alpha tocopherol, ergost-5-en-3-ol, and beta-elemene. Rodent tuber mutant clones are very potential to be developed into anticancer drugs.

  8. PAMP-induced defense responses in potato require both salicylic acid and jasmonic acid.

    Science.gov (United States)

    Halim, Vincentius A; Altmann, Simone; Ellinger, Dorothea; Eschen-Lippold, Lennart; Miersch, Otto; Scheel, Dierk; Rosahl, Sabine

    2009-01-01

    To elucidate the molecular mechanisms underlying pathogen-associated molecular pattern (PAMP)-induced defense responses in potato (Solanum tuberosum), the role of the signaling compounds salicylic acid (SA) and jasmonic acid (JA) was analyzed. Pep-13, a PAMP from Phytophthora, induces the accumulation of SA, JA and hydrogen peroxide, as well as the activation of defense genes and hypersensitive-like cell death. We have previously shown that SA is required for Pep-13-induced defense responses. To assess the importance of JA, RNA interference constructs targeted at the JA biosynthetic genes, allene oxide cyclase and 12-oxophytodienoic acid reductase, were expressed in transgenic potato plants. In addition, expression of the F-box protein COI1 was reduced by RNA interference. Plants expressing the RNA interference constructs failed to accumulate the respective transcripts in response to wounding or Pep-13 treatment, neither did they contain significant amounts of JA after elicitation. In response to infiltration of Pep-13, the transgenic plants exhibited a highly reduced accumulation of reactive oxygen species as well as reduced hypersensitive cell death. The ability of the JA-deficient plants to accumulate SA suggests that SA accumulation is independent or upstream of JA accumulation. These data show that PAMP responses in potato require both SA and JA and that, in contrast to Arabidopsis, these compounds act in the same signal transduction pathway. Despite their inability to fully respond to PAMP treatment, the transgenic RNA interference plants are not altered in their basal defense against Phytophthora infestans.

  9. Inherited phenotype instability of inflorescence and floral organ development in homeotic barley double mutants and its specific modification by auxin inhibitors and 2,4-D.

    Science.gov (United States)

    Šiukšta, Raimondas; Vaitkūnienė, Virginija; Kaselytė, Greta; Okockytė, Vaiva; Žukauskaitė, Justina; Žvingila, Donatas; Rančelis, Vytautas

    2015-03-01

    Barley (Hordeum vulgare) double mutants Hv-Hd/tw2, formed by hybridization, are characterized by inherited phenotypic instability and by several new features, such as bract/leaf-like structures, long naked gaps in the spike, and a wide spectrum of variations in the basic and ectopic flowers, which are absent in single mutants. Several of these features resemble those of mutations in auxin distribution, and thus the aim of this study was to determine whether auxin imbalances are related to phenotypic variations and instability. The effects of auxin inhibitors and 2,4-D (2,4-dichlorophenoxyacetic acid) on variation in basic and ectopic flowers were therefore examined, together with the effects of 2,4-D on spike structure. The character of phenotypic instability and the effects of auxin inhibitors and 2,4-D were compared in callus cultures and intact plants of single homeotic Hv-tw2 and Hv-Hooded/Kap (in the BKn3 gene) mutants and alternative double mutant lines: offspring from individual plants in distal hybrid generations (F9-F10) that all had the same BKn3 allele as determined by DNA sequencing. For intact plants, two auxin inhibitors, 9-hydroxyfluorene-9-carboxylic acid (HFCA) and p-chlorophenoxyisobutyric acid (PCIB), were used. Callus growth and flower/spike structures of the Hv-tw2 mutant differed in their responses to HFCA and PCIB. An increase in normal basic flowers after exposure to auxin inhibitors and a decrease in their frequencies caused by 2,4-D were observed, and there were also modifications in the spectra of ectopic flowers, especially those with sexual organs, but the effects depended on the genotype. Exposure to 2,4-D decreased the frequency of short gaps and lodicule transformations in Hv-tw2 and of long naked gaps in double mutants. The effects of auxin inhibitors and 2,4-D suggest that ectopic auxin maxima or deficiencies arise in various regions of the inflorescence/flower primordia. Based on the phenotypic instability observed, definite

  10. A novel Arabidopsis CHITIN ELICITOR RECEPTOR KINASE 1 (CERK1) mutant with enhanced pathogen-induced cell death and altered receptor processing.

    Science.gov (United States)

    Petutschnig, Elena K; Stolze, Marnie; Lipka, Ulrike; Kopischke, Michaela; Horlacher, Juliane; Valerius, Oliver; Rozhon, Wilfried; Gust, Andrea A; Kemmerling, Birgit; Poppenberger, Brigitte; Braus, Gerhard H; Nürnberger, Thorsten; Lipka, Volker

    2014-12-01

    Plants detect pathogens by sensing microbe-associated molecular patterns (MAMPs) through pattern recognition receptors. Pattern recognition receptor complexes also have roles in cell death control, but the underlying mechanisms are poorly understood. Here, we report isolation of cerk1-4, a novel mutant allele of the Arabidopsis chitin receptor CERK1 with enhanced defense responses. We identified cerk1-4 in a forward genetic screen with barley powdery mildew and consequently characterized it by pathogen assays, mutant crosses and analysis of defense pathways. CERK1 and CERK1-4 proteins were analyzed biochemically. The cerk1-4 mutation causes an amino acid exchange in the CERK1 ectodomain. Mutant plants maintain chitin signaling capacity but exhibit hyper-inducible salicylic acid concentrations and deregulated cell death upon pathogen challenge. In contrast to chitin signaling, the cerk1-4 phenotype does not require kinase activity and is conferred by the N-terminal part of the receptor. CERK1 undergoes ectodomain shedding, a well-known process in animal cell surface proteins. Wild-type plants contain the full-length CERK1 receptor protein as well as a soluble form of the CERK1 ectodomain, whereas cerk1-4 plants lack the N-terminal shedding product. Our work suggests that CERK1 may have a chitin-independent role in cell death control and is the first report of ectodomain shedding in plants. © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

  11. Sugar signaling regulation by Arabidopsis SIZ1-driven sumoylation is independent of salicylic acid

    DEFF Research Database (Denmark)

    Castro, Pedro Humberto Araújo R F; Verde, Nuno; Tavares, Rui Manuel

    2018-01-01

    inefficient responses to nutrient imbalance in phosphate, nitrate and copper. Recently, we reported that siz1 also displays altered responses to exogenous sugar supplementation. The siz1 mutant is a salicylic acid (SA) accumulator, and SA may interfere with sugar-dependent responses and signaling events. Here......, we extended our previous studies to determine the importance of SA in the SIZ1 response to sugars, by introducing the bacterial salicylate hydroxylase NahG into the siz1 background. Results demonstrate that siz1 phenotypes involving delayed germination are partially dependent of SA levels, whereas...

  12. Studies on auxotrophic mutants of Auricularia auricula and Auricularia fuscosucinea induced by irradiation

    International Nuclear Information System (INIS)

    Han Xincai; Yang Xinmei

    1991-01-01

    The induction of auxotrophic mutants of Auricularia auricula and Auricularia fuscosucinea from monokaryotic basidiospore by means of 60 Co-γ ray irradiation was reported. Under the irradiations of 10 krad-200 krad, 9 auxotrophic mutant strains were obtained, including 8 strains of A. auricula and 1 strain of A. fuscosucinea. The frequency of mutagenesis was 2.38 x 10 -3 -44.4 x 10 -3 . It was found that the optimum irradiation dose for A. auricula was 200 krad and for A. fuscosucinea was 10 krad. Biochemical and physiological researches indicated that the colony morphology, the hyphae growth speed, the contents of amino acid and the pattern of esterase isozyme of the mutants were different from those of the prototrophic strains

  13. Abscisic Acid Synthesis and Response

    Science.gov (United States)

    Finkelstein, Ruth

    2013-01-01

    Abscisic acid (ABA) is one of the “classical” plant hormones, i.e. discovered at least 50 years ago, that regulates many aspects of plant growth and development. This chapter reviews our current understanding of ABA synthesis, metabolism, transport, and signal transduction, emphasizing knowledge gained from studies of Arabidopsis. A combination of genetic, molecular and biochemical studies has identified nearly all of the enzymes involved in ABA metabolism, almost 200 loci regulating ABA response, and thousands of genes regulated by ABA in various contexts. Some of these regulators are implicated in cross-talk with other developmental, environmental or hormonal signals. Specific details of the ABA signaling mechanisms vary among tissues or developmental stages; these are discussed in the context of ABA effects on seed maturation, germination, seedling growth, vegetative stress responses, stomatal regulation, pathogen response, flowering, and senescence. PMID:24273463

  14. Acid tolerance response and survival by oral bacteria.

    Science.gov (United States)

    Svensäter, G; Larsson, U B; Greif, E C; Cvitkovitch, D G; Hamilton, I R

    1997-10-01

    Using 21 species of oral bacteria, representing six acidogenic genera, we undertook to determine whether the pH-limiting exponential growth is related to the ability of the organisms to generate an acid-tolerance response that results in enhanced survival at low pH. The lower pH limit of exponential growth varied by more than two units with that of Neisseria A182 at pH 6.34; growth of Lactobacillus casei RB1014 stopped at pH 3.81, with species of Actinomyces, Enterococcus, Prevotella and Streptococcus falling between these limits. The working hypothesis was that the organisms with the higher pH limits for growth are unable to respond to acidic environments in order to survive, whereas the more aciduric organisms would possess or acquire acid tolerance. Adaptation to acid tolerance was tested by determining whether the prior exposure of exponential-phase cells to a low, sub-lethal pH would trigger the induction of a mechanism that would enhance survival at a pH killing pH 7.5 control cells. The killing pH varied from pH 4.5 for Prevotella intermedia ATCC 25611 to pH 2.3 for the three Lactobacillus casei strains in the study, with the three Streptococcus mutans strains killed at pH 3.0 for 3 h. The adaptation experiments revealed three groups of organisms: non-acid-responders, generally representing strains with the highest terminal pH values; weak acid-responders in the middle of the pH list, generating low numbers of survivors at one or two pH values, and the aciduric, strong responders generating a high number of survivors at pH values in the range 6.0 to 3.5, but not at pH 7.5. Predominant among the latter group were the S. mutans and Lactobacilli casei strains, with the most significant adaptive response exhibited by S. mutans LT11 and S. mutans Ingbritt, involving a process that required protein synthesis. Time course experiments with the latter organisms indicated that 90-120 min was required after exposure to the triggering pH before the acid response was

  15. Activities of native and tyrosine-69 mutant phospholipases A2 on phospholipid analogues. A reevaluation of the minimal substrate requirements.

    Science.gov (United States)

    Kuipers, O P; Dekker, N; Verheij, H M; de Haas, G H

    1990-06-26

    The role of Tyr-69 of porcine pancreatic phospholipase A2 in substrate binding was studied with the help of proteins modified by site-directed mutagenesis and phospholipid analogues with a changed head-group geometry. Two mutants were used containing Phe and Lys, respectively, at position 69. Modifications in the phospholipids included introduction of a sulfur at the phosphorus (thionophospholipids), removal of the negative charge at phosphorus (phosphatidic acid dimethyl ester), and reduction (phosphonolipids) or extension (diacylbutanetriol choline phosphate) of the distance between the phosphorus and the acyl ester bond. Replacement of Tyr-69 by Lys reduces enzymatic activity, but the mutant enzyme retains both the stereospecificity and positional specificity of native phospholipase A2. The Phe-69 mutant not only hydrolyzes the Rp isomer of thionophospholipids more efficiently than the wild-type enzyme, but the Sp thiono isomer is hydrolyzed too, although at a low (approximately 4%) rate. Phosphonolipids are hydrolyzed by native phospholipase A2 about 7 times more slowly than natural phospholipids, with retention of positional specificity and a (partial) loss of stereospecificity. The dimethyl ester of phosphatidic acid is degraded efficiently in a calcium-dependent and positional-specific way by native phospholipase A2 and by the mutants, indicating that a negative charge at phosphorus is not an absolute substrate requirement. The activities on the phosphatidic acid dimethyl ester of native enzyme and the Lys-69 mutant are lower than those on the corresponding lecithin, in contrast to the Phe-69 mutant, which has equal activities on both substrates.(ABSTRACT TRUNCATED AT 250 WORDS)

  16. Comparative proteomic analysis of a high-tillering dwarf mutant induced by spaceflight at different tillering stages

    International Nuclear Information System (INIS)

    Wang Wei; Wei Lijun; Wang Junmin; Xu Jianlong; Sun Yeqing

    2011-01-01

    To investigate changes of proteins during rice tiller development, a comparative proteomic analysis between a high-tillering dwarf mutant R955 induced by spaceflight and its ground control was performed at three developmental stages during the vegetative growth, i. e. at days 14 (no tiller appearance), 21 (initial tillering) and 55 (maximum tillering) after sowing. Analysis of the protein spots on two-dimensional fluorescence difference gel electrophoresis 2-D DIGE images revealed 97 proteins that were differentially expressed at the three stages. Among them, 59 unique proteins were successfully identified by mass spectrometry. Through functional and quantitative analysis of proteomic data, it was found that biological processes including energy pathway, photosynthesis, protein metabolism, nitrogen assimilation, amino acid metabolism and stimulus response were mainly involved in tiller development in mutant plant. K-means clustering revealed that proteins regulated at different stages tended to be involved in different biological processes. Two-way analysis of variance (Two-way ANOVA) showed that S-like was directly correlated RNase with the high-tillering ability. (authors)

  17. The small ethylene response factor ERF96 is involved in the regulation of the abscisic acid response in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Xiaoping eWang

    2015-11-01

    Full Text Available Ethylene regulates many aspects of plant growth and development including seed germination, leaf senescence, and fruit ripening, and of plant responses to environmental stimuli including both biotic and abiotic stresses. Ethylene Response Factors (ERFs are plant-specific transcription factors and are a subfamily of the AP2 (APETALA2/ERF transcription factor family. The function of many members in this large gene family remains largely unknown. ERF96, a member of the Group IX ERF family transcription factors, has recently been shown to be a transcriptional activator that is involved in plant defense response in Arabidopsis. Here we provide evidence that ERF96 is a positive regulator of abscisic acid (ABA responses. Bioinformatics analysis indicated that there are a total four small ERFs in Arabidopsis including ERF95, ERF96, ERF97 and ERF98, and that ERF96 forms a cluster with ERF95 and ERF97. By using quantitative RT-PCR, we found that ERF96 is expressed in all tissues and organs examined except roots, with relatively high expression in flowers and seeds. Results from the protoplast transfection assay results indicated that the EDLL motif-containing C-terminal domain is responsible for ERF96’s transcriptional activity. Although loss-of-function mutant of ERF96 was morphologically similar to wild type plants, transgenic plants overexpressing ERF96 had smaller rosette size and were delayed in flowering time. In ABA sensitivity assays, we found that ERF96 overexpression plants were hypersensitive to ABA in terms of ABA inhibition of seed germination, early seedling development and root elongation. Consistent with these observations, elevated transcript levels of some ABA-responsive genes including RD29A, ABI5, ABF3, ABF4, P5CS and COR15A were observed in the transgenic plants in the presence of ABA. However, in the absence of ABA treatment, the transcript levels of these ABA-responsive genes remained largely unchanged. Our experiments also showed

  18. Phenotypic characterization of adenovirus type 12 temperature-sensitive mutants in productive infection and transformation.

    Science.gov (United States)

    Hama, S; Kimura, G

    1980-01-01

    Eleven temperature-sensitive mutants of adenovirus type 12, capable of forming plaques in human cells at 33 C but not at 39.5 C, were isolated from a stock of a wild-type strain after treatment with either nitrous acid or hydroxylamine. Complementation tests in doubly infected human cells permitted a tentative assignment of eight of these mutants to six complementation groups. Temperature-shift experiments revealed that one mutant is affected early and most of the other mutants are affected late. Only the early mutant, H12ts505, was temperature sensitive in viral DNA replication. Infectious virions of all the mutants except H12ts505 and two of the late mutants produced at 33 C, appeared to be more heat labile than those of the wild type. Only H12ts505 was temperature sensitive for the establishment of transformation of rat 3Y1 cells. One of the late mutants (H12ts504) had an increased transforming ability at the permissive temperature. Results of temperature-shift transformation experiments suggest that a viral function affected in H12ts505 is required for "initiation" of transformation. Some of the growth properties of H12ts505-transformed cells were also temperature dependent, suggesting that a functional expression of a gene mutation in H12ts505 is required to maintain at least some aspects of the transformed state.

  19. The putative E3 ubiquitin ligase ECERIFERUM9 regulates abscisic acid biosynthesis and response during seed germination and postgermination growth in arabidopsis

    KAUST Repository

    Zhao, Huayan

    2014-05-08

    The ECERIFERUM9 (CER9) gene encodes a putative E3 ubiquitin ligase that functions in cuticle biosynthesis and the maintenance of plant water status. Here, we found that CER9 is also involved in abscisic acid (ABA) signaling in seeds and young seedlings of Arabidopsis (Arabidopsis thaliana). The germinated embryos of the mutants exhibited enhanced sensitivity to ABA during the transition from reversible dormancy to determinate seedling growth. Expression of the CER9 gene is closely related to ABA levels and displays a similar pattern to that of ABSCISIC ACID-INSENSITIVE5 (ABI5), which encodes a positive regulator of ABA responses in seeds. cer9 mutant seeds exhibited delayed germination that is independent of seed coat permeability. Quantitative proteomic analyses showed that cer9 seeds had a protein profile similar to that of the wild type treated with ABA. Transcriptomics analyses revealed that genes involved in ABA biosynthesis or signaling pathways were differentially regulated in cer9 seeds. Consistent with this, high levels of ABA were detected in dry seeds of cer9. Blocking ABA biosynthesis by fluridone treatment or by combining an ABA-deficient mutation with cer9 attenuated the phenotypes of cer9. Whereas introduction of the abi1-1, abi3-1, or abi4-103 mutation could completely eliminate the ABA hypersensitivity of cer9, introduction of abi5 resulted only in partial suppression. These results indicate that CER9 is a novel negative regulator of ABA biosynthesis and the ABA signaling pathway during seed germination. © 2014 American Society of Plant Biologists. All Rights Reserved.

  20. The Putative E3 Ubiquitin Ligase ECERIFERUM9 Regulates Abscisic Acid Biosynthesis and Response during Seed Germination and Postgermination Growth in Arabidopsis.

    Science.gov (United States)

    Zhao, Huayan; Zhang, Huoming; Cui, Peng; Ding, Feng; Wang, Guangchao; Li, Rongjun; Jenks, Matthew A; Lü, Shiyou; Xiong, Liming

    2014-07-01

    The ECERIFERUM9 (CER9) gene encodes a putative E3 ubiquitin ligase that functions in cuticle biosynthesis and the maintenance of plant water status. Here, we found that CER9 is also involved in abscisic acid (ABA) signaling in seeds and young seedlings of Arabidopsis (Arabidopsis thaliana). The germinated embryos of the mutants exhibited enhanced sensitivity to ABA during the transition from reversible dormancy to determinate seedling growth. Expression of the CER9 gene is closely related to ABA levels and displays a similar pattern to that of ABSCISIC ACID-INSENSITIVE5 (ABI5), which encodes a positive regulator of ABA responses in seeds. cer9 mutant seeds exhibited delayed germination that is independent of seed coat permeability. Quantitative proteomic analyses showed that cer9 seeds had a protein profile similar to that of the wild type treated with ABA. Transcriptomics analyses revealed that genes involved in ABA biosynthesis or signaling pathways were differentially regulated in cer9 seeds. Consistent with this, high levels of ABA were detected in dry seeds of cer9. Blocking ABA biosynthesis by fluridone treatment or by combining an ABA-deficient mutation with cer9 attenuated the phenotypes of cer9. Whereas introduction of the abi1-1, abi3-1, or abi4-103 mutation could completely eliminate the ABA hypersensitivity of cer9, introduction of abi5 resulted only in partial suppression. These results indicate that CER9 is a novel negative regulator of ABA biosynthesis and the ABA signaling pathway during seed germination. © 2014 American Society of Plant Biologists. All Rights Reserved.

  1. Blood ketone response to norepinephrine-induced free fatty acid in diabetes

    Energy Technology Data Exchange (ETDEWEB)

    Blackard, W G; Omori, Yoshiaki

    1963-04-18

    During 90-minute norepinephrine infusions, blood free fatty acid and ketone responses of Japanese nondiabetic and diabetic subjects were determined. Nonobese diabetic subjects with and without fasting hyperglycemia demonstrated significantly greater blood ketone elevations than nondiabetics. An inverse correlation between obesity and blood ketone response to nonrepinephrine was observed in diabetics. This correlation could not be attributed to varying degrees of fasting hyperglycemia or free fatty acid elevation. Nonobese diabetics with mild fasting hyperglycemia (90 to 150 mg%) exhibited an unexpected greater increase in blood ketones than nonobese diabetics with moderate fasting hyperglycemia (150 to 250 mg%). Differences in free fatty acid elevations were not responsible for this apparent paradox. The magnitude of the hyperketonemic response, though dependent on free fatty elevation, seemed more sensitive to the degree of obesity and the fasting blood glucose level. Fractional ketone body measurements attributed the blood ketone elevations predominantly to ..beta..-hydroxybutyric acid increases. 43 references, 6 figures, 1 table.

  2. Fusarium graminearum PKS14 is involved in orsellinic acid and orcinol synthesis

    DEFF Research Database (Denmark)

    Jørgensen, Simon Hartung; Frandsen, Rasmus John Normand; Nielsen, Kristian Fog

    2014-01-01

    and cultivated two of the resulting mutants on RM medium. This led to the production of two compounds, which were only detected in the PKS14 overexpressing mutants and not in the wild type or PKS14 deletion mutants. The two compounds were tentatively identified as orsellinic acid and orcinol by comparing...... spectroscopic data (mass spectroscopy and chromatography) to authentic standards. NMR analysis of putative orcinol isolated from the PKS14 overexpressing mutant supported our identification. Orcinol and orsellinic acid, not previously detected in Fusarium, have primarily been detected in lichen fungi...

  3. Characterization of the Symbiotic Nitrogen-Fixing Common Bean Low Phytic Acid (lpa1) Mutant Response to Water Stress.

    Science.gov (United States)

    Chiozzotto, Remo; Ramírez, Mario; Talbi, Chouhra; Cominelli, Eleonora; Girard, Lourdes; Sparvoli, Francesca; Hernández, Georgina

    2018-02-15

    The common bean ( Phaseolus vulgaris L.) low phytic acid ( lpa1 ) biofortified genotype produces seeds with improved nutritional characteristics and does not display negative pleiotropic effects. Here we demonstrated that lpa1 plants establish an efficient nitrogen-fixing symbiosis with Rhizobium etli CE3. The lpa1 nodules showed a higher expression of nodule-function related genes than the nodules of the parental wild type genotype (BAT 93). We analyzed the response to water stress of lpa1 vs. BAT 93 plants grown under fertilized or under symbiotic N₂-fixation conditions. Water stress was induced by water withholding (up to 14% soil moisture) to fertilized or R. etli nodulated plants previously grown with normal irrigation. The fertilized lpa1 plants showed milder water stress symptoms during the water deployment period and after the rehydration recovery period when lpa1 plants showed less biomass reduction. The symbiotic water-stressed lpa1 plants showed decreased nitrogenase activity that coincides with decreased sucrose synthase gene expression in nodules; lower turgor weight to dry weight (DW) ratio, which has been associated with higher drought resistance index; downregulation of carbon/nitrogen (C/N)-related and upregulation of stress-related genes. Higher expression of stress-related genes was also observed in bacteroids of stressed lpa1 plants that also displayed very high expression of the symbiotic cbb ₃ oxidase ( fixN d).

  4. X-ray induced degradation of DNA in radiosensitive mutants of Anacystis nidulans

    Energy Technology Data Exchange (ETDEWEB)

    Zhukas, K I; Vorontsova, G V; Groshev, V V; Shestakov, S V [Moskovskij Gosudarstvennyj Univ. (USSR). Biologo-Pochvennyj Fakul' tet

    1975-01-01

    In irradiated Cyanophyceae (Anacystis nidulans) cells there occurs a process of DNA degeneration to acid-soluble products which is linked with protein synthesis and stimulated by caffeine and acriflavine. The degree of DNA degeneration increases with x-ray dose, is not very dependent on the composition of the incubation medium and is weakly linked with photosynthesis. In the cells of a radiation-resistant mutant the degree of DNA degeneration is slighter, and in the cells of radiosensitive mutants larger, than in ordinary cells. The role of DNA degradation in the radiation detruction of cells is discussed.

  5. Structural features and activity of Brazzein and its mutants upon substitution of a surfaced exposed alanine.

    Science.gov (United States)

    Ghanavatian, Parisa; Khalifeh, Khosrow; Jafarian, Vahab

    2016-12-01

    Brazzein (Brz) is a member of sweet-tasting protein containing four disulfide bonds. It was reported as a compact and heat-resistant protein. Here, we have used site-directed mutagenesis and replaced a surface-exposed alanine with aspartic acid (A19D mutant), lysine (A19K mutant) and glycine (A19G mutant). Activity comparisons of wild-type (WT) and mutants using taste panel test procedure showed that A19G variant has the same activity as WT protein. However, introduction of a positive charge in A19K mutant led to significant increase in Brz's sweetness, while A19D has reduced sweetness compared to WT protein. Docking studies showed that mutation at position 19 results in slight chain mobility of protein at the binding surface and changing the patterns of interactions toward more effective binding of E9K variant in the concave surface of sweet taste receptor. Far-UV CD data spectra have a characteristic shape of beta structure for all variants, however different magnitudes of spectra suggest that beta-sheet structure in WT and A19G is more stable than that of A19D and A19K. Equilibrium unfolding studies with fluorescence spectroscopy and using urea and dithiothritol (DTT) as chemical denaturants indicates that A19G mutant gains more stability against urea denaturation; while conformational stability of A19D and A19K decreases when compared with WT and A19G variants. We concluded that the positive charge at the surface of protein is important factor responsible for the interaction of protein with the human sweet receptor and Ala 19 can be considered as a key region for investigating the mechanism of the interaction of Brz with corresponding receptor. Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  6. Differential Impact of LPG-and PG-Deficient Leishmania major Mutants on the Immune Response of Human Dendritic Cells.

    Directory of Open Access Journals (Sweden)

    Michelle A Favila

    2015-12-01

    Full Text Available Leishmania major infection induces robust interleukin-12 (IL12 production in human dendritic cells (hDC, ultimately resulting in Th1-mediated immunity and clinical resolution. The surface of Leishmania parasites is covered in a dense glycocalyx consisting of primarily lipophosphoglycan (LPG and other phosphoglycan-containing molecules (PGs, making these glycoconjugates the likely pathogen-associated molecular patterns (PAMPS responsible for IL12 induction.Here we explored the role of parasite glycoconjugates on the hDC IL12 response by generating L. major Friedlin V1 mutants defective in LPG alone, (FV1 lpg1-, or generally deficient for all PGs, (FV1 lpg2-. Infection with metacyclic, infective stage, L. major or purified LPG induced high levels of IL12B subunit gene transcripts in hDCs, which was abrogated with FV1 lpg1- infections. In contrast, hDC infections with FV1 lpg2- displayed increased IL12B expression, suggesting other PG-related/LPG2 dependent molecules may act to dampen the immune response. Global transcriptional profiling comparing WT, FV1 lpg1-, FV1 lpg2- infections revealed that FV1 lpg1- mutants entered hDCs in a silent fashion as indicated by repression of gene expression. Transcription factor binding site analysis suggests that LPG recognition by hDCs induces IL-12 in a signaling cascade resulting in Nuclear Factor κ B (NFκB and Interferon Regulatory Factor (IRF mediated transcription.These data suggest that L. major LPG is a major PAMP recognized by hDC to induce IL12-mediated protective immunity and that there is a complex interplay between PG-baring Leishmania surface glycoconjugates that result in modulation of host cellular IL12.

  7. The Dynamics of Embolism Refilling in Abscisic Acid (ABA-Deficient Tomato Plants

    Directory of Open Access Journals (Sweden)

    Francesca Secchi

    2012-12-01

    Full Text Available Plants are in danger of embolism formation in xylem vessels when the balance between water transport capacity and transpirational demand is compromised. To maintain this delicate balance, plants must regulate the rate of transpiration and, if necessary, restore water transport in embolized vessels. Abscisic acid (ABA is the dominant long-distance signal responsible for plant response to stress, and it is possible that it plays a role in the embolism/refilling cycle. To test this idea, a temporal analysis of embolism and refilling dynamics, transpiration rate and starch content was performed on ABA-deficient mutant tomato plants. ABA-deficient mutants were more vulnerable to embolism formation than wild-type plants, and application of exogenous ABA had no effect on vulnerability. However, mutant plants treated with exogenous ABA had lower stomatal conductance and reduced starch content in the xylem parenchyma cells. The lower starch content could have an indirect effect on the plant’s refilling activity. The results confirm that plants with high starch content (moderately stressed mutant plants were more likely to recover from loss of water transport capacity than plants with low starch content (mutant plants with application of exogenous ABA or plants experiencing severe water stress. This study demonstrates that ABA most likely does not play any direct role in embolism refilling, but through the modulation of carbohydrate content, it could influence the plant’s capacity for refilling.

  8. The Arabidopsis thaliana cysteine-rich receptor-like kinase CRK20 modulates host responses to Pseudomonas syringae pv. tomato DC3000 infection

    KAUST Repository

    Ederli, Luisa; Madeo, Laura; Calderini, Ornella; Gehring, Christoph A; Moretti, Chiaraluce; Buonaurio, Roberto; Paolocci, Francesco; Pasqualini, Stefania

    2011-01-01

    by pathogens, salicylic acid and ozone (O3). However, its role in responses to biotic and abiotic stress remains to be elucidated. To determine the function of CRK20 in such responses, two CRK20 loss-of-function mutants, crk20-1 and crk20-2, were isolated from

  9. Comparison of arabidopsis stomatal density mutants indicates variation in water stress responses and potential epistatic effects

    Science.gov (United States)

    Shaneka S. Lawson; Paula M. Pijut; Charles H. Michler

    2014-01-01

    Recent physiological analysis of Arabidopsis stomatal density (SD) mutants indicated that SD was not the major factor controlling aboveground biomass accumulation. Despite the general theory that plants with fewer stomata have limited biomass acquisition capabilities, epf1 and several other Arabidopsis mutants varied significantly in leaf fresh...

  10. Gamma rays induced mutation for low phytic acid and trypsin inhibitor content in soybean

    International Nuclear Information System (INIS)

    Gupta, S.K.; Manjaya, J.G.

    2017-01-01

    Soybean (Glycine max (L.) Merrill) is an important source of vegetable protein and is used as a food, feed and health supplement. However, consumption of soybean as food is limited because of the presence of many anti-nutritional factors. Trypsin inhibitors and phytic acid are two major anti-nutritional factors present in soybean that need to be removed for increasing the soybean consumption as food. Trypsin inhibitor is known to inhibit the trypsin/chymotrpsin activity and phytic acid reduces the bioavailability of essential micronutrients in digestive tract, resulting in adverse effect on health. Therefore, developing soybean cultivars having low trypsin inhibitors and phytic acid content is highly desirable. Soybean cultivar JS 93-05 was irradiated with 250 Gy gamma rays to induce mutation for various morphological and biochemical characters. A large number of mutants with altered morphological characters were identified. Ninety true breeding mutant lines in M6 generation were screened for trypsin inhibitor and phytic acid content. The phytic acid content was estimated using modified colorimetric method and trypsin inhibitor concentration was estimated using BAPNA as substrate in colorimetric method. The phytic acid content in the mutants varied from 7.59 to 24.14 mg g -1 . Two mutants lines TSG - 62 (7.59 mg g -1 ) and TSG - 66 (9.62 mg g -1 ) showed significant low phytic acid content as compared to the parent JS 93-05 (20.19 mg g -1 ). The trypsin inhibitor concentration in the mutants varied from 19.92 to 53.64 TIU mg -1 and one mutant line (TSG -14) was found with the lowest trypsin inhibitor concentration of 19.92 TIU mg -1 compared to parent JS 93-05 (50.90 TIU mg -1 ). The mutant lines identified in this study will serve as important genetic resources for developing low phytic acid and low trypsin inhibitor cultivars in soybean. (author)

  11. Escherichia coli ArgR mutants defective in cer/Xer recombination, but not in DNA binding.

    Science.gov (United States)

    Sénéchal, Hélène; Delesques, Jérémy; Szatmari, George

    2010-04-01

    The Escherichia coli arginine repressor (ArgR) is an L-arginine-dependent DNA-binding protein that controls the expression of the arginine biosynthetic genes and is required as an accessory factor for Xer site-specific recombination at cer and related recombination sites in plasmids. We used the technique of pentapeptide scanning mutagenesis to isolate a series of ArgR mutants that were considerably reduced in cer recombination, but were still able to repress an argA::lacZ fusion. DNA sequence analysis showed that all of the mutants mapped to the same nucleotide, resulting in a five amino acid insertion between residues 149 and 150 of ArgR, corresponding to the end of the alpha6 helix. A truncated ArgR containing a stop codon at residue 150 displayed the same phenotype as the protein with the five amino acid insertion, and both mutants displayed sequence-specific DNA-binding activity that was L-arginine dependent. These results show that the C-terminus of ArgR is more important in cer/Xer site-specific recombination than in DNA binding.

  12. Leaf and canopy photosynthesis of a chlorophyll deficient soybean mutant.

    Science.gov (United States)

    Sakowska, Karolina; Alberti, Giorgio; Genesio, Lorenzo; Peressotti, Alessandro; Delle Vedove, Gemini; Gianelle, Damiano; Colombo, Roberto; Rodeghiero, Mirco; Panigada, Cinzia; Juszczak, Radosław; Celesti, Marco; Rossini, Micol; Haworth, Matthew; Campbell, Benjamin W; Mevy, Jean-Philippe; Vescovo, Loris; Cendrero-Mateo, M Pilar; Rascher, Uwe; Miglietta, Franco

    2018-03-02

    The photosynthetic, optical, and morphological characteristics of a chlorophyll-deficient (Chl-deficient) "yellow" soybean mutant (MinnGold) were examined in comparison with 2 green varieties (MN0095 and Eiko). Despite the large difference in Chl content, similar leaf photosynthesis rates were maintained in the Chl-deficient mutant by offsetting the reduced absorption of red photons by a small increase in photochemical efficiency and lower non-photochemical quenching. When grown in the field, at full canopy cover, the mutants reflected a significantly larger proportion of incoming shortwave radiation, but the total canopy light absorption was only slightly reduced, most likely due to a deeper penetration of light into the canopy space. As a consequence, canopy-scale gross primary production and ecosystem respiration were comparable between the Chl-deficient mutant and the green variety. However, total biomass production was lower in the mutant, which indicates that processes other than steady state photosynthesis caused a reduction in biomass accumulation over time. Analysis of non-photochemical quenching relaxation and gas exchange in Chl-deficient and green leaves after transitions from high to low light conditions suggested that dynamic photosynthesis might be responsible for the reduced biomass production in the Chl-deficient mutant under field conditions. © 2018 John Wiley & Sons Ltd.

  13. Characterization of reduced height mutant of emmer wheat var. NP200 (Triticum dicoccum)

    International Nuclear Information System (INIS)

    Suman, Sud; Nayeem, K.A.; Bhagwat, S.G.

    2006-01-01

    Full text: Emmer wheat commonly known as Khapli is cultivated on limited area in Tamil Nadu, Andhra Pradesh, Karnataka, Maharashtra and Gujarat. Although cultivation of emmer wheat is confirmed to a small area, improvement work in this species is gaining importance because of its potential for diabetic patients and high dietary fibre in comparison to durum and bread wheats. Emmer wheat cultivar NP200 is a selection from local wheats of Andhra Pradesh. The cultivar NP200 is tall and is prone to lodging leading to yield loss. Therefore, systematic effort to improve cultivar NP200 is needed with the objective to reduce height and introduce lodging tolerance and to improve harvest index. The cultivar NP200 was irradiated with γ-rays. A reduced height mutant with vigorous growth and high tillering was found in M2 population. The mutant was designated as HW1095. The progeny of mutant in M3 showed 35.7 percent reduction in height as compared to parent. The HW1095 mutant was subjected to gibberellic acid treatment at seedling stage and was found to be insensitive to gibberellic acid. An allele specific marker for major dwarfing gene Rht B1b was used to check the status of dwarfing gene in semi dwarf emmer (DDK1009, DDK1025, HW5013, HW5301 and MACS2961) and tall emmer (Np200 and NP201), semi dwarf durums (HD4502, HD4530, MACS2846) along with dwarf mutant (HW1095). The validity of primer in semi dwarf durums and emmer for Rht B 1b gene was found to be perfect. The parent variety NP200 showed presence of wild type allele (Rht B1a) with the primer pair BF-WR1. All semi dwarf emmer showed a band of 237 bp with primer pair BF-MR1. However, mutant (HW1095) showed absence of amplification for both Rht B1a and Rht B1b alleles with respective primer pairs. The results indicated that the reduced height mutant carried a mutation different than from the existing allele (Rht B1b)

  14. Responsive Boronic Acid-Decorated (Co)polymers: From Glucose Sensors to Autonomous Drug Delivery.

    Science.gov (United States)

    Vancoillie, Gertjan; Hoogenboom, Richard

    2016-10-19

    Boronic acid-containing (co)polymers have fascinated researchers for decades, garnering attention for their unique responsiveness toward 1,2- and 1,3-diols, including saccharides and nucleotides. The applications of materials that exert this property are manifold including sensing, but also self-regulated drug delivery systems through responsive membranes or micelles. In this review, some of the main applications of boronic acid containing (co)polymers are discussed focusing on the role of the boronic acid group in the response mechanism. We hope that this summary, which highlights the importance and potential of boronic acid-decorated polymeric materials, will inspire further research within this interesting field of responsive polymers and polymeric materials.

  15. A Molecular and Cellular Context-Dependent Role for Ir76b in Detection of Amino Acid Taste

    Directory of Open Access Journals (Sweden)

    Anindya Ganguly

    2017-01-01

    Full Text Available Amino acid taste is expected to be a universal property among animals. Although sweet, bitter, salt, and water tastes have been well characterized in insects, the mechanisms underlying amino acid taste remain elusive. From a Drosophila RNAi screen, we identify an ionotropic receptor, Ir76b, as necessary for yeast preference. Using calcium imaging, we identify Ir76b+ amino acid taste neurons in legs, overlapping partially with sweet neurons but not those that sense other tastants. Ir76b mutants have reduced responses to amino acids, which are rescued by transgenic expression of Ir76b and a mosquito ortholog AgIr76b. Co-expression of Ir20a with Ir76b is sufficient for conferring amino acid responses in sweet-taste neurons. Notably, Ir20a also serves to block salt response of Ir76b. Our study establishes the role of a highly conserved receptor in amino acid taste and suggests a mechanism for mutually exclusive roles of Ir76b in salt- and amino-acid-sensing neurons.

  16. Analysis of two potential long-distance signaling molecules, LjCLE-RS1/2 and jasmonic acid, in a hypernodulating mutant too much love.

    Science.gov (United States)

    Magori, Shimpei; Kawaguchi, Masayoshi

    2010-04-01

    Legume plants tightly control the number and development of root nodules. This is partly regulated by a long-distance signaling known as auto-regulation of nodulation (AON). AON signaling involves at least two potential long-distance signals: root-derived signal and shoot-derived signal. However, their molecular characteristics and the mode of action remain unclear. In our recent study, we isolated a novel Lotus japonicus hypernodulating mutant too much love (tml). Based on several grafting experiments, we concluded that its causative gene TML functions as a receptor of the shoot-derived signal. This finding prompted us to ask how the candidates of the long-distance signal molecules, LjCLE-RS1/2 and jasmonic acid (JA), are affected in tml mutants. Expression analysis revealed that rapid induction of LjCLE-RS1/2 upon rhizobial inoculation is still intact in tml, supporting that TML plays a role in reception of the shoot-derived signal but not in generation of the root-derived signal. Furthermore, physiological analysis showed that JA, a candidate of the shoot-derived signal, can suppress tml hypernodulation. Therefore, contrary to the previous report, JA might not be a component of AON signaling.

  17. Recombination Phenotypes of Escherichia coli greA Mutants

    Directory of Open Access Journals (Sweden)

    Poteete Anthony R

    2011-03-01

    Full Text Available Abstract Background The elongation factor GreA binds to RNA polymerase and modulates transcriptional pausing. Some recent research suggests that the primary role of GreA may not be to regulate gene expression, but rather, to promote the progression of replication forks which collide with RNA polymerase, and which might otherwise collapse. Replication fork collapse is known to generate dsDNA breaks, which can be recombinogenic. It follows that GreA malfunction could have consequences affecting homologous recombination. Results Escherichia coli mutants bearing substitutions of the active site acidic residues of the transcription elongation factor GreA, D41N and E44K, were isolated as suppressors of growth inhibition by a toxic variant of the bacteriophage lambda Red-beta recombination protein. These mutants, as well as a D41A greA mutant and a greA deletion, were tested for proficiency in recombination events. The mutations were found to increase the efficiency of RecA-RecBCD-mediated and RecA-Red-mediated recombination, which are replication-independent, and to decrease the efficiency of replication-dependent Red-mediated recombination. Conclusion These observations provide new evidence for a role of GreA in resolving conflicts between replication and transcription.

  18. Physiological responses of root-less epiphytic plants to acid rain.

    Science.gov (United States)

    Kováčik, Jozef; Klejdus, Bořivoj; Bačkor, Martin; Stork, František; Hedbavny, Josef

    2011-03-01

    Selected physiological responses of Tillandsia albida (Bromeliaceae) and two lichens (Hypogymnia physodes and Xanthoria parietina) exposed to simulated acid rain (AR) over 3 months were studied. Pigments were depressed in all species being affected the most in Tillandsia. Amounts of hydrogen peroxide and superoxide were elevated and soluble proteins decreased only in AR-exposed Hypogymnia. Free amino acids were slightly affected among species and only glutamate sharply decreased in AR-exposed Xanthoria. Slight increase in soluble phenols but decrease in flavonoids in almost all species suggests that the latter are not essential for tolerance to AR. Almost all phenolic acids in Tillandsia leaves decreased in response to AR and activities of selected enzymes (phenylalanine ammonia-lyase, polyphenol oxidase, ascorbate- and guaiacol-peroxidase) were enhanced by AR. In lichens, considerable increase in metabolites (physodalic acid, atranorin and parietin) in response to AR was found but amount of ergosterol was unchanged. Macronutrients (K, Ca, Mg) decreased more pronouncedly in comparison with micronutrients in all species. Xanthoria showed higher tolerance in comparison with Hypogymnia, suggesting that could be useful for long-term biomonitoring.

  19. Phosphorylation of KasB Regulates Virulence and Acid-Fastness in Mycobacterium tuberculosis

    Science.gov (United States)

    Vilchèze, Catherine; Molle, Virginie; Carrère-Kremer, Séverine; Leiba, Jade; Mourey, Lionel; Shenai, Shubhada; Baronian, Grégory; Tufariello, Joann; Hartman, Travis; Veyron-Churlet, Romain; Trivelli, Xavier; Tiwari, Sangeeta; Weinrick, Brian; Alland, David; Guérardel, Yann; Jacobs, William R.; Kremer, Laurent

    2014-01-01

    Mycobacterium tuberculosis bacilli display two signature features: acid-fast staining and the capacity to induce long-term latent infections in humans. However, the mechanisms governing these two important processes remain largely unknown. Ser/Thr phosphorylation has recently emerged as an important regulatory mechanism allowing mycobacteria to adapt their cell wall structure/composition in response to their environment. Herein, we evaluated whether phosphorylation of KasB, a crucial mycolic acid biosynthetic enzyme, could modulate acid-fast staining and virulence. Tandem mass spectrometry and site-directed mutagenesis revealed that phosphorylation of KasB occurred at Thr334 and Thr336 both in vitro and in mycobacteria. Isogenic strains of M. tuberculosis with either a deletion of the kasB gene or a kasB_T334D/T336D allele, mimicking constitutive phosphorylation of KasB, were constructed by specialized linkage transduction. Biochemical and structural analyses comparing these mutants to the parental strain revealed that both mutant strains had mycolic acids that were shortened by 4–6 carbon atoms and lacked trans-cyclopropanation. Together, these results suggested that in M. tuberculosis, phosphorylation profoundly decreases the condensing activity of KasB. Structural/modeling analyses reveal that Thr334 and Thr336 are located in the vicinity of the catalytic triad, which indicates that phosphorylation of these amino acids would result in loss of enzyme activity. Importantly, the kasB_T334D/T336D phosphomimetic and deletion alleles, in contrast to the kasB_T334A/T336A phosphoablative allele, completely lost acid-fast staining. Moreover, assessing the virulence of these strains indicated that the KasB phosphomimetic mutant was attenuated in both immunodeficient and immunocompetent mice following aerosol infection. This attenuation was characterized by the absence of lung pathology. Overall, these results highlight for the first time the role of Ser/Thr kinase

  20. Phosphorylation of KasB regulates virulence and acid-fastness in Mycobacterium tuberculosis.

    Directory of Open Access Journals (Sweden)

    Catherine Vilchèze

    2014-05-01

    Full Text Available Mycobacterium tuberculosis bacilli display two signature features: acid-fast staining and the capacity to induce long-term latent infections in humans. However, the mechanisms governing these two important processes remain largely unknown. Ser/Thr phosphorylation has recently emerged as an important regulatory mechanism allowing mycobacteria to adapt their cell wall structure/composition in response to their environment. Herein, we evaluated whether phosphorylation of KasB, a crucial mycolic acid biosynthetic enzyme, could modulate acid-fast staining and virulence. Tandem mass spectrometry and site-directed mutagenesis revealed that phosphorylation of KasB occurred at Thr334 and Thr336 both in vitro and in mycobacteria. Isogenic strains of M. tuberculosis with either a deletion of the kasB gene or a kasB_T334D/T336D allele, mimicking constitutive phosphorylation of KasB, were constructed by specialized linkage transduction. Biochemical and structural analyses comparing these mutants to the parental strain revealed that both mutant strains had mycolic acids that were shortened by 4-6 carbon atoms and lacked trans-cyclopropanation. Together, these results suggested that in M. tuberculosis, phosphorylation profoundly decreases the condensing activity of KasB. Structural/modeling analyses reveal that Thr334 and Thr336 are located in the vicinity of the catalytic triad, which indicates that phosphorylation of these amino acids would result in loss of enzyme activity. Importantly, the kasB_T334D/T336D phosphomimetic and deletion alleles, in contrast to the kasB_T334A/T336A phosphoablative allele, completely lost acid-fast staining. Moreover, assessing the virulence of these strains indicated that the KasB phosphomimetic mutant was attenuated in both immunodeficient and immunocompetent mice following aerosol infection. This attenuation was characterized by the absence of lung pathology. Overall, these results highlight for the first time the role of

  1. Whole genome-wide transcript profiling to identify differentially expressed genes associated with seed field emergence in two soybean low phytate mutants.

    Science.gov (United States)

    Yuan, Fengjie; Yu, Xiaomin; Dong, Dekun; Yang, Qinghua; Fu, Xujun; Zhu, Shenlong; Zhu, Danhua

    2017-01-18

    Seed germination is important to soybean (Glycine max) growth and development, ultimately affecting soybean yield. A lower seed field emergence has been the main hindrance for breeding soybeans low in phytate. Although this reduction could be overcome by additional breeding and selection, the mechanisms of seed germination in different low phytate mutants remain unknown. In this study, we performed a comparative transcript analysis of two low phytate soybean mutants (TW-1 and TW-1-M), which have the same mutation, a 2 bp deletion in GmMIPS1, but show a significant difference in seed field emergence, TW-1-M was higher than that of TW-1 . Numerous genes analyzed by RNA-Seq showed markedly different expression levels between TW-1-M and TW-1 mutants. Approximately 30,000-35,000 read-mapped genes and ~21000-25000 expressed genes were identified for each library. There were ~3900-9200 differentially expressed genes (DEGs) in each contrast library, the number of up-regulated genes was similar with down-regulated genes in the mutant TW-1and TW-1-M. Gene ontology functional categories of DEGs indicated that the ethylene-mediated signaling pathway, the abscisic acid-mediated signaling pathway, response to hormone, ethylene biosynthetic process, ethylene metabolic process, regulation of hormone levels, and oxidation-reduction process, regulation of flavonoid biosynthetic process and regulation of abscisic acid-activated signaling pathway had high correlations with seed germination. In total, 2457 DEGs involved in the above functional categories were identified. Twenty-two genes with 20 biological functions were the most highly up/down- regulated (absolute value Log2FC >5) in the high field emergence mutant TW-1-M and were related to metabolic or signaling pathways. Fifty-seven genes with 36 biological functions had the greatest expression abundance (FRPM >100) in germination-related pathways. Seed germination in the soybean low phytate mutants is a very complex process

  2. Diversion of phagosome trafficking by pathogenic Rhodococcus equi depends on mycolic acid chain length.

    Science.gov (United States)

    Sydor, Tobias; von Bargen, Kristine; Hsu, Fong-Fu; Huth, Gitta; Holst, Otto; Wohlmann, Jens; Becken, Ulrike; Dykstra, Tobias; Söhl, Kristina; Lindner, Buko; Prescott, John F; Schaible, Ulrich E; Utermöhlen, Olaf; Haas, Albert

    2013-03-01

    Rhodococcus equi is a close relative of Mycobacterium spp. and a facultative intracellular pathogen which arrests phagosome maturation in macrophages before the late endocytic stage. We have screened a transposon mutant library of R. equi for mutants with decreased capability to prevent phagolysosome formation. This screen yielded a mutant in the gene for β-ketoacyl-(acyl carrier protein)-synthase A (KasA), a key enzyme of the long-chain mycolic acid synthesizing FAS-II system. The longest kasA mutant mycolic acid chains were 10 carbon units shorter than those of wild-type bacteria. Coating of non-pathogenic E. coli with purified wild-type trehalose dimycolate reduced phagolysosome formation substantially which was not the case with shorter kasA mutant-derived trehalose dimycolate. The mutant was moderately attenuated in macrophages and in a mouse infection model, but was fully cytotoxic.Whereas loss of KasA is lethal in mycobacteria, R. equi kasA mutant multiplication in broth was normal proving that long-chain mycolic acid compounds are not necessarily required for cellular integrity and viability of the bacteria that typically produce them. This study demonstrates a central role of mycolic acid chain length in diversion of trafficking by R. equi. © 2012 Blackwell Publishing Ltd.

  3. Biosynthesis of myristic acid in luminescent bacteria

    International Nuclear Information System (INIS)

    Byers, D.M.

    1987-01-01

    In vivo pulse-label studies have demonstrated that luminescent bacteria can provide myritic acid (14:0) required for the synthesis of the luciferase substrate myristyl aldehyde. Luminescent wild type Vibrio harveyi incubated with [ 14 C] acetate in a nutrient-depleted medium accumulated substantial tree [ 14 C]fatty acid (up to 20% of the total lipid label). Radio-gas chromatography revealed that > 75% of the labeled fatty acid is 14:0. No free fatty acid was detected in wild type cells labeled prior to the development of bioluminescence in the exponential growth phase, or in a dark mutant of V. harveyi (mutant M17) that requires exogenous 14:0 for light emission. The preferential accumulation of 14:0 was not observed when wild type cells were labeled with [ 14 C]acetate in regular growth medium. Moreover, all V. harveyi strains exhibited similar fatty acid mass compositions regardless of the state of bioluminescence. Since earlier work has shown that a luminescence-related acyltransferase (defective in the M17 mutant) can catalyze the deacylation of fatty acyl-acyl carrier protein in vitro, the present results are consistent with a model in which this enzyme diverts 14:0 to the luminescence system during fatty acid biosynthesis. Under normal conditions, the supply of 14:0 by this pathway is tightly regulated such that bioluminescence development does not significantly alter the total fatty acid composition

  4. An extra early mutant of pigeonpea

    International Nuclear Information System (INIS)

    Ravikesavan, R.; Kalaimagal, T.; Rathnaswamy, R.

    2001-01-01

    The redgram (Cajanus cajan (L.) Huth) variety 'Prabhat DT' was gamma irradiated with 100, 200, 300 and 400 Gy doses. Several mutants have been identified viz., extra early mutants, monostem mutants, obcordifoliate mutants and bi-stigmatic mutants. The extra early mutant was obtained when treated with 100 Gy dose. The mutant was selfed and forwarded from M 2 to M 4 generation. In the M 4 generation the mutant line was raised along with the parental variety. Normal cultural practices were followed and the biometrical observations were recorded. It was observed that for the characters viz., total number of branches per plant, number of pods per plants, seeds per pod, 100 seed weight and seed yield per plant there was no difference between the mutant and parent variety. Whereas, regarding the days to flowering and maturity the mutants were earlier than the parents. The observation was recorded from two hundred plants each. The mutant gives the same yield in 90 days as that of the parent variety in 107 days, which make it an economic mutant

  5. Radio-sensitivity analysis and selection of useful mutants of rape (Brassica napus L.) by gamma irradiation

    International Nuclear Information System (INIS)

    Goh, Eun Jeong; Kim, Wook Jin; Kim, Jin Baek; Kim, Dong Sub; Kim, Sang Hoon; Kang, Si Yong

    2010-01-01

    Rape (Brassica napus L.) plants are one of the major oilseed crops. The main components of rapeseed are oil (35 to 47%) and protein (15 to 32%). For the biodiesel production, the development of a new variety of rape plant with high biomass and/or oleic acid contents is required. In order to determine the optimum dose of gamma-ray irradiation, the rape seeds of cvs. Hanra (Hr), Youngsan (Ys), Tammi (Tm), and Tamra (Tr) were irradiated with a 100 ∼ 4,000 Gy dose range of gamma-rays. Considering the growth factors, the optimum doses were determined to be within the range of 600 ∼ 1,000 Gy for the selection of useful mutant lines. Six-hundred and eighty eight (688) M 2 mutant lines were obtained from 600 ∼ 1,000 Gy gamma-ray-irradiated M 1 plants through selfing. The growth characteristics, leaf shape, early flowering, and flower color were all investigated. The selected mutant numbers of early flowering, leaf shape, and flower color were 34, 52, and 3 from the four cultivars, respectively. These mutant lines will be used for the development of a new variety of rape plant with high biomass and oleic acid contents

  6. The effect of Aspergillus niger mutagenization on citric acid biosynthesis

    Directory of Open Access Journals (Sweden)

    Stanisław Walisch

    2014-08-01

    Full Text Available The industrial A. niger strain producing citric acid was mutagenized with the use of new chemical mutagens: free nitroxyl radicals. Strains of higher citric acid production yield were obtained. Citric acid was produced in a shorter time compared to the initial strain. During 6-12 months of storage most of the strains preserved their positive features which proves that mutants with profitable biotechnological properties were obtained. These mutants are used in industrial process.

  7. Absence-like and tonic seizures in aspartoacylase/attractin double-mutant mice.

    Science.gov (United States)

    Gohma, Hiroshi; Kuramoto, Takashi; Matalon, Reuben; Surendran, Sankar; Tyring, Stephen; Kitada, Kazuhiro; Sasa, Masashi; Serikawa, Tadao

    2007-04-01

    The Spontaneously Epileptic Rat (SER), a double-mutant for tremor and zitter mutations, shows spontaneous occurrences of absence-like and tonic seizures. Several lines of evidence suggest that the combined effect of Aspa and Atrn mutations is the most likely cause of the epileptic phenotype of the SER. To address this issue, we produced a new double-mutant mouse line carrying both homozygous Aspa-knockout and Atrn(mg-3J) mutant alleles. The Aspa/Atrn double-mutant mice exhibited absence-like and tonic seizures that were characterized by the appearance of 5-7 Hz spike-wave-like complexes and low voltage fast waves on EEGs. These results demonstrate directly that the simultaneous loss of the Aspa and Atrn gene functions causes epileptic seizures in the mouse and suggest that both Aspa and Atrn deficiencies might be responsible for epileptic seizures in the SER.

  8. E-2-hexenal promotes susceptibility to Pseudomonas syringae by activating jasmonic acid pathways in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Alessandra eScala

    2013-04-01

    Full Text Available Green Leaf Volatiles (GLVs are C6-molecules - alcohols, aldehydes and esters - produced by plants upon herbivory or during pathogen infection. Exposure to this blend of volatiles induces defence-related responses in neighboring undamaged plants, thus assigning a role to GLVs in regulating plant defences. Here we compared Arabidopsis thaliana ecotype Ler with a hydroperoxide lyase line, hpl1, unable to synthesize GLVs, for susceptibility to Pseudomonas syringae pv. tomato (DC3000. We found that the growth of DC3000 was significantly reduced in the hpl1 mutant. This phenomenon correlated with lower jasmonic acid (JA levels and higher salicylic acid (SA levels in the hpl1 mutant. Furthermore, upon infection, the JA-responsive genes VSP2 and LEC were only slightly or not induced, respectively, in hpl1. This suggests that the reduced growth of DC3000 in hpl1 plants is due to the constraint of JA-dependent responses. Treatment of hpl1 plants with E-2-hexenal, one of the more reactive GLVs, prior to infection with DC3000, resulted in increased growth of DC3000 in hpl1, thus complementing this mutant. Interestingly, the growth of DC3000 also increased in Ler plants treated with E-2-hexenal. This stronger growth was not dependent on the JA-signaling component MYC2, but on ORA59, an integrator of JA and ethylene signaling pathways, and on the production of coronatine by DC3000. GLVs may have multiple effects on plant-pathogen interactions, in this case reducing resistance to P. syringae via JA and ORA59.

  9. Cloning a T-DNA-Linked Phosphate Gene that mediates Salt Tolerance on Mutant of Arabidopsis thaliana

    International Nuclear Information System (INIS)

    Njoroge, N.C; Tremblay, L.; Lefebvre, D.D.

    2006-01-01

    T-DNA insertionally mutagenized seeds of Arabidopsis thaliana were used to unravel genetic mechanisms underlying salt tolerance in plants. Over a period of two weeks, kanamycin homozygous (KK) seeds of the mutant NN143 attain germination levels of 65% and 77% on 175mM Nacl and 300mM mannitol respectively. Under these conditions of osmotic stress, the wild type seeds were incapable of germination. The mutant was also capable of germination on a medium containing 2μM abscisic acid (ABA). After two weeks on 2μM ABA, it attained 100% germination and the wild type did not germinate. The ABA level in the mutant was 40% higher than the wild type. Segregation analysis indicated that salt tolerance in the mutant is T-DNA linked. Genetic analysis of the F1 and F2 generations indicated that the salt tolerance trait in the mutant is dominant. The putative salt tolerance gene of mutant NN143 was cloned by plasmid rescue and sequence data indicated involvement of a protein phosphatase. The possible mechanism underlying salt tolerance in the mutant is discussed.(author)

  10. Gr and hp-1 tomato mutants unveil unprecedented interactions between arbuscular mycorrhizal symbiosis and fruit ripening.

    Science.gov (United States)

    Chialva, Matteo; Zouari, Inès; Salvioli, Alessandra; Novero, Mara; Vrebalov, Julia; Giovannoni, James J; Bonfante, Paola

    2016-07-01

    Systemic responses to an arbuscular mycorrhizal fungus reveal opposite phenological patterns in two tomato ripening mutants depending whether ethylene or light reception is involved. The availability of tomato ripening mutants has revealed many aspects of the genetics behind fleshy fruit ripening, plant hormones and light signal reception. Since previous analyses revealed that arbuscular mycorrhizal symbiosis influences tomato berry ripening, we wanted to test the hypothesis that an interplay might occur between root symbiosis and fruit ripening. With this aim, we screened seven tomato mutants affected in the ripening process for their responsiveness to the arbuscular mycorrhizal fungus Funneliformis mosseae. Following their phenological responses we selected two mutants for a deeper analysis: Green ripe (Gr), deficient in fruit ethylene perception and high-pigment-1 (hp-1), displaying enhanced light signal perception throughout the plant. We investigated the putative interactions between ripening processes, mycorrhizal establishment and systemic effects using biochemical and gene expression tools. Our experiments showed that both mutants, notwithstanding a normal mycorrhizal phenotype at root level, exhibit altered arbuscule functionality. Furthermore, in contrast to wild type, mycorrhization did not lead to a higher phosphate concentration in berries of both mutants. These results suggest that the mutations considered interfere with arbuscular mycorrhiza inducing systemic changes in plant phenology and fruits metabolism. We hypothesize a cross talk mechanism between AM and ripening processes that involves genes related to ethylene and light signaling.

  11. Towards the Identification of New Genes Involved in ABA-Dependent Abiotic Stresses Using Arabidopsis Suppressor Mutants of abh1 Hypersensitivity to ABA during Seed Germination

    Directory of Open Access Journals (Sweden)

    Iwona Szarejko

    2013-06-01

    Full Text Available Abscisic acid plays a pivotal role in the abiotic stress response in plants. Although great progress has been achieved explaining the complexity of the stress and ABA signaling cascade, there are still many questions to answer. Mutants are a valuable tool in the identification of new genes or new alleles of already known genes and in elucidating their role in signaling pathways. We applied a suppressor mutation approach in order to find new components of ABA and abiotic stress signaling in Arabidopsis. Using the abh1 (ABA hypersensitive 1 insertional mutant as a parental line for EMS mutagenesis, we selected several mutants with suppressed hypersensitivity to ABA during seed germination. Here, we present the response to ABA and a wide range of abiotic stresses during the seed germination and young seedling development of two suppressor mutants—soa2 (suppressor of abh1 hypersensitivity to ABA 2 and soa3 (suppressor of abh1 hypersensitivity to ABA 3. Generally, both mutants displayed a suppression of the hypersensitivity of abh1 to ABA, NaCl and mannitol during germination. Both mutants showed a higher level of tolerance than Columbia-0 (Col-0—the parental line of abh1 in high concentrations of glucose. Additionally, soa2 exhibited better root growth than Col-0 in the presence of high ABA concentrations. soa2 and soa3 were drought tolerant and both had about 50% fewer stomata per mm2 than the wild-type but the same number as their parental line—abh1. Taking into account that suppressor mutants had the same genetic background as their parental line—abh1, it was necessary to backcross abh1 with Landsberg erecta four times for the map-based cloning approach. Mapping populations, derived from the cross of abh1 in the Landsberg erecta background with each suppressor mutant, were created. Map based cloning in order to identify the suppressor genes is in progress.

  12. Transcriptomic profiling-based mutant screen reveals three new transcription factors mediating menadione resistance in Neurospora crassa.

    Science.gov (United States)

    Zhu, Jufen; Yu, Xinxu; Xie, Baogui; Gu, Xiaokui; Zhang, Zhenying; Li, Shaojie

    2013-06-01

    To gain insight into the regulatory mechanisms of oxidative stress responses in filamentous fungi, the genome-wide transcriptional response of Neurospora crassa to menadione was analysed by digital gene expression (DGE) profiling, which identified 779 upregulated genes and 576 downregulated genes. Knockout mutants affecting 130 highly-upregulated genes were tested for menadione sensitivity, which revealed that loss of the transcription factor siderophore regulation (SRE) (a transcriptional repressor for siderophore biosynthesis), catatase-3, cytochrome c peroxidase or superoxide dismutase 1 copper chaperone causes hypersensitivity to menadione. Deletion of sre dramatically increased transcription of the siderophore biosynthesis gene ono and the siderophore iron transporter gene sit during menadione stress, suggesting that SRE is required for repression of iron uptake under oxidative stress conditions. Contrary to its phenotype, the sre deletion mutant showed higher transcriptional levels of genes encoding reactive oxygen species (ROS) scavengers than wild type during menadione stress, which implies that the mutant suffers a higher level of oxidative stress than wild type. Uncontrolled iron uptake in the sre mutant might exacerbate cellular oxidative stress. This is the first report of a negative regulator of iron assimilation participating in the fungal oxidative stress response. In addition to SRE, eight other transcription factor genes were also menadione-responsive but their single gene knockout mutants showed wild-type menadione sensitivity. Two of them, named as mit-2 (menadione induced transcription factor-2) and mit-4 (menadione induced transcription factor-4), were selected for double mutant analysis. The double mutant was hypersensitive to menadione. Similarly, the double mutation of mit-2 and sre also had additive effects on menadione sensitivity, suggesting multiple transcription factors mediate oxidative stress resistance in an additive manner

  13. The octadecanoid signalling pathway in plants mediates a response to ultraviolet radiation

    International Nuclear Information System (INIS)

    Conconi, A.; Smerdon, M.J.; Howe, G.A.; Ryan, C.A.

    1996-01-01

    Many plant genes that respond to environmental and developmental changes are regulated by jasmonic acid, which is derived from linolenic acid via the octadecanoid pathway. Linolenic acid is an important fatty-acid constituent of membranes in most plant species and its intracellular levels increase in response to certain signals. Here we report that irradiation of tomato leaves with ultraviolet light induces the expression of several plant defensive genes that are normally activated through the octadecanoid pathway after wounding. The response to ultraviolet light is blocked by an inhibitor of the octadecanoid pathway and it does not occur in a tomato mutant defective in this pathway. The ultraviolet irradiation maximally induces the defence genes at levels where cyclobutane pyrimidine dimer formation, an indicator of DNA damage, is less than 0.2 dimers per gene. Our evidence indicates that this plant defence response to certain wavelengths of ultraviolet radiation requires the activation of the octadecanoid defence signalling pathway. (author)

  14. Hepatitis C Virus Frameshift/Alternate Reading Frame Protein Suppresses Interferon Responses Mediated by Pattern Recognition Receptor Retinoic-Acid-Inducible Gene-I.

    Directory of Open Access Journals (Sweden)

    Seung Bum Park

    Full Text Available Hepatitis C virus (HCV actively evades host interferon (IFN responses but the mechanisms of how it does so are not completely understood. In this study, we present evidence for an HCV factor that contributes to the suppression of retinoic-acid-inducible gene-I (RIG-I-mediated IFN induction. Expression of frameshift/alternate reading frame protein (F/ARFP from HCV -2/+1 frame in Huh7 hepatoma cells suppressed type I IFN responses stimulated by HCV RNA pathogen-associated molecular pattern (PAMP and poly(IC. The suppression occurred independently of other HCV factors; and activation of interferon stimulated genes, TNFα, IFN-λ1, and IFN-λ2/3 was likewise suppressed by HCV F/ARFP. Point mutations in the full-length HCV sequence (JFH1 genotype 2a strain were made to introduce premature termination codons in the -2/+1 reading frame coding for F/ARFP while preserving the original reading frame, which enhanced IFNα and IFNβ induction by HCV. The potentiation of IFN response by the F/ARFP mutations was diminished in Huh7.5 cells, which already have a defective RIG-I, and by decreasing RIG-I expression in Huh7 cells. Furthermore, adding F/ARFP back via trans-complementation suppressed IFN induction in the F/ARFP mutant. The F/ARFP mutants, on the other hand, were not resistant to exogenous IFNα. Finally, HCV-infected human liver samples showed significant F/ARFP antibody reactivity, compared to HCV-uninfected control livers. Therefore, HCV F/ARFP likely cooperates with other viral factors to suppress type I and III IFN induction occurring through the RIG-I signaling pathway. This study identifies a novel mechanism of pattern recognition receptor modulation by HCV and suggests a biological function of the HCV alternate reading frame in the modulation of host innate immunity.

  15. Mutant heterosis in rice

    International Nuclear Information System (INIS)

    1987-01-01

    In the variety TKM6 a high yielding semidwarf mutant has been induced. This TKM6 mutant was used in test crosses with a number of other varieties and mutants to examine the extent of heterosis of dwarfs in rice and to select superior crosses. An excerpt of the published data is given. It appears from the backcross of the mutant with its original variety, that an increase in number of productive tillers occurs in the hybrid, leading to a striking grain yield increase, while the semi-dwarf culm length (the main mutant character) reverts to the normal phenotype. In the cross with IR8 on the other hand, there is only a minimal increase in tiller number but a substantial increase in TGW leading to more than 30% yield increase over the better parent

  16. Nonbehavioral Selection for Pawns, Mutants of PARAMECIUM AURELIA with Decreased Excitability

    Science.gov (United States)

    Schein, Stanley J.

    1976-01-01

    The reversal response in Paramecium aurelia is mediated by calcium which carries the inward current during excitation. Electrophysiological studies indicate that strontium and barium can also carry the inward current. Exposure to high concentrations of barium rapidly paralyzes and later kills wild-type paramecia. Following mutagenesis with nitrosoguanidine, seven mutants which continued to swim in the `high-barium' solution were selected. All of the mutants show decreased reversal behavior, with phenotypes ranging from extremely non-reversing (`extreme' pawns) to nearly wild-type reversal behavior (`partial' pawns). The mutations fall into three complementation groups, identical to the pwA, pwB, and pwC genes of Kung et al. (1975). All of the pwA and pwB mutants withstand longer exposure to barium, the pwB mutants surviving longer than the pwA mutants. Among mutants of each gene, survival is correlated with loss of reversal behavior. Double mutants (A–B, A–C, B–C), identified in the exautogamous progeny of crosses between `partial' mutants, exhibited a more extreme non-reversing phenotype than either of their single-mutant (`partial' pawn) parents.———Inability to reverse could be expected from an alteration in the calcium-activated reversal mechanism or in excitation. A normal calcium-activated structure was demonstrated in all pawns by chlorpromazine treatment. In a separate report (Schein, Bennett and Katz 1976) the results of electrophysiological investigations directly demonstrate decreased excitability in all of the mutants, a decrease due to an altered calcium activation. The studies of the genetics, the survival in barium and the electro-physiology of the pawns demonstrate that the pwA and pwB genes have different effects on calcium activation. PMID:1001878

  17. Stress Tolerance in Doughs of Saccharomyces cerevisiae Trehalase Mutants Derived from Commercial Baker’s Yeast

    Science.gov (United States)

    Shima, Jun; Hino, Akihiro; Yamada-Iyo, Chie; Suzuki, Yasuo; Nakajima, Ryouichi; Watanabe, Hajime; Mori, Katsumi; Takano, Hiroyuki

    1999-01-01

    Accumulation of trehalose is widely believed to be a critical determinant in improving the stress tolerance of the yeast Saccharomyces cerevisiae, which is commonly used in commercial bread dough. To retain the accumulation of trehalose in yeast cells, we constructed, for the first time, diploid homozygous neutral trehalase mutants (Δnth1), acid trehalase mutants (Δath1), and double mutants (Δnth1 ath1) by using commercial baker’s yeast strains as the parent strains and the gene disruption method. During fermentation in a liquid fermentation medium, degradation of intracellular trehalose was inhibited with all of the trehalase mutants. The gassing power of frozen doughs made with these mutants was greater than the gassing power of doughs made with the parent strains. The Δnth1 and Δath1 strains also exhibited higher levels of tolerance of dry conditions than the parent strains exhibited; however, the Δnth1 ath1 strain exhibited lower tolerance of dry conditions than the parent strain exhibited. The improved freeze tolerance exhibited by all of the trehalase mutants may make these strains useful in frozen dough. PMID:10388673

  18. The cellular Mre11 protein interferes with adenovirus E4 mutant DNA replication

    International Nuclear Information System (INIS)

    Mathew, Shomita S.; Bridge, Eileen

    2007-01-01

    Adenovirus type 5 (Ad5) relocalizes and degrades the host DNA repair protein Mre11, and efficiently initiates viral DNA replication. Mre11 associates with Ad E4 mutant DNA replication centers and is important for concatenating viral genomes. We have investigated the role of Mre11 in the E4 mutant DNA replication defect. RNAi-mediated knockdown of Mre11 dramatically rescues E4 mutant DNA replication in cells that do or do not concatenate viral genomes, suggesting that Mre11 inhibits DNA replication independent of genome concatenation. The mediator of DNA damage checkpoint 1 (Mdc1) protein is involved in recruiting and sustaining Mre11 at sites of DNA damage following ionizing radiation. We observe foci formation by Mdc1 in response to viral infection, indicating that this damage response protein is activated. However, knockdown of Mdc1 does not prevent Mre11 from localizing at viral DNA replication foci or rescue E4 mutant DNA replication. Our results are consistent with a model in which Mre11 interferes with DNA replication when it is localized at viral DNA replication foci

  19. ISOELECTRIC FOCUSING OF MEMBRANE PROTEINS OF PROBIOTIC B. COAGULANS AND ITS BACTERIOPHAGE RESISTANT MUTANTS

    Directory of Open Access Journals (Sweden)

    Kavita Rajesh Pandey

    2016-09-01

    Full Text Available Bacteriophages are the most notorious type of infection in the probiotic and dairy fermentations. Two phage resistant mutants viz. B. co PIII and B. co MIII (B. coagulans mutants PIII and MIII obtained in previous studies (Dubey and Vakil, 2010, were further characterized for their protein profile in comparison with the parental probiotic strain –B. coagulans. The cell lysates were subjected to ultra-centrifugation and the purified membrane fractions were resolved using 2D gel electrophoresis. The Isoelectric focussing showed 187, 202 and 154 protein spots for the parental strain, mutant B. co PIII and mutant B. co MIII, respectively. Ten and 18 protein spots were missing as compared to parent for mutants B.co PIII and B.co MIII whereas there were 21 and 14 new spots noticed for these two mutants. Eight membrane proteins present only in the phage sensitive parental culture could be tentatively identified by comparison with the complete proteome of B. coagulans by use of UniprotKB and then CELLO database It is quite likely that some of these identified membrane proteins may be also functioning as receptors for phage adsorption followed by entry of nucleic acid into the phage sensitive host cell.

  20. Structural analysis of Herbaspirillum seropedicae lipid-A and of two mutants defective to colonize maize roots.

    Science.gov (United States)

    Serrato, Rodrigo V; Balsanelli, Eduardo; Sassaki, Guilherme L; Carlson, Russell W; Muszynski, Artur; Monteiro, Rose A; Pedrosa, Fábio O; Souza, Emanuel M; Iacomini, Marcello

    2012-11-01

    Lipid-A was isolated by mild acid hydrolysis from lipopolysaccharides extracted from cells of Herbaspirillum seropedicae, strain SMR1, and from two mutants deficient in the biosynthesis of rhamnose (rmlB⁻ and rmlC⁻). Structural analyzes were carried out using MALDI-TOF and derivatization by per-O-trimethylsilylation followed by GC-MS in order to determine monosaccharide and fatty acid composition. De-O-acylation was also performed to determine the presence of N-linked fatty acids. Lipid-A from H. seropedicae SMR1 showed a major structure comprising 2-amino-2-deoxy-glucopyranose-(1→6)-2-amino-2-deoxy-glucopyranose phosphorylated at C4' and C1 positions, each carrying a unit of 4-amino-4-deoxy-arabinose. C2 and C2' positions were substituted by amide-linked 3-hydroxy-dodecanoic acids. Both rhamnose-defective mutants showed similar structure for their lipid-A moieties, except for the lack of 4-amino-4-deoxy-arabinose units attached to phosphoryl groups. Copyright © 2012 Elsevier B.V. All rights reserved.

  1. Jasmonic acid signaling modulates ozone-induced hypersensitive cell death.

    Science.gov (United States)

    Rao, M V; Lee, H; Creelman, R A; Mullet, J E; Davis, K R

    2000-09-01

    Recent studies suggest that cross-talk between salicylic acid (SA)-, jasmonic acid (JA)-, and ethylene-dependent signaling pathways regulates plant responses to both abiotic and biotic stress factors. Earlier studies demonstrated that ozone (O(3)) exposure activates a hypersensitive response (HR)-like cell death pathway in the Arabidopsis ecotype Cvi-0. We now have confirmed the role of SA and JA signaling in influencing O(3)-induced cell death. Expression of salicylate hydroxylase (NahG) in Cvi-0 reduced O(3)-induced cell death. Methyl jasmonate (Me-JA) pretreatment of Cvi-0 decreased O(3)-induced H(2)O(2) content and SA concentrations and completely abolished O(3)-induced cell death. Cvi-0 synthesized as much JA as did Col-0 in response to O(3) exposure but exhibited much less sensitivity to exogenous Me-JA. Analyses of the responses to O(3) of the JA-signaling mutants jar1 and fad3/7/8 also demonstrated an antagonistic relationship between JA- and SA-signaling pathways in controlling the magnitude of O(3)-induced HR-like cell death.

  2. First discovery of two polyketide synthase genes for mitorubrinic acid and mitorubrinol yellow pigment biosynthesis and implications in virulence of Penicillium marneffei.

    Directory of Open Access Journals (Sweden)

    Patrick C Y Woo

    Full Text Available BACKGROUND: The genome of P. marneffei, the most important thermal dimorphic fungus causing respiratory, skin and systemic mycosis in China and Southeast Asia, possesses 23 polyketide synthase (PKS genes and 2 polyketide synthase nonribosomal peptide synthase hybrid (PKS-NRPS genes, which is of high diversity compared to other thermal dimorphic pathogenic fungi. We hypothesized that the yellow pigment in the mold form of P. marneffei could also be synthesized by one or more PKS genes. METHODOLOGY/PRINCIPAL FINDINGS: All 23 PKS and 2 PKS-NRPS genes of P. marneffei were systematically knocked down. A loss of the yellow pigment was observed in the mold form of the pks11 knockdown, pks12 knockdown and pks11pks12 double knockdown mutants. Sequence analysis showed that PKS11 and PKS12 are fungal non-reducing PKSs. Ultra high performance liquid chromatography-photodiode array detector/electrospray ionization-quadruple time of flight-mass spectrometry (MS and MS/MS analysis of the culture filtrates of wild type P. marneffei and the pks11 knockdown, pks12 knockdown and pks11pks12 double knockdown mutants showed that the yellow pigment is composed of mitorubrinic acid and mitorubrinol. The survival of mice challenged with the pks11 knockdown, pks12 knockdown and pks11pks12 double knockdown mutants was significantly better than those challenged with wild type P. marneffei (P<0.05. There was also statistically significant decrease in survival of pks11 knockdown, pks12 knockdown and pks11pks12 double knockdown mutants compared to wild type P. marneffei in both J774 and THP1 macrophages (P<0.05. CONCLUSIONS/SIGNIFICANCE: The yellow pigment of the mold form of P. marneffei is composed of mitorubrinol and mitorubrinic acid. This represents the first discovery of PKS genes responsible for mitorubrinol and mitorubrinic acid biosynthesis. pks12 and pks11 are probably responsible for sequential use in the biosynthesis of mitorubrinol and mitorubrinic acid

  3. Identification of Vitis vinifera L. grape berry skin color mutants and polyphenolic profile.

    Science.gov (United States)

    Ferreira, Vanessa; Fernandes, Fátima; Pinto-Carnide, Olinda; Valentão, Patrícia; Falco, Virgílio; Martín, Juan Pedro; Ortiz, Jesús María; Arroyo-García, Rosa; Andrade, Paula B; Castro, Isaura

    2016-03-01

    A germplasm set of twenty-five grapevine accessions, forming eleven groups of possible berry skin color mutants, were genotyped with twelve microsatellite loci, being eleven of them identified as true color mutants. The polyphenolic profiling of the confirmed mutant cultivars revealed a total of twenty-four polyphenols, comprising non-colored compounds (phenolic acids, flavan-3-ols, flavonols and a stilbene) and anthocyanins. Results showed differences in the contribution of malvidin-3-O-glucoside to the characteristic Pinot Noir anthocyanins profile. Regarding the two Pique-Poul colored variants, the lighter variant was richer than the darker one in all classes of compounds, excepting anthocyanins. In Moscatel Galego Roxo the F3'H pathway seems to be more active than F3'5'H, resulting in higher amounts of cyanidin, precursor of the cyanidin derivatives. As far as we are aware, this is the first time that a relationship between the content of polyphenolic compounds is established in groups of grape berry skin color mutant cultivars. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Arg156 in the AP2-domain exhibits the highest binding activity among the 20 individuals to the GCC box in BnaERF-B3-hy15, a mutant ERF transcription factor from Brassica napus

    Directory of Open Access Journals (Sweden)

    Jing Zhuang

    2016-10-01

    Full Text Available To develop mutants of the ERF factor with more binding activities to the GCC box, we performed in vitro directed evolution by using DNA shuffling and screened mutants through yeast one-hybrid assay. Here, a series of mutants were obtained and used to reveal key amino acids that induce changes in the DNA binding activity of the BnaERF-B3 protein. With the BnaERF-B3-hy15 as the template, we produced 12 mutants which host individual mutation of potential key residues. We found that amino acid 156 is the key site, and the other 18 mutants host the 18 corresponding individual amino acid residues at site 156. Among the 20 individuals comprising WT (Gly156, Mu3 (Arg156, and 18 mutants with other 18 amino acid residues, Arg156 in the AP2-domain is the amino acid residue with the highest binding activity to the GCC box. The structure of the α-helix in the AP2-domain affects the binding activity. Other residues within AP2-domain modulated binding activity of ERF protein, suggesting that these positions are important for binding activity. Comparison of the mutant and wild-type transcription factors revealed the relationship of protein function and sequence modification. Our result provides a potential useful resource for understanding the trans-activation of ERF proteins.

  5. Physiological and transcriptional response of Lactobacillus casei ATCC 334 to acid stress.

    Science.gov (United States)

    Broadbent, Jeff R; Larsen, Rebecca L; Deibel, Virginia; Steele, James L

    2010-05-01

    This study investigated features of the acid tolerance response (ATR) in Lactobacillus casei ATCC 334. To optimize ATR induction, cells were acid adapted for 10 or 20 min at different pH values (range, 3.0 to 5.0) and then acid challenged at pH 2.0. Adaptation over a broad range of pHs improved acid tolerance, but the highest survival was noted in cells acid adapted for 10 or 20 min at pH 4.5. Analysis of cytoplasmic membrane fatty acids (CMFAs) in acid-adapted cells showed that they had significantly (P L. casei survival at pH 2.5 was improved at least 100-fold by chemical induction of the stringent response or by the addition of 30 mM malate or 30 mM histidine to the acid challenge medium. To our knowledge, this is the first report that intracellular histidine accumulation may be involved in bacterial acid resistance.

  6. Role of a single amino acid substitution of VP3 H142D for increased acid resistance of foot-and-mouth disease virus serotype A.

    Science.gov (United States)

    Biswal, Jitendra K; Das, Biswajit; Sharma, Gaurav K; Khulape, Sagar A; Pattnaik, Bramhadev

    2016-04-01

    Foot-and-mouth disease virus (FMDV) particles lose infectivity due to their dissociation into pentamers at pH value below 6.5. After the uptake of FMDV by receptor-mediated endocytosis, the acid-dependent dissociation process is required for the release of FMDV genome inside endosomes. Nevertheless, dissociation of FMDV particles in mildly acidic conditions renders the inactivated FMD vaccine less effective. To improve the acid stability of inactivated FMD vaccine during the manufacturing process, a serotype A IND 40/2000 (in-use vaccine strain) mutant with increased resistance to acid inactivation was generated through reverse genetics approach. Based upon the earlier reports, the crucial amino acid residue, H142 of VP3 capsid protein was substituted separately to various amino acid residues Arg (R), Phe (F), Ala (A), and Asp (D) on the full-genome length cDNA clone. While the H142 → R or H142 → F or H142 → A substitutions resulted in non-infectious FMDV, H142 → D mutation on VP3 protein (H3142D) resulted in the generation of mutant virus with enhanced resistance to acid-induced inactivation. In addition, H3142D substitution did not alter the replication ability and antigenicity of mutant as compared to the parental virus. However, the virus competition experiments revealed that the H3142D substitution conferred a loss of fitness for the mutant virus. Results from this study demonstrate that the H3142D substitution is the molecular determinant of acid-resistant phenotype in FMDV serotype A.

  7. A novel p53 mutational hotspot in skin tumors from UV-irradiated Xpc mutant mice alters transactivation functions.

    Science.gov (United States)

    Inga, Alberto; Nahari, Dorit; Velasco-Miguel, Susana; Friedberg, Errol C; Resnick, Michael A

    2002-08-22

    A mutation in codon 122 of the mouse p53 gene resulting in a T to L amino acid substitution (T122-->L) is frequently associated with skin cancer in UV-irradiated mice that are both homozygous mutant for the nucleotide excision repair (NER) gene Xpc (Xpc(-/-)) and hemizygous mutant for the p53 gene. We investigated the functional consequences of the mouse T122-->L mutation when expressed either in mammalian cells or in the yeast Saccharomyces cerevisiae. Similar to a non-functional allele, high expression of the T122-->L allele in p53(-/-) mouse embryo fibroblasts and human Saos-2 cells failed to suppress growth. However, the T122-->L mutant p53 showed wild-type transactivation levels with Bax and MDM2 promoters when expressed in either cell type and retained transactivation of the p21 and the c-Fos promoters in one cell line. Using a recently developed rheostatable p53 induction system in yeast we assessed the T122-->L transactivation capacity at low levels of protein expression using 12 different p53 response elements (REs). Compared to wild-type p53 the T122-->L protein manifested an unusual transactivation pattern comprising reduced and enhanced activity with specific REs. The high incidence of the T122-->L mutant allele in the Xpc(-/-) background suggests that both genetic and epigenetic conditions may facilitate the emergence of particular functional p53 mutations. Furthermore, the approach that we have taken also provides for the dissection of functions that may be retained in many p53 tumor alleles.

  8. Protein-membrane interaction and fatty acid transfer from intestinal fatty acid-binding protein to membranes. Support for a multistep process.

    Science.gov (United States)

    Falomir-Lockhart, Lisandro J; Laborde, Lisandro; Kahn, Peter C; Storch, Judith; Córsico, Betina

    2006-05-19

    Fatty acid transfer from intestinal fatty acid-binding protein (IFABP) to phospholipid membranes occurs during protein-membrane collisions. Electrostatic interactions involving the alpha-helical "portal" region of the protein have been shown to be of great importance. In the present study, the role of specific lysine residues in the alpha-helical region of IFABP was directly examined. A series of point mutants in rat IFABP was engineered in which the lysine positive charges in this domain were eliminated or reversed. Using a fluorescence resonance energy transfer assay, we analyzed the rates and mechanism of fatty acid transfer from wild type and mutant proteins to acceptor membranes. Most of the alpha-helical domain mutants showed slower absolute fatty acid transfer rates to zwitterionic membranes, with substitution of one of the lysines of the alpha2 helix, Lys27, resulting in a particularly dramatic decrease in the fatty acid transfer rate. Sensitivity to negatively charged phospholipid membranes was also reduced, with charge reversal mutants in the alpha2 helix the most affected. The results support the hypothesis that the portal region undergoes a conformational change during protein-membrane interaction, which leads to release of the bound fatty acid to the membrane and that the alpha2 segment is of particular importance in the establishment of charge-charge interactions between IFABP and membranes. Cross-linking experiments with a phospholipid-photoactivable reagent underscored the importance of charge-charge interactions, showing that the physical interaction between wild-type intestinal fatty acid-binding protein and phospholipid membranes is enhanced by electrostatic interactions. Protein-membrane interactions were also found to be enhanced by the presence of ligand, suggesting different collisional complex structures for holo- and apo-IFABP.

  9. Biological response to purification and acid functionalization of carbon nanotubes

    Science.gov (United States)

    Figarol, Agathe; Pourchez, Jérémie; Boudard, Delphine; Forest, Valérie; Tulliani, Jean-Marc; Lecompte, Jean-Pierre; Cottier, Michèle; Bernache-Assollant, Didier; Grosseau, Philippe

    2014-07-01

    Acid functionalization has been considered as an easy way to enhance the dispersion and biodegradation of carbon nanotubes (CNT). However, inconsistencies between toxicity studies of acid functionalized CNT remain unexplained. This could be due to a joint effect of the main physicochemical modifications resulting from an acid functionalization: addition of surface acid groups and purification from catalytic metallic impurities. In this study, the impact on CNT biotoxicity of these two physiochemical features was assessed separately. The in vitro biological response of RAW 264.7 macrophages was evaluated after exposure to 15-240 µg mL-1 of two types of multi-walled CNT. For each type of CNT (small: 20 nm diameter, and big: 90 nm diameter), three different surface chemical properties were studied (total of six CNT samples): pristine, acid functionalized and desorbed. Desorbed CNT were purified by the acid functionalization but presented a very low amount of surface acid groups due to a thermal treatment under vacuum. A Janus effect of acid functionalization with two opposite impacts is highlighted. The CNT purification decreased the overall toxicity, while the surface acid groups intensified it when present at a specific threshold. These acid groups especially amplified the pro-inflammatory response. The threshold mechanism which seemed to regulate the impact of acid groups should be further studied to determine its value and potential link to the other physicochemical state of the CNT. The results suggest that, for a safer-design approach, the benefit-risk balance of an acid functionalization has to be considered, depending on the CNT primary state of purification. Further research should be conducted in this direction.

  10. A two component system is involved in acid adaptation of Lactobacillus delbrueckii subsp. bulgaricus.

    Science.gov (United States)

    Cui, Yanhua; Liu, Wei; Qu, Xiaojun; Chen, Zhangting; Zhang, Xu; Liu, Tong; Zhang, Lanwei

    2012-05-20

    The Gram-positive bacterium Lactobacillus delbrueckii subsp. bulgaricus is of vital importance to the food industry, especially to the dairy industry. Two component systems (TCSs) are one of the most important mechanisms for environmental sensing and signal transduction in the majority of Gram-positive and Gram-negative bacteria. A typical TCS consists of a histidine protein kinase (HPK) and a cytoplasmic response regulator (RR). To investigate the functions of TCSs during acid adaptation in L. bulgaricus, we used quantitative PCR to reveal how TCSs expression changes during acid adaptation. Two TCSs (JN675228/JN675229 and JN675230/JN675231) and two HPKs (JN675236 and JN675240) were induced during acid adaptation. These TCSs were speculated to be related with the acid adaptation ability of L. bulgaricus. The mutants of JN675228/JN675229 were constructed in order to investigate the functions of JN675228/JN675229. The mutants showed reduced acid adaptation compared to that of wild type, and the complemented strains were similar to the wild-type strain. These observations suggested that JN675228 and JN675229 were involved in acid adaptation in L. bulgaricus. The interaction between JN675228 and JN675229 was identified by means of yeast two-hybrid system. The results indicated there is interaction between JN675228 and JN675229. Crown Copyright © 2011. Published by Elsevier GmbH. All rights reserved.

  11. Mutations in type 3 reovirus that determine binding to sialic acid are contained in the fibrous tail domain of viral attachment protein sigma1.

    Science.gov (United States)

    Chappell, J D; Gunn, V L; Wetzel, J D; Baer, G S; Dermody, T S

    1997-03-01

    The reovirus attachment protein, sigma1, determines numerous aspects of reovirus-induced disease, including viral virulence, pathways of spread, and tropism for certain types of cells in the central nervous system. The sigma1 protein projects from the virion surface and consists of two distinct morphologic domains, a virion-distal globular domain known as the head and an elongated fibrous domain, termed the tail, which is anchored into the virion capsid. To better understand structure-function relationships of sigma1 protein, we conducted experiments to identify sequences in sigma1 important for viral binding to sialic acid, a component of the receptor for type 3 reovirus. Three serotype 3 reovirus strains incapable of binding sialylated receptors were adapted to growth in murine erythroleukemia (MEL) cells, in which sialic acid is essential for reovirus infectivity. MEL-adapted (MA) mutant viruses isolated by serial passage in MEL cells acquired the capacity to bind sialic acid-containing receptors and demonstrated a dependence on sialic acid for infection of MEL cells. Analysis of reassortant viruses isolated from crosses of an MA mutant virus and a reovirus strain that does not bind sialic acid indicated that the sigma1 protein is solely responsible for efficient growth of MA mutant viruses in MEL cells. The deduced sigma1 amino acid sequences of the MA mutant viruses revealed that each strain contains a substitution within a short region of sequence in the sigma1 tail predicted to form beta-sheet. These studies identify specific sequences that determine the capacity of reovirus to bind sialylated receptors and suggest a location for a sialic acid-binding domain. Furthermore, the results support a model in which type 3 sigma1 protein contains discrete receptor binding domains, one in the head and another in the tail that binds sialic acid.

  12. Structure of the lamin A/C R482W mutant responsible for dominant familial partial lipodystrophy (FPLD)

    Energy Technology Data Exchange (ETDEWEB)

    Magracheva, Eugenia; Kozlov, Serguei; Stewart, Colin L.; Wlodawer, Alexander; Zdanov, Alexander; (NCI)

    2009-08-07

    Proteins of the A-type lamin family, which consists of two members, lamin A and lamin C, are the major components of a thin proteinaceous filamentous meshwork, the lamina, that underlies the inner nuclear membrane. A-type lamins have recently become the focus of extensive functional studies as a consequence of the linking of at least eight congenital diseases to mutations in the lamin A/C gene (LMNA). This spectrum of pathologies, which mostly manifest themselves as dominant traits, includes muscle dystrophies, dilated cardiomyopathies, the premature aging syndrome Hutchinson-Guilford progeria and familial partial lipodystrophy (FPLD). The crystal structure of the lamin A/C mutant R482W, a variant that causes FPLD, has been determined at 1.5 {angstrom} resolution. A completely novel aggregation state of the C-terminal globular domain and the position of the mutated amino-acid residue suggest means by which the mutation may affect lamin A/C-protein and protein-DNA interactions.

  13. Evaluation of Drought response in Some Rice Mutant Lines Using Stress Tolerance Indices

    Directory of Open Access Journals (Sweden)

    H Aminpanah

    2018-05-01

    Full Text Available Introduction Drought is a major problem that limits the adoption of high-yielding rice varieties in drought-prone rainfed rice environments. To improve crop productivity, it is necessary to understand the mechanism of plant responses to drought conditions with the ultimate goal of improving crop performance in the vast areas of the world where rainfall is limiting or unreliable. Safaei Chaeikar et al. (2008 reported that MP, GMP, HM and STI indices, which showed the highest correlation with grain yield under both optimal and stress conditions, can be used as the best indices to introduce drought-tolerant genotypes in rice breeding programs. They also were introduced Nemat, Sepidrood, IR64, IR50 and Bejar genotypes as tolerant varieties. The present study was conducted to determine how drought affects grain yield in rice mutant lines and also to test this hypothesis in order to identify the most suitable indices/genotypes. Materials and Methods A field trial was conducted at Iranian Rice Research Centers in North of Iran, Rasht (latitude 37◦28', longitude 49◦28'E and altitude 7m below the sea level, during the 2014-2015 growing season. The seeds were sown in a nursery on the 10 May and 25 day old seedlings were transplanted to the field. Two separately experiment was carried out under reproductive stage drought stress and controlled conditions based on randomized complete block design with three replications, in four-row plots of three m length. Transplanting was done using 1 seedling per hill; at hill spacing of 25 cm × 25 cm. 18 rice genotypes were consisted 14 M5 mutant lines and their four parental cultivars. Results and Discussion Analysis of variance indicated significant effects of drought stress, genotype and interaction effects of two factors on grain yield, plant height, flag leaf area, tiller number and grain fertility percentage. Drought stress at reproductive stage caused reduction in grain yield (59.47%, grain fertility

  14. Preconception folic acid use modulates estradiol and follicular responses to ovarian stimulation.

    Science.gov (United States)

    Twigt, John M; Hammiche, Fatima; Sinclair, Kevin D; Beckers, Nicole G; Visser, Jenny A; Lindemans, Jan; de Jong, Frank H; Laven, Joop S E; Steegers-Theunissen, Régine P

    2011-02-01

    Folate is a methyl donor. Availability of folate affects DNA methylation profiles and thereby gene expression profiles. We investigated the effects of low-dose folic acid use (0.4 mg/d) on the ovarian response to mild and conventional ovarian stimulation in women. In a randomized trial among subfertile women, 24 and 26 subjects received conventional and mild ovarian stimulation, respectively. Blood samples were taken during the early follicular phase of the cycle prior to treatment and on the day of human chorionic gonadotropin administration for determination of serum total homocysteine, anti-Müllerian hormone (AMH), estradiol, and folate. Folic acid use was validated by questionnaire and serum folate levels. Preovulatory follicles were visualized, counted, and diameters recorded using transvaginal ultrasound. The relation between folic acid use and ovarian response was assessed using linear regression analysis. Folic acid use modified the ovarian response to ovarian stimulation treatment. The estradiol response was higher in nonfolic acid users receiving conventional treatment [β(interaction) = 0.52 (0.07-0.97); P = 0.03], and this effect was independent of serum AMH levels and the preovulatory follicle count. In the conventional treatment, the mean follicle number was also greater in nonusers compared with the users group (14.1 vs. 8.9, P = 0.03). Low-dose folic acid use attenuates follicular and endocrine responses to conventional stimulation, independent of AMH and follicle count. The nature of this observation suggests that the effect of folic acid is most prominent during early follicle development, affecting immature follicles. Deleterious effects of folate deficiency, like DNA hypomethylation and oxidative stress, can help to explain our observations.

  15. Phosphorylation of InhA inhibits mycolic acid biosynthesis and growth of Mycobacterium tuberculosis

    Energy Technology Data Exchange (ETDEWEB)

    Molle, Virginie; Gulten, Gulcin; Vilchèze, Catherine; Veyron-Churlet, Romain; Zanella-Cléon, Isabelle; Sacchettini, James C.; Jacobs, Jr, William R.; Kremer, Laurent (CNRS-UMR); (Einstein); (TAM)

    2011-08-24

    The remarkable survival ability of Mycobacterium tuberculosis in infected hosts is related to the presence of cell wall-associated mycolic acids. Despite their importance, the mechanisms that modulate expression of these lipids in response to environmental changes are unknown. Here we demonstrate that the enoyl-ACP reductase activity of InhA, an essential enzyme of the mycolic acid biosynthetic pathway and the primary target of the anti-tubercular drug isoniazid, is controlled via phosphorylation. Thr-266 is the unique kinase phosphoacceptor, both in vitro and in vivo. The physiological relevance of Thr-266 phosphorylation was demonstrated using inhA phosphoablative (T266A) or phosphomimetic (T266D/E) mutants. Enoyl reductase activity was severely impaired in the mimetic mutants in vitro, as a consequence of a reduced binding affinity to NADH. Importantly, introduction of inhA{_}T266D/E failed to complement growth and mycolic acid defects of an inhA-thermosensitive Mycobacterium smegmatis strain, in a similar manner to what is observed following isoniazid treatment. This study suggests that phosphorylation of InhA may represent an unusual mechanism that allows M. tuberculosis to regulate its mycolic acid content, thus offering a new approach to future anti-tuberculosis drug development.

  16. Phosphorylation of InhA inhibits mycolic acid biosynthesis and growth of Mycobacterium tuberculosis.

    Science.gov (United States)

    Molle, Virginie; Gulten, Gulcin; Vilchèze, Catherine; Veyron-Churlet, Romain; Zanella-Cléon, Isabelle; Sacchettini, James C; Jacobs, William R; Kremer, Laurent

    2010-12-01

    The remarkable survival ability of Mycobacterium tuberculosis in infected hosts is related to the presence of cell wall-associated mycolic acids. Despite their importance, the mechanisms that modulate expression of these lipids in response to environmental changes are unknown. Here we demonstrate that the enoyl-ACP reductase activity of InhA, an essential enzyme of the mycolic acid biosynthetic pathway and the primary target of the anti-tubercular drug isoniazid, is controlled via phosphorylation. Thr-266 is the unique kinase phosphoacceptor, both in vitro and in vivo. The physiological relevance of Thr-266 phosphorylation was demonstrated using inhA phosphoablative (T266A) or phosphomimetic (T266D/E) mutants. Enoyl reductase activity was severely impaired in the mimetic mutants in vitro, as a consequence of a reduced binding affinity to NADH. Importantly, introduction of inhA_T266D/E failed to complement growth and mycolic acid defects of an inhA-thermosensitive Mycobacterium smegmatis strain, in a similar manner to what is observed following isoniazid treatment. This study suggests that phosphorylation of InhA may represent an unusual mechanism that allows M. tuberculosis to regulate its mycolic acid content, thus offering a new approach to future anti-tuberculosis drug development. © 2010 Blackwell Publishing Ltd.

  17. Improved sugar yields from biomass sorghum feedstocks: comparing low-lignin mutants and pretreatment chemistries.

    Science.gov (United States)

    Godin, Bruno; Nagle, Nick; Sattler, Scott; Agneessens, Richard; Delcarte, Jérôme; Wolfrum, Edward

    2016-01-01

    For biofuel production processes to be economically efficient, it is essential to maximize the production of monomeric carbohydrates from the structural carbohydrates of feedstocks. One strategy for maximizing carbohydrate production is to identify less recalcitrant feedstock cultivars by performing some type of experimental screening on a large and diverse set of candidate materials, or by identifying genetic modifications (random or directed mutations or transgenic plants) that provide decreased recalcitrance. Economic efficiency can also be increased using additional pretreatment processes such as deacetylation, which uses dilute NaOH to remove the acetyl groups of hemicellulose prior to dilute acid pretreatment. In this work, we used a laboratory-scale screening tool that mimics relevant thermochemical pretreatment conditions to compare the total sugar yield of three near-isogenic brown midrib ( bmr ) mutant lines and the wild-type (WT) sorghum cultivar. We then compared results obtained from the laboratory-scale screening pretreatment assay to a large-scale pretreatment system. After pretreatment and enzymatic hydrolysis, the bmr mutants had higher total sugar yields than the WT sorghum cultivar. Increased pretreatment temperatures increased reactivity for all sorghum samples reducing the differences observed at lower reaction temperatures. Deacetylation prior to dilute acid pretreatment increased the total sugar yield for all four sorghum samples, and reduced the differences in total sugar yields among them, but solubilized a sizable fraction of the non-structural carbohydrates. The general trends of increased total sugar yield in the bmr mutant compared to the WT seen at the laboratory scale were observed at the large-scale system. However, in the larger reactor system, the measured total sugar yields were lower and the difference in total sugar yield between the WT and bmr sorghum was larger. Sorghum bmr mutants, which have a reduced lignin content showed

  18. Renal acidification responses to respiratory acid-base disorders.

    Science.gov (United States)

    Madias, Nicolaos E

    2010-01-01

    Respiratory acid-base disorders are those abnormalities in acid-base equilibrium that are expressed as primary changes in the arterial carbon dioxide tension (PaCO2). An increase in PaCO2 (hypercapnia) acidifies body fluids and initiates the acid-base disturbance known as respiratory acidosis. By contrast, a decrease in PaCO2 (hypocapnia) alkalinizes body fluids and initiates the acid-base disturbance known as respiratory alkalosis. The impact on systemic acidity of these primary changes in PaCO2 is ameliorated by secondary, directional changes in plasma [HCO3¯] that occur in 2 stages. Acutely, hypercapnia or hypocapnia yields relatively small changes in plasma [HCO3¯] that originate virtually exclusively from titration of the body's nonbicarbonate buffers. During sustained hypercapnia or hypocapnia, much larger changes in plasma [HCO3¯] occur that reflect adjustments in renal acidification mechanisms. Consequently, the deviation of systemic acidity from normal is smaller in the chronic forms of these disorders. Here we provide an overview of the renal acidification responses to respiratory acid-base disorders. We also identify gaps in knowledge that require further research.

  19. Methylammonium-resistant mutants of Nicotiana plumbaginifolia are affected in nitrate transport.

    Science.gov (United States)

    Godon, C; Krapp, A; Leydecker, M T; Daniel-Vedele, F; Caboche, M

    1996-02-25

    This work reports the isolation and preliminary characterization of Nicotiana plumbaginifolia mutants resistant to methylammonium. Nicotiana plumbaginifolia plants cannot grow on low levels of nitrate in the presence of methylammonium. Methylammonium is not used as a nitrogen source, although it can be efficiently taken up by Nicotiana plumbaginifolia cells and converted into methylglutamine, an analog of glutamine. Glutamine is known to repress the expression of the enzymes that mediate the first two steps in the nitrate assimilatory pathway, nitrate reductase (NR) and nitrite reductase (NiR). Methylammonium has therefore been used, in combination with low concentrations of nitrate, as a selective agent in order to screen for mutants in which the nitrate pathway is de-repressed. Eleven semi-dominant mutants, all belonging to the same complementation group, were identified. The mutant showing the highest resistance to methylammonium was not affected either in the utilization of ammonium, accumulation of methylammonium or in glutamine synthase activity. A series of experiments showed that utilization of nitrite by the wild-type and the mutant was comparable, in the presence or the absence of methylammonium, thus suggesting that the mutation specifically affected nitrate transport or reduction. Although NR mRNA levels were less repressed by methylammonium treatment of the wild-type than the mutant, NR activities of the mutant remained comparable with or without methylammonium, leading to the hypothesis that modified expression of NR is probably not responsible for resistance to methylammonium. Methylammonium inhibited nitrate uptake in the wild-type but had only a limited effect in the mutant. The implications of these results are discussed.

  20. Selection of gamma-ray induced salt tolerant rice mutants by in vitro mutagenesis

    International Nuclear Information System (INIS)

    Kim, Dong Sub; Chun, Jae Beom; Lee, Kyung Jun; Kim, Jin Baek; Kim, Sang Hoon; Yun, Song Jong; Kang, Si Yong

    2010-01-01

    The present study had been performed to select the salt tolerant rice mutant lines through an in vivo and in vitro mutagenesis with a gamma-ray. The physiological responses such as MDA and chlorophyll of the selected salt mutant lines were investigated under salt stress. For the selection of the salt tolerant rice mutants by in vitro mutagenesis with gamma-ray, we conducted a second selection procedure with 1,500 mutant lines induced from the original cv. Dongan (wild-type, WT): Ist, selection under a nutrient solution with 171 mM NaCI: 2nd, selection under in vitro conditions. Based on a growth comparison of the entries, out of mutant lines, the putative 2 salt tolerant rice mutant lines, ST-495 and ST-532, were selected. The 2 ST-lines had a lower malonaldehyde (MDA) contents than wild-type (WT) during salt stress. The survival rate of the WT, ST-495 and ST-532 were 36.6%, 70% and 50% in 171 mM NaCI, respectively. The chlorophyll and carotenoid contents were decreased more in a WT plant than the two selected mutant lines. These rice mutant lines will be released for cultivation at the reclaimed land and used as a control plot for genetic research about salt tolerance

  1. Selection of gamma-ray induced salt tolerant rice mutants by in vitro mutagenesis

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Dong Sub; Chun, Jae Beom; Lee, Kyung Jun; Kim, Jin Baek; Kim, Sang Hoon; Yun, Song Jong; Kang, Si Yong [Korea Atomic Energy Research Institute, Jeongeup (Korea, Republic of)

    2010-06-15

    The present study had been performed to select the salt tolerant rice mutant lines through an in vivo and in vitro mutagenesis with a gamma-ray. The physiological responses such as MDA and chlorophyll of the selected salt mutant lines were investigated under salt stress. For the selection of the salt tolerant rice mutants by in vitro mutagenesis with gamma-ray, we conducted a second selection procedure with 1,500 mutant lines induced from the original cv. Dongan (wild-type, WT): Ist, selection under a nutrient solution with 171 mM NaCI: 2nd, selection under in vitro conditions. Based on a growth comparison of the entries, out of mutant lines, the putative 2 salt tolerant rice mutant lines, ST-495 and ST-532, were selected. The 2 ST-lines had a lower malonaldehyde (MDA) contents than wild-type (WT) during salt stress. The survival rate of the WT, ST-495 and ST-532 were 36.6%, 70% and 50% in 171 mM NaCI, respectively. The chlorophyll and carotenoid contents were decreased more in a WT plant than the two selected mutant lines. These rice mutant lines will be released for cultivation at the reclaimed land and used as a control plot for genetic research about salt tolerance.

  2. The Citrus transcription factor, CitERF13, regulates citric acid accumulation via a protein-protein interaction with the vacuolar proton pump, CitVHA-c4.

    Science.gov (United States)

    Li, Shao-jia; Yin, Xue-ren; Xie, Xiu-lan; Allan, Andrew C; Ge, Hang; Shen, Shu-ling; Chen, Kun-song

    2016-02-03

    Organic acids are essential to fruit flavor. The vacuolar H(+) transporting adenosine triphosphatase (V-ATPase) plays an important role in organic acid transport and accumulation. However, less is known of V-ATPase interacting proteins and their relationship with organic acid accumulation. The relationship between V-ATPase and citric acid was investigated, using the citrus tangerine varieties 'Ordinary Ponkan (OPK)' and an early maturing mutant 'Zaoshu Ponkan (ZPK)'. Five V-ATPase genes (CitVHA) were predicted as important to citric acid accumulation. Among the genes, CitVHA-c4 was observed, using a yeast two-hybrid screen, to interact at the protein level with an ethylene response factor, CitERF13. This was verified using bimolecular fluorescence complementation assays. A similar interaction was also observed between Arabidopsis AtERF017 (a CitERF13 homolog) and AtVHA-c4 (a CitVHA-c4 homolog). A synergistic effect on citric acid levels was observed between V-ATPase proteins and interacting ERFs when analyzed using transient over-expression in tobacco and Arabidopsis mutants. Furthermore, the transcript abundance of CitERF13 was concomitant with CitVHA-c4. CitERF13 or AtERF017 over-expression leads to significant citric acid accumulation. This accumulation was abolished in an AtVHA-c4 mutant background. ERF-VHA interactions appear to be involved in citric acid accumulation, which was observed in both citrus and Arabidopsis.

  3. The transcription factor ABI4 Is required for the ascorbic acid-dependent regulation of growth and regulation of jasmonate-dependent defense signaling pathways in Arabidopsis.

    Science.gov (United States)

    Kerchev, Pavel I; Pellny, Till K; Vivancos, Pedro Diaz; Kiddle, Guy; Hedden, Peter; Driscoll, Simon; Vanacker, Hélène; Verrier, Paul; Hancock, Robert D; Foyer, Christine H

    2011-09-01

    Cellular redox homeostasis is a hub for signal integration. Interactions between redox metabolism and the ABSCISIC ACID-INSENSITIVE-4 (ABI4) transcription factor were characterized in the Arabidopsis thaliana vitamin c defective1 (vtc1) and vtc2 mutants, which are defective in ascorbic acid synthesis and show a slow growth phenotype together with enhanced abscisic acid (ABA) levels relative to the wild type (Columbia-0). The 75% decrease in the leaf ascorbate pool in the vtc2 mutants was not sufficient to adversely affect GA metabolism. The transcriptome signatures of the abi4, vtc1, and vtc2 mutants showed significant overlap, with a large number of transcription factors or signaling components similarly repressed or induced. Moreover, lincomycin-dependent changes in LIGHT HARVESTING CHLOROPHYLL A/B BINDING PROTEIN 1.1 expression were comparable in these mutants, suggesting overlapping participation in chloroplast to nucleus signaling. The slow growth phenotype of vtc2 was absent in the abi4 vtc2 double mutant, as was the sugar-insensitive phenotype of the abi4 mutant. Octadecanoid derivative-responsive AP2/ERF-domain transcription factor 47 (ORA47) and AP3 (an ABI5 binding factor) transcripts were enhanced in vtc2 but repressed in abi4 vtc2, suggesting that ABI4 and ascorbate modulate growth and defense gene expression through jasmonate signaling. We conclude that low ascorbate triggers ABA- and jasmonate-dependent signaling pathways that together regulate growth through ABI4. Moreover, cellular redox homeostasis exerts a strong influence on sugar-dependent growth regulation.

  4. Drosophila fatty acid taste signals through the PLC pathway in sugar-sensing neurons.

    Directory of Open Access Journals (Sweden)

    Pavel Masek

    Full Text Available Taste is the primary sensory system for detecting food quality and palatability. Drosophila detects five distinct taste modalities that include sweet, bitter, salt, water, and the taste of carbonation. Of these, sweet-sensing neurons appear to have utility for the detection of nutritionally rich food while bitter-sensing neurons signal toxicity and confer repulsion. Growing evidence in mammals suggests that taste for fatty acids (FAs signals the presence of dietary lipids and promotes feeding. While flies appear to be attracted to fatty acids, the neural basis for fatty acid detection and attraction are unclear. Here, we demonstrate that a range of FAs are detected by the fly gustatory system and elicit a robust feeding response. Flies lacking olfactory organs respond robustly to FAs, confirming that FA attraction is mediated through the gustatory system. Furthermore, flies detect FAs independent of pH, suggesting the molecular basis for FA taste is not due to acidity. We show that low and medium concentrations of FAs serve as an appetitive signal and they are detected exclusively through the same subset of neurons that sense appetitive sweet substances, including most sugars. In mammals, taste perception of sweet and bitter substances is dependent on phospholipase C (PLC signaling in specialized taste buds. We find that flies mutant for norpA, a Drosophila ortholog of PLC, fail to respond to FAs. Intriguingly, norpA mutants respond normally to other tastants, including sucrose and yeast. The defect of norpA mutants can be rescued by selectively restoring norpA expression in sweet-sensing neurons, corroborating that FAs signal through sweet-sensing neurons, and suggesting PLC signaling in the gustatory system is specifically involved in FA taste. Taken together, these findings reveal that PLC function in Drosophila sweet-sensing neurons is a conserved molecular signaling pathway that confers attraction to fatty acids.

  5. Dipeptidyl peptidase IV is involved in the cellulose-responsive induction of cellulose biomass-degrading enzyme genes in Aspergillus aculeatus.

    Science.gov (United States)

    Tani, Shuji; Yuki, Shota; Kunitake, Emi; Sumitani, Jun-Ichi; Kawaguchi, Takashi

    2017-06-01

    We screened for factors involved in the cellulose-responsive induction of cellulose biomass-degrading enzyme genes from approximately 12,000 Aspergillus aculeatus T-DNA insertion mutants harboring a transcriptional fusion between the FIII-avicelase gene (cbhI) promoter and the orotidine 5'-monophosphate decarboxylase gene. Analysis of 5-fluoroorodic acid (5-FOA) sensitivity, cellulose utilization, and cbhI expression of the mutants revealed that a mutant harboring T-DNA at the dipeptidyl peptidase IV (dppIV) locus had acquired 5-FOA resistance and was deficient in cellulose utilization and cbhI expression. The deletion of dppIV resulted in a significant reduction in the cellulose-responsive expression of both cbhI as well as genes controlled by XlnR-independent and XlnR-dependent signaling pathways at an early phase in A. aculeatus. In contrast, the dppIV deletion did not affect the xylose-responsive expression of genes under the control of XlnR. These results demonstrate that DppIV participates in cellulose-responsive induction in A. aculeatus.

  6. Gamma-radiation Mutagenesis in Genetically Unstable Barley Mutants. Pt. 2. Comparison of Various Mutants

    International Nuclear Information System (INIS)

    Balchiuniene, L.

    1995-01-01

    Spontaneous and gamma-induced mutability was compared in two groups of genetically unstable barley ear structure mutants - tweaky spike (tw) and branched ear (be). Instability in different loci causes different levels of spontaneous and gamma-induced mutability. A high spontaneous level of chlorophyll mutations is peculiar to be-ust mutants. It is suggested that the high level of induced chlorophyll mutations in allelic tw mutants is a result of better surviving of chlorophyll mutation carriers in the genotypical-physiological environment created by mutant tw alleles. (author). 6 refs., 2 tabs

  7. Lipid composition of cAMP-dependent protein kinase mutants of Aspergillus niger.

    Science.gov (United States)

    Jernejc, Katarina; Bencina, Mojca

    2003-08-29

    Lipid composition of cAMP-dependent protein kinase (PKA) Aspergillus niger mutants with overexpressed or deleted genes for either regulatory and/or the catalytic subunit of PKA was analyzed. Disruption of the gene encoding the PKA regulatory subunit resulted in 20% less total lipids, 30% less neutral lipids, four times more glycolipids and two-fold higher triacylglycerol lipase activity compared to the control strain. Concomitantly a five-fold decrease in phosphatidylcholine, accompanied with 1.5-, 1.8- and 2.8-fold increases in phosphatidylethanolamine, lysophosphatidylethanolamine and phosphatidylinositol, was determined, respectively. The lack of PKA activity, due to the disruption of a gene encoding the PKA catalytic subunit, resulted in a 1.6-times increase in total lipids with two times more neutral lipids associated with lower triacylglycerol lipase activity and a decrease in phospholipids. The mutants with unrestricted PKA activity synthesized twice as much citric acid as the control strain and three times more than strains lacking PKA activity. The results indicate the involvement of cAMP-mediated PKA activity in regulation of lipid biosynthesis as well as citric acid synthesis.

  8. Effects of fatty acid activation on photosynthetic production of fatty acid-based biofuels in Synechocystis sp. PCC6803

    Directory of Open Access Journals (Sweden)

    Gao Qianqian

    2012-03-01

    Full Text Available Abstract Background Direct conversion of solar energy and carbon dioxide to drop in fuel molecules in a single biological system can be achieved from fatty acid-based biofuels such as fatty alcohols and alkanes. These molecules have similar properties to fossil fuels but can be produced by photosynthetic cyanobacteria. Results Synechocystis sp. PCC6803 mutant strains containing either overexpression or deletion of the slr1609 gene, which encodes an acyl-ACP synthetase (AAS, have been constructed. The complete segregation and deletion in all mutant strains was confirmed by PCR analysis. Blocking fatty acid activation by deleting slr1609 gene in wild-type Synechocystis sp. PCC6803 led to a doubling of the amount of free fatty acids and a decrease of alkane production by up to 90 percent. Overexpression of slr1609 gene in the wild-type Synechocystis sp. PCC6803 had no effect on the production of either free fatty acids or alkanes. Overexpression or deletion of slr1609 gene in the Synechocystis sp. PCC6803 mutant strain with the capability of making fatty alcohols by genetically introducing fatty acyl-CoA reductase respectively enhanced or reduced fatty alcohol production by 60 percent. Conclusions Fatty acid activation functionalized by the slr1609 gene is metabolically crucial for biosynthesis of fatty acid derivatives in Synechocystis sp. PCC6803. It is necessary but not sufficient for efficient production of alkanes. Fatty alcohol production can be significantly improved by the overexpression of slr1609 gene.

  9. TALEN-mediated targeted mutagenesis of fatty acid desaturase 2 (FAD2) in peanut (Arachis hypogaea L.) promotes the accumulation of oleic acid.

    Science.gov (United States)

    Wen, Shijie; Liu, Hao; Li, Xingyu; Chen, Xiaoping; Hong, Yanbin; Li, Haifen; Lu, Qing; Liang, Xuanqiang

    2018-05-01

    A first creation of high oleic acid peanut varieties by using transcription activator-like effecter nucleases (TALENs) mediated targeted mutagenesis of Fatty Acid Desaturase 2 (FAD2). Transcription activator like effector nucleases (TALENs), which allow the precise editing of DNA, have already been developed and applied for genome engineering in diverse organisms. However, they are scarcely used in higher plant study and crop improvement, especially in allopolyploid plants. In the present study, we aimed to create targeted mutagenesis by TALENs in peanut. Targeted mutations in the conserved coding sequence of Arachis hypogaea fatty acid desaturase 2 (AhFAD2) were created by TALENs. Genetic stability of AhFAD2 mutations was identified by DNA sequencing in up to 9.52 and 4.11% of the regeneration plants at two different targeted sites, respectively. Mutation frequencies among AhFAD2 mutant lines were significantly correlated to oleic acid accumulation. Genetically, stable individuals of positive mutant lines displayed a 0.5-2 fold increase in the oleic acid content compared with non-transgenic controls. This finding suggested that TALEN-mediated targeted mutagenesis could increase the oleic acid content in edible peanut oil. Furthermore, this was the first report on peanut genome editing event, and the obtained high oleic mutants could serve for peanut breeding project.

  10. Homeostatic responses to amino acid insufficiency

    Directory of Open Access Journals (Sweden)

    Tracy G. Anthony

    2015-09-01

    Full Text Available This article provides a brief overview describing how two key signaling pathways, namely the integrated stress response and the mammalian target of rapamycin complex 1, work together to facilitate cellular adaptation to dietary amino acid insufficiency. A deeper understanding of these mechanisms is leading to identification of novel targets which aid in disease treatments, improve stress recovery and increase health span through slowed aging and enhanced metabolic fitness.

  11. Mutant p53 protein localized in the cytoplasm inhibits autophagy.

    Science.gov (United States)

    Morselli, Eugenia; Tasdemir, Ezgi; Maiuri, Maria Chiara; Galluzzi, Lorenzo; Kepp, Oliver; Criollo, Alfredo; Vicencio, José Miguel; Soussi, Thierry; Kroemer, Guido

    2008-10-01

    The knockout, knockdown or chemical inhibition of p53 stimulates autophagy. Moreover, autophagy-inducing stimuli such as nutrient depletion, rapamycin or lithium cause the depletion of cytoplasmic p53, which in turn is required for the induction of autophagy. Here, we show that retransfection of p53(-/-) HCT 116 colon carcinoma cells with wild type p53 decreases autophagy down to baseline levels. Surprisingly, one third among a panel of 22 cancer-associated p53 single amino acid mutants also inhibited autophagy when transfected into p53(-/-) cells. Those variants of p53 that preferentially localize to the cytoplasm effectively repressed autophagy, whereas p53 mutants that display a prominently nuclear distribution failed to inhibit autophagy. The investigation of a series of deletion mutants revealed that removal of the DNA-binding domain from p53 fails to interfere with its role in the regulation of autophagy. Altogether, these results identify the cytoplasmic localization of p53 as the most important feature for p53-mediated autophagy inhibition. Moreover, the structural requirements for the two biological activities of extranuclear p53, namely induction of apoptosis and inhibition of autophagy, are manifestly different.

  12. Studies on induced partially resistant mutants of barley against powdery mildew

    International Nuclear Information System (INIS)

    Roebbelen, G.; Abdel-Hafez, A.G.; Reinhold, M.; Kwon, H.J.; Neuhaus-Steinmetz, J.P.; Heun, M.

    1983-01-01

    After mutagenic seed treatment of three partially resistant cultivars of spring barley with EMS and NaN 3 , 45 mutants in a first and 16 in a second experiment were selected in the M 2 -M 4 generations. The screening was done alternatively under natural infection in the field or controlled infection with a single pathotype in the greenhouse. These mutants exhibited a higher resistance and a higher susceptibility, respectively, than the initial cultivars Asse, Bomi and Vada. Some mutants expressed their altered resistance behaviour particularly during certain stages of development. High-level resistance was conditioned by mutation in the ml-o locus in three cases. For several Bomi mutants pathotype specificity with and without reversed ranking was proven as well as pathotype non-specificity in comparison with the reaction of the original cultivar. In 14 cases studied the inheritance of the involved mutants was monogenic recessive. The laevigatum locus responsible for the intermediate mildew resistance of Bomi was not affected by the mutations. Detection of groups of allelic mutants showed that there are at least two regions in the barley genome in which mutations for mildew resistance can occur rather frequently. In total, the past ten years of this mutation research have given convincing evidence that the strategies of mutant screening applied have yielded promising new material both for breeding and for progress in basic understanding of host-pathogen interactions. (author)

  13. Promising rice mutants

    International Nuclear Information System (INIS)

    Hakim, L.; Azam, M.A.; Miah, A.J.; Mansur, M.A.; Akanda, H.R.

    1988-01-01

    Two induced mutants namely, Mut NS 1 (tall) and Mut NS 5 (semi-dwarf) derived from rice variety Nizersail were evaluated for various agronomic characters at four locations in Bangladesh. Both the mutants matured about three weeks earlier and yielded significantly higher than the parent variety Nizersail. (author). 3 tabs., 9 refs

  14. Low oleic acid-derived repression of jasmonic acid-inducible defense responses requires the WRKY50 and WRKY51 proteins

    Science.gov (United States)

    Signaling induced upon a reduction in oleic acid (18:1) levels simultaneously up-regulates salicylic acid (SA)-mediated responses and inhibits jasmonic acid (JA)-inducible defenses, resulting in enhanced resistance to biotrophs but increased susceptibility to necrotrophs. SA and the signaling compon...

  15. Deletion map of CYC1 mutants and its correspondence to mutationally altered iso-1-cytochromes c of yeast

    International Nuclear Information System (INIS)

    Sherman, F.; Jackson, M.; Liebman, S.W.; Schweingruber, A.M.; Stewart, J.W.

    1975-01-01

    Mutants arising spontaneously from sporulated cultures of certain strains of yeast, Saccharomyces cerevisiae, contained deletions of the CYC1 gene which controls the primary structure of iso-1-cytochrome c. At least 60 different kinds of deletions were uncovered among the 104 deletions examined and these ranged in length from those encompassing only two adjacent point mutants to those encompassing at least the entire CYC1 gene. X-ray-induced recombination rates of crosses involving these deletions and cyc1 point mutants resulted in the assignment of 211 point mutants to 47 mutational sites and made it possible to unambiguously order 40 of these 47 sites. Except for one mutant, cyc1-15, there was a strict colinear relationship between the deletion map and the positions of 13 sites that were previously determined by amino acid alterations in iso-1-cytochromes c from intragenic revertants

  16. Evaluation of Some Chemical Characteristics of barley Mutants induced by Gamma Irradiation

    International Nuclear Information System (INIS)

    Abdeldaiem, M.H.; Ali, H.G.M.

    2011-01-01

    This study aims to evaluate the antioxidant activity of acetonic extract from some barley mutations (P1, P2 and P3 varieties) induced by gamma irradiation as compared with local barley variety (Hordeum vulgare L.) as control. Barley samples were obtained from Plant Breeding Unit, Plant Research Department, Nuclear Research Centre, Atomic Energy Authority, Egypt. The measurements of the antioxidant activity using a radical scavenging capacity against 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ?-carotene bleaching assay were assessed in the barley acetonic extract. Furthermore, amino acids composition of barley mutant samples was determined. The results indicated that the acetonic extract of barley varieties under investigation possess marked antioxidant and anti radical capacities. The data showed that the acetonic extract of barley mutant P1 possessed the higher antioxidant activity as compared with the antioxidant activities of acetonic extract from control and other barley mutant samples. Meanwhile, the flour of barley mutations under investigation contained trace elements of iron, copper and manganese. GC and mass analyses were used to identify the active compound of extract of control and mutant barley samples. The results illustrated that the main components of the control sample of barely extract was pentane, 3 methyl (47.73%) while gamma irradiation caused noticeable change in the relative percentage of some components of acetonic extract from barley mutant samples. Moreover, the results presented that changes were disappeared, and some compounds of the acetonic extract from mutant barley samples were appeared. Furthermore, the results exhibited that barley flour supplemented with wheat flour at 30% level produced acceptable cookies. Accordingly, the phenolic constituents of barley acetonic extract induced by gamma irradiation, especially samples of P1 mutant, may have a future role as ingredients in the development of functional foods.

  17. Kinetics, improved activity and thermostability of endoglucanase and beta glucosidase from a mutant-derivative of aspergillus niger ms82

    International Nuclear Information System (INIS)

    Sohail, M.; Ahmad, A.; Khan, S.A.; Uddin, F.

    2013-01-01

    A mutant MS301 of Aspergillus niger MS82 showed 1.5 to 2.5-fold improved endoglucanase and beta-glucosidase activity when grown on crude lignocellulosic substrates under solid-state and submerged conditions. Indicators of thermal stability of enzymes (Tm and T1/2) showed that the wild type and mutant endoglucanase was more heat-resistant compared to beta-glucosidase. However, mutant and parent enzymes shared almost the same values for melting temperatures and half-lives. Endoglucanase and beta-glucosidase from both the strains showed optimum activity under acidic pH. Energy of activation (Ea) of mutant beta-glucosidase was substantially lower than the parent enzyme while Ea of mutant endoglucanase was slightly less than the parent. The lowered Ea values can be attributed to the improved beta-glucosidase activity of the mutant strain. Moreover, the MS301 enzymes were better in hydrolyzing purified and crude cellulosic materials than the parent MS82. (author)

  18. Engineering a pH responsive pore forming protein.

    Science.gov (United States)

    Kisovec, Matic; Rezelj, Saša; Knap, Primož; Cajnko, Miša Mojca; Caserman, Simon; Flašker, Ajda; Žnidaršič, Nada; Repič, Matej; Mavri, Janez; Ruan, Yi; Scheuring, Simon; Podobnik, Marjetka; Anderluh, Gregor

    2017-02-08

    Listeriolysin O (LLO) is a cytolysin capable of forming pores in cholesterol-rich lipid membranes of host cells. It is conveniently suited for engineering a pH-governed responsiveness, due to a pH sensor identified in its structure that was shown before to affect its stability. Here we introduced a new level of control of its hemolytic activity by making a variant with hemolytic activity that was pH-dependent. Based on detailed structural analysis coupled with molecular dynamics and mutational analysis, we found that the bulky side chain of Tyr406 allosterically affects the pH sensor. Molecular dynamics simulation further suggested which other amino acid residues may also allosterically influence the pH-sensor. LLO was engineered to the point where it can, in a pH-regulated manner, perforate artificial and cellular membranes. The single mutant Tyr406Ala bound to membranes and oligomerized similarly to the wild-type LLO, however, the final membrane insertion step was pH-affected by the introduced mutation. We show that the mutant toxin can be activated at the surface of artificial membranes or living cells by a single wash with slightly acidic pH buffer. Y406A mutant has a high potential in development of novel nanobiotechnological applications such as controlled release of substances or as a sensor of environmental pH.

  19. Engineering a pH responsive pore forming protein

    Science.gov (United States)

    Kisovec, Matic; Rezelj, Saša; Knap, Primož; Cajnko, Miša Mojca; Caserman, Simon; Flašker, Ajda; Žnidaršič, Nada; Repič, Matej; Mavri, Janez; Ruan, Yi; Scheuring, Simon; Podobnik, Marjetka; Anderluh, Gregor

    2017-02-01

    Listeriolysin O (LLO) is a cytolysin capable of forming pores in cholesterol-rich lipid membranes of host cells. It is conveniently suited for engineering a pH-governed responsiveness, due to a pH sensor identified in its structure that was shown before to affect its stability. Here we introduced a new level of control of its hemolytic activity by making a variant with hemolytic activity that was pH-dependent. Based on detailed structural analysis coupled with molecular dynamics and mutational analysis, we found that the bulky side chain of Tyr406 allosterically affects the pH sensor. Molecular dynamics simulation further suggested which other amino acid residues may also allosterically influence the pH-sensor. LLO was engineered to the point where it can, in a pH-regulated manner, perforate artificial and cellular membranes. The single mutant Tyr406Ala bound to membranes and oligomerized similarly to the wild-type LLO, however, the final membrane insertion step was pH-affected by the introduced mutation. We show that the mutant toxin can be activated at the surface of artificial membranes or living cells by a single wash with slightly acidic pH buffer. Y406A mutant has a high potential in development of novel nanobiotechnological applications such as controlled release of substances or as a sensor of environmental pH.

  20. X-ray induced degradation of DNA in radiosensitive mutants of Anacystis nidulans x-rays

    International Nuclear Information System (INIS)

    Zhukas, K.I.; Vorontsova, G.V.; Groshev, V.V.; Shestakov, S.V.

    1975-01-01

    In irradiated Cyanophyceae (Anacystis nidulans) cells there occurs a process of DNA degeneration to acid-soluble products which is linked with protein synthesis and stimulated by caffeine and acriflavine. The degree of DNA degeneration increases with X-ray dose, is not very dependent on the composition of the incubation medium and is weakly linked with photosynthesis. In the cells of a radiation-resistant mutant the degree of DNA degeneration is slighter, and in the cells of radiosensitive mutants larger, than in ordinary cells. The role of DNA degradation in the radiation detruction of cells is discussed. (author)

  1. Generation of comprehensive transposon insertion mutant library for the model archaeon, Haloferax volcanii, and its use for gene discovery.

    Science.gov (United States)

    Kiljunen, Saija; Pajunen, Maria I; Dilks, Kieran; Storf, Stefanie; Pohlschroder, Mechthild; Savilahti, Harri

    2014-12-09

    Archaea share fundamental properties with bacteria and eukaryotes. Yet, they also possess unique attributes, which largely remain poorly characterized. Haloferax volcanii is an aerobic, moderately halophilic archaeon that can be grown in defined media. It serves as an excellent archaeal model organism to study the molecular mechanisms of biological processes and cellular responses to changes in the environment. Studies on haloarchaea have been impeded by the lack of efficient genetic screens that would facilitate the identification of protein functions and respective metabolic pathways. Here, we devised an insertion mutagenesis strategy that combined Mu in vitro DNA transposition and homologous-recombination-based gene targeting in H. volcanii. We generated an insertion mutant library, in which the clones contained a single genomic insertion. From the library, we isolated pigmentation-defective and auxotrophic mutants, and the respective insertions pinpointed a number of genes previously known to be involved in carotenoid and amino acid biosynthesis pathways, thus validating the performance of the methodologies used. We also identified mutants that had a transposon insertion in a gene encoding a protein of unknown or putative function, demonstrating that novel roles for non-annotated genes could be assigned. We have generated, for the first time, a random genomic insertion mutant library for a halophilic archaeon and used it for efficient gene discovery. The library will facilitate the identification of non-essential genes behind any specific biochemical pathway. It represents a significant step towards achieving a more complete understanding of the unique characteristics of halophilic archaea.

  2. Photoproduct formation and repair capacity in a mutant of Bacillus cereus 569 producing UV-sensitive spores

    International Nuclear Information System (INIS)

    Weinberger, S.; Evenchik, Z.; Hertman, I.; Bar-Ilan Univ., Ramat-Gan

    1982-01-01

    A mutant of Bacillus cereus 569 UV sensitive in both vegetative and sporal stages was isolated by N-methyl-N'-nitro-N-nitrosoguanidine (NTG) mutagenesis followed by selection on mitomycin C. The UV-sensitive mutant designated as B. cereus 2422 exhibited normal content of dipicolinic acid (DPA) and resistance to X-rays and ethyl methanesulphonate. The photoproduct type and amount, induced by a given UV dose, was similar in either cells or spores of both the mutant 2422 and the wild-type ancestor. The mutant 2422 excised cyclobutane thymine dimers only to a limited extent (20%) as compared with 80% removal in the wild type. Removal of a spore-specific photoproduct (TDHT) during germination proceeded to a similar extent in B. cereus 2422 and the wild-type parent. However, under growing conditions, an additional removal of the TDHT was observed only in the wild-type strain. Liquid holding recovery occurred in irradiated wild-type cells, but not in mutant cells. Spontaneous revertants were isolated that regained UV resistance simultaneously in both the vegetative and sporal stage. (orig./AJ)

  3. The Putative E3 Ubiquitin Ligase ECERIFERUM9 Regulates Abscisic Acid Biosynthesis and Response during Seed Germination and Postgermination Growth in Arabidopsis1[W][OPEN

    Science.gov (United States)

    Zhao, Huayan; Zhang, Huoming; Cui, Peng; Ding, Feng; Wang, Guangchao; Li, Rongjun; Jenks, Matthew A.; Lü, Shiyou; Xiong, Liming

    2014-01-01

    The ECERIFERUM9 (CER9) gene encodes a putative E3 ubiquitin ligase that functions in cuticle biosynthesis and the maintenance of plant water status. Here, we found that CER9 is also involved in abscisic acid (ABA) signaling in seeds and young seedlings of Arabidopsis (Arabidopsis thaliana). The germinated embryos of the mutants exhibited enhanced sensitivity to ABA during the transition from reversible dormancy to determinate seedling growth. Expression of the CER9 gene is closely related to ABA levels and displays a similar pattern to that of ABSCISIC ACID-INSENSITIVE5 (ABI5), which encodes a positive regulator of ABA responses in seeds. cer9 mutant seeds exhibited delayed germination that is independent of seed coat permeability. Quantitative proteomic analyses showed that cer9 seeds had a protein profile similar to that of the wild type treated with ABA. Transcriptomics analyses revealed that genes involved in ABA biosynthesis or signaling pathways were differentially regulated in cer9 seeds. Consistent with this, high levels of ABA were detected in dry seeds of cer9. Blocking ABA biosynthesis by fluridone treatment or by combining an ABA-deficient mutation with cer9 attenuated the phenotypes of cer9. Whereas introduction of the abi1-1, abi3-1, or abi4-103 mutation could completely eliminate the ABA hypersensitivity of cer9, introduction of abi5 resulted only in partial suppression. These results indicate that CER9 is a novel negative regulator of ABA biosynthesis and the ABA signaling pathway during seed germination. PMID:24812105

  4. Studies on cytological, physiological and genetic characteristics in somatic mutant strains of Sugi (Cryptomeria japonica D. Don)

    International Nuclear Information System (INIS)

    Maeta, T.; Somegou, M.; Nakahira, K.; Miyazaki, Y.; Kondo, T.

    1982-01-01

    From microscopic observation of the pollen of induced mutant strains in Sugi (Cryptomeria japonica D. Don), it was found that there were large differences in pollen fertility among the mutant strains, and that it deviated year to year from the mother plants. The large differences in frequency of sterile pollen among mutant strains depended on the genetic characteristics of each mutant strain. Higher frequencies of sterile pollen were observed at the terminal part of branchlets in some mutant strains, and this was considered to be induced by the lateness of flower-bud formation at low temperature conditions in late summer. Delayed formation and gibberellic acid treatment applied for flower induction resulted in low fertility and abnormality of pollen in mutant strains. Chromosome aberration in mutant strains was caused either by gamma irradiation or by some mutational events that responded to environmental conditions. In the former case, aberration might have been maintained for a long period through vegetative propagation. Some of the irregularities were due to mitotic cell division, because cells with micronuclei at the pacytene stage in pollen mother cells and with fragments at MI were observed. Somatic mutability of Kuma-sugi mutants after re-irradiation was investigated. From waxless mutants morphological somatic mutations, which have fat or stout stems and thick and short needles, were frequently produced, whereas from morphological mutants the lowest somatic mutation frequency was induced. In some mutant strains higher rooting ability than the mother plants was found, and the possibility of character improvement was pointed out. (author)

  5. Identification of genes potentially involved in solute stress response in Sphingomonas wittichii RW1 by transposon mutant recovery

    Directory of Open Access Journals (Sweden)

    Edith eCoronado

    2014-11-01

    Full Text Available The term water stress refers to the effects of low water availability on microbial growth and physiology. Water availability has been proposed as a major constraint for the use of microorganisms in contaminated sites with the purpose of bioremediation. Sphingomonas wittichii RW1 is a bacterium capable of degrading the xenobiotic compounds dibenzofuran and dibenzo-p-dioxin, and has potential to be used for targeted bioremediation. The aim of the current work was to identify genes implicated in water stress in RW1 by means of transposon mutagenesis and mutant growth experiments. Conditions of low water potential were mimicked by adding NaCl to the growth media. Three different mutant selection or separation method were tested, which, however recovered different mutants. Recovered transposon mutants with poorer growth under salt-induced water stress carried insertions in genes involved in proline and glutamate biosynthesis, and further in a gene putatively involved in aromatic compound catabolism. Transposon mutants growing poorer on medium with lowered water potential also included ones that had insertions in genes involved in more general functions such as transcriptional regulation, elongation factor, cell division protein, RNA polymerase β or an aconitase.

  6. Identification and quantification of flavonoids in yellow grain mutant of rice (Oryza sativa L.).

    Science.gov (United States)

    Kim, Backki; Woo, Sunmin; Kim, Mi-Jung; Kwon, Soon-Wook; Lee, Joohyun; Sung, Sang Hyun; Koh, Hee-Jong

    2018-02-15

    Flavonoids are naturally occurring phenolic compounds with potential health-promoting activities. Although anthocyanins and phenolic acids in coloured rice have been investigated, few studies have focused on flavonoids. Herein, we analysed flavonoids in a yellow grain rice mutant using UHPLC-DAD-ESI-Q-TOF-MS, and identified 19 flavonoids by comparing retention times and accurate mass measurements. Among them, six flavonoids, isoorientin, isoorientin 2″-O-glucoside, vitexin 2″-O-glucoside, isovitexin, isoscoparin 2″-O-glucoside and isoscoparin, were isolated and fully identified from the yellow grain rice mutant, and the levels were significantly higher than wild-type, with isoorientin particularly abundant in mutant embryo. Significant differences in total phenolic compounds and antioxidant activity were observed in mutant rice by DPPH, FRAP and TEAC assays. The results suggest that the representative six flavonoids may play an important role in colouration and antioxidant activity of embryo and endosperm tissue. The findings provide insight into flavonoid biosynthesis and the possibility of improving functionality in rice. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. A comparative transcriptome analysis of a wild purple potato and its red mutant provides insight into the mechanism of anthocyanin transformation.

    Directory of Open Access Journals (Sweden)

    Fang Liu

    Full Text Available In this study, a red mutant was obtained through in vitro regeneration of a wild purple potato. High-performance liquid chromatography and Mass spectrometry analysis revealed that pelargonidin-3-O-glucoside and petunidin-3-O-glucoside were main anthocyanins in the mutant and wild type tubers, respectively. In order to thoroughly understand the mechanism of anthocyanin transformation in two materials, a comparative transcriptome analysis of the mutant and wild type was carried out through high-throughput RNA sequencing, and 295 differentially expressed genes (DEGs were obtained. Real-time qRT-PCR validation of DEGs was consistent with the transcriptome date. The DEGs mainly influenced biological and metabolic pathways, including phenylpropanoid biosynthesis and translation, and biosynthesis of flavone and flavonol. In anthocyanin biosynthetic pathway, the analysis of structural genes expressions showed that three genes, one encoding phenylalanine ammonia-lyase, one encoding 4-coumarate-CoA ligase and one encoding flavonoid 3',5'-hydroxylasem were significantly down-regulated in the mutant; one gene encoding phenylalanine ammonia-lyase was significantly up-regulated. Moreover, the transcription factors, such as bZIP family, MYB family, LOB family, MADS family, zf-HD family and C2H2 family, were significantly regulated in anthocyanin transformation. Response proteins of hormone, such as gibberellin, abscisic acid and brassinosteroid, were also significantly regulated in anthocyanin transformation. The information contributes to discovering the candidate genes in anthocyanin transformation, which can serve as a comprehensive resource for molecular mechanism research of anthocyanin transformation in potatoes.

  8. Mutant DnaAs of Escherichia coli that are refractory to negative control.

    Science.gov (United States)

    Chodavarapu, Sundari; Felczak, Magdalena M; Simmons, Lyle A; Murillo, Alec; Kaguni, Jon M

    2013-12-01

    DnaA is the initiator of DNA replication in bacteria. A mutant DnaA named DnaAcos is unusual because it is refractory to negative regulation. We developed a genetic method to isolate other mutant DnaAs that circumvent regulation to extend our understanding of mechanisms that control replication initiation. Like DnaAcos, one mutant bearing a tyrosine substitution for histidine 202 (H202Y) withstands the regulation exerted by datA, hda and dnaN (β clamp), and both DnaAcos and H202Y resist inhibition by the Hda-β clamp complex in vitro. Other mutant DnaAs carrying G79D, E244K, V303M or E445K substitutions are either only partially sensitive or refractory to inhibition by the Hda-β clamp complex in vitro but are responsive to hda expression in vivo. All mutant DnaAs remain able to interact directly with Hda. Of interest, both DnaAcos and DnaAE244K bind more avidly to Hda. These mutants, by sequestrating Hda, may limit its availability to regulate other DnaA molecules, which remain active to induce extra rounds of DNA replication. Other evidence suggests that a mutant bearing a V292M substitution hyperinitiates by escaping the effect of an unknown regulatory factor. Together, our results provide new insight into the mechanisms that regulate replication initiation in Escherichia coli.

  9. Grain Filling Characteristics and Their Relations with Endogenous Hormones in Large- and Small-Grain Mutants of Rice.

    Science.gov (United States)

    Zhang, Weiyang; Cao, Zhuanqin; Zhou, Qun; Chen, Jing; Xu, Gengwen; Gu, Junfei; Liu, Lijun; Wang, Zhiqin; Yang, Jianchang; Zhang, Hao

    2016-01-01

    This study determined if the variation in grain filling parameters between two different spikelet types of rice (Oryza sativa L.) is regulated by the hormonal levels in the grains. Two rice mutants, namely, a large-grain mutant (AZU-M) and a small-grain mutant (ZF802-M), and their respective wild types (AZU-WT and ZF802-WT) were grown in the field. The endosperm cell division rate, filling rate, and hormonal levels: zeatin + zeatin riboside (Z+ZR), indo-3-acetic acid (IAA), polyamines (PAs), and abscisic acid (ABA) were determined. The results showed that there was no significant difference between the filling and endosperm cell division rates. These rates were synchronous between the superior and inferior spikelets for both mutants. However, the abovementioned parameters were significantly different between the two spikelet types for the two wild types. The superior spikelets filled faster and their filling rate was higher compared to the inferior ones. Changes in the concentrations of plant hormones were consistent with the observed endosperm cell division rate and the filling rate for both types of spikelets of mutant and wild type plants. Regression analysis showed a significant positive correlation between cell division and filling rates with the concentrations of the investigated hormones. Exogenous chemical application verified the role of ABA, IAA, and PAs in grain filling. The results indicate that poor filling of inferior spikelets in rice occurs primarily due to the reduced hormone concentrations therein, leading to lower division rate of endosperm cells, fewer endosperm cells, slower filling rate, and smaller grain weight.

  10. Grain Filling Characteristics and Their Relations with Endogenous Hormones in Large- and Small-Grain Mutants of Rice.

    Directory of Open Access Journals (Sweden)

    Weiyang Zhang

    Full Text Available This study determined if the variation in grain filling parameters between two different spikelet types of rice (Oryza sativa L. is regulated by the hormonal levels in the grains. Two rice mutants, namely, a large-grain mutant (AZU-M and a small-grain mutant (ZF802-M, and their respective wild types (AZU-WT and ZF802-WT were grown in the field. The endosperm cell division rate, filling rate, and hormonal levels: zeatin + zeatin riboside (Z+ZR, indo-3-acetic acid (IAA, polyamines (PAs, and abscisic acid (ABA were determined. The results showed that there was no significant difference between the filling and endosperm cell division rates. These rates were synchronous between the superior and inferior spikelets for both mutants. However, the abovementioned parameters were significantly different between the two spikelet types for the two wild types. The superior spikelets filled faster and their filling rate was higher compared to the inferior ones. Changes in the concentrations of plant hormones were consistent with the observed endosperm cell division rate and the filling rate for both types of spikelets of mutant and wild type plants. Regression analysis showed a significant positive correlation between cell division and filling rates with the concentrations of the investigated hormones. Exogenous chemical application verified the role of ABA, IAA, and PAs in grain filling. The results indicate that poor filling of inferior spikelets in rice occurs primarily due to the reduced hormone concentrations therein, leading to lower division rate of endosperm cells, fewer endosperm cells, slower filling rate, and smaller grain weight.

  11. RodZ and PgsA play intertwined roles in membrane homeostasis of Bacillus subtilis and resistance to weak organic acid stress

    Directory of Open Access Journals (Sweden)

    Johan Willem Albertus Van Beilen

    2016-10-01

    Full Text Available Weak organic acids like sorbic and acetic acid are widely used to prevent growth of spoilage organisms such as Bacilli. To identify genes involved in weak acid stress tolerance we screened a transposon mutant library of Bacillus subtilis for sorbic acid sensitivity. Mutants of the rodZ (ymfM gene were found to be hypersensitive to the lipophilic weak organic acid. RodZ is involved in determining the cell’s rod-shape and believed to interact with the bacterial actin-like MreB cytoskeleton. Since rodZ lies upstream in the genome of the essential gene pgsA (phosphatidylglycerol phosphate synthase we hypothesized that expression of the latter might also be affected in rodZ mutants and hence contribute to the phenotype observed. We show that both genes are co-transcribed and that both the rodZ::mini-Tn10 mutant and a conditional pgsA mutant, under conditions of minimal pgsA expression, were sensitive to sorbic and acetic acid. Both strains displayed a severely altered membrane composition. Compared to the wild-type strain, phosphatidylglycerol and cardiolipin levels were lowered and the average acyl chain length was elongated. Induction of rodZ expression from a plasmid in our transposon mutant led to no recovery of weak acid susceptibility comparable to wild-type levels. However, pgsA overexpression in the same mutant partly restored sorbic acid susceptibility and fully restored acetic acid sensitivity. A construct containing both rodZ and pgsA as on the genome led to some restored growth as well. We propose that RodZ and PgsA play intertwined roles in membrane homeostasis and resistance to weak organic acid stress.

  12. Genetic analysis of amino acid transport in the facultatively heterotrophic cyanobacterium Synechocystis sp. Strain 6803

    International Nuclear Information System (INIS)

    Labarre, J.; Thuriaux, P.; Chauvat, F.

    1987-01-01

    The existence of active transport systems (permeases) operating on amino acids in the photoautotrophic cyanobacterium Synechocystis sp. strain 6803 was demonstrated by following the initial rates of uptake with 14 C-labeled amino acids, measuring the intracellular pools of amino acids, and isolating mutants resistant to toxic amino acids. One class of mutants (Pfa1) corresponds to a regulatory defect in the biosynthesis of the aromatic amino acids, but two other classes (Can1 and Aza1) are defective in amino acid transport. The Can1 mutants are defective in the active transport of three basic amino acids (arginine, histidine, and lysine) and in one of two transport systems operating on glutamine. The Aza1 mutants are not affected in the transport of the basic amino acids but have lost the capacity to transport all other amino acids except glutamate. The latter amino acid is probably transported by a third permease which could be identical to the Can1-independent transport operating on glutamine. Thus, genetic evidence suggests that strain 6803 has only a small number of amino acid transport systems with fairly broad specificity and that, with the exception of glutamine, each amino acid is accumulated by only one major transport system. Compared with heterotrophic bacteria such as Escherichia coli, these permeases are rather inefficient in terms of affinity (apparent K/sub m/ ranging from 6 to 60 μM) and of V/sub max/

  13. Gamma ray induced mutants in Coleus

    Energy Technology Data Exchange (ETDEWEB)

    Vasudevan, K; Jos, J S [Central Tuber Crops Research Institute, Trivandrum, Kerala (India)

    1988-07-01

    The germplasm collection of Chinese potato (Coleus parviflorus Benth) contains almost no variation for yield contributing traits. The crop does not produce seeds. Treatment of underground tubers with 1 kR, 2 kR, 3 kR and 4 kR gamma rays resulted in 50 morphologically different mutants which are maintained as mutant clones. In the M{sub 1}V{sub 1} generation, suspected mutant sprouts, were carefully removed and grown separately. The most interesting mutant types are the following: (i) erect mutant with spoon shaped light green leaves, 30 cm long inflorescences against 20 cm in the control, cylindrical tubers measuring ca. 7.0 cm long and 3 cm girth against 4 cm and 2.5 cm in the control (ii) early mutants 1 and 2, one having less leaf serration, the other having light green small leaves and dwarf type (iii) fleshy leaf mutant, dark green, thick and smooth leaves. Control plants spread almost in 1 m{sup 2} area and bear tubers from the nodes of branches. In the early mutants tuber formation is mainly restricted to the base of the plant, which makes harvest easier. The crop usually matures within 150 - 160 days, the early mutants are ready for harvest 100 days after planting. As the mutants are less spreading, the yield could be increased by closer spacing.

  14. Gamma ray induced mutants in Coleus

    International Nuclear Information System (INIS)

    Vasudevan, K.; Jos, J.S.

    1988-01-01

    The germplasm collection of Chinese potato (Coleus parviflorus Benth) contains almost no variation for yield contributing traits. The crop does not produce seeds. Treatment of underground tubers with 1 kR, 2 kR, 3 kR and 4 kR gamma rays resulted in 50 morphologically different mutants which are maintained as mutant clones. In the M 1 V 1 generation, suspected mutant sprouts, were carefully removed and grown separately. The most interesting mutant types are the following: (i) erect mutant with spoon shaped light green leaves, 30 cm long inflorescences against 20 cm in the control, cylindrical tubers measuring ca. 7.0 cm long and 3 cm girth against 4 cm and 2.5 cm in the control (ii) early mutants 1 and 2, one having less leaf serration, the other having light green small leaves and dwarf type (iii) fleshy leaf mutant, dark green, thick and smooth leaves. Control plants spread almost in 1 m 2 area and bear tubers from the nodes of branches. In the early mutants tuber formation is mainly restricted to the base of the plant, which makes harvest easier. The crop usually matures within 150 - 160 days, the early mutants are ready for harvest 100 days after planting. As the mutants are less spreading, the yield could be increased by closer spacing

  15. Efficacy of hepatitis B vaccine against antiviral drug-resistant hepatitis B virus mutants in the chimpanzee model.

    Science.gov (United States)

    Kamili, Saleem; Sozzi, Vitini; Thompson, Geoff; Campbell, Katie; Walker, Christopher M; Locarnini, Stephen; Krawczynski, Krzysztof

    2009-05-01

    Hepatitis B virus (HBV) mutants resistant to treatment with nucleoside or nucleotide analogs and those with the ability to escape from HBV-neutralizing antibody have the potential to infect HBV-vaccinated individuals. To address this potential serious public health challenge, we tested the efficacy of immunity induced by a commercial hepatitis B vaccine against a tissue culture-derived, clonal HBV polymerase mutant in HBV seronegative chimpanzees. The polymerase gene mutant contained a combination of three mutations (rtV173L, rtL180M, rtM204V), two of which resulted in changes to the overlapping viral envelope of the hepatitis B surface antigen (sE164D, sI195M). Prior to the HBV mutant challenge of vaccinated chimpanzees, we established virologic, serologic, and pathologic characteristics of infections resulting from intravenous inoculation of the HBV polymerase gene mutant and the sG145R vaccine-escape surface gene mutant. Cloning and sequencing experiments determined that the three mutations in the polymerase gene mutant remained stable and that the single mutation in the surface gene mutant reverted to the wild-type sequence. Immunological evidence of HBV replication was observed in the vaccinated chimpanzees after challenge with the polymerase gene mutant as well as after rechallenge with serum-derived wild-type HBV (5,000 chimpanzee infectious doses administered intravenously), despite robust humoral and cellular anti-HBV immune responses after hepatitis B vaccination. Our data showing successful experimental infection by HBV mutants despite the presence of high anti-HBs levels considered protective in the vaccinated host are consistent with clinical reports on breakthrough infection in anti-HBs-positive patients infected with HBV mutants. In the absence of a protective humoral immunity, adaptive cellular immune responses elicited by infection may limit HBV replication and persistence.

  16. A mutation of Aspergillus niger for hyper-production of citric acid from corn meal hydrolysate in a bioreactor*

    Science.gov (United States)

    Hu, Wei; Liu, Jing; Chen, Ji-hong; Wang, Shu-yang; Lu, Dong; Wu, Qing-hua; Li, Wen-jian

    2014-01-01

    The properties of the screened mutants for hyper-production of citric acid induced by carbon (12C6+) ion beams and X-ray irradiation were investigated in our current study. Among these mutants, mutant H4002 screened from 12C6+ ion irradiation had a higher yield of citric acid production than the parental strain in a 250-ml shaking flash. These expanded submerged experiments in a bioreactor were also carried out for mutant H4002. The results showed that (177.7–196.0) g/L citric acid was accumulated by H4002 through exploiting corn meal hydrolysate (containing initial 200.0–235.7 g/L sugar) with the productivity of (2.96–3.27) g/(L∙h). This was especially true when the initial sugar concentration was 210 g/L, and the best economical citric acid production reached (187.5±0.7) g/L with a productivity of 3.13 g/(L∙h). It was observed that mutant H4002 can utilize low-cost corn meal as a feedstock to efficiently produce citric acid. These results imply that the H4002 strain has the industrial production potentiality for citric acid and offers strong competition for the citric acid industry. PMID:25367793

  17. A mutation of Aspergillus niger for hyper-production of citric acid from corn meal hydrolysate in a bioreactor.

    Science.gov (United States)

    Hu, Wei; Liu, Jing; Chen, Ji-hong; Wang, Shu-yang; Lu, Dong; Wu, Qing-hua; Li, Wen-jian

    2014-11-01

    The properties of the screened mutants for hyper-production of citric acid induced by carbon ((12)C(6+)) ion beams and X-ray irradiation were investigated in our current study. Among these mutants, mutant H4002 screened from (12)C(6+) ion irradiation had a higher yield of citric acid production than the parental strain in a 250-ml shaking flash. These expanded submerged experiments in a bioreactor were also carried out for mutant H4002. The results showed that (177.7-196.0) g/L citric acid was accumulated by H4002 through exploiting corn meal hydrolysate (containing initial 200.0-235.7 g/L sugar) with the productivity of (2.96-3.27) g/(L∙h). This was especially true when the initial sugar concentration was 210 g/L, and the best economical citric acid production reached (187.5±0.7) g/L with a productivity of 3.13 g/(L∙h). It was observed that mutant H4002 can utilize low-cost corn meal as a feedstock to efficiently produce citric acid. These results imply that the H4002 strain has the industrial production potentiality for citric acid and offers strong competition for the citric acid industry.

  18. Renal response to acute acid loading--an organ physiological approach

    DEFF Research Database (Denmark)

    Osther, P J; Engel, K; Kildeberg, P

    2004-01-01

    , as the extracellular acid-base status would be expected to be the key physiological trigger for renal NAE. The object of this study was to investigate the renal response to acute non-carbonic acid loading using a quantitative organ physiological approach. MATERIAL AND METHODS: Five-h NH4Cl loading studies were...

  19. RecA-independent resistance to irradiation with u.v. light in acid-habituated Escherichia coli

    International Nuclear Information System (INIS)

    Goodson, M.; Rowbury, R.J.

    1991-01-01

    Growth of Escherichia coli 1829 ColV, I-K94 at pH 5.0 led to an increase in u.v. resistance compared with cells grown at pH 7.0. This was due to a phenotypic change, since organisms grown at pH 7.0 showed increased resistance after only 2.5-5.0 min incubation at the mildly acid pH. Other E. coli K12 derivatives became more u.v.-resistant at pH 5.0 including uvrA, recA and polAl mutants. Organisms grown at pH 5.0 also showed increased Weigle reactivation of u.v. irradiated λ phage and this applied to the repair-deficient mutants as well as the parent strains. Both the increased u.v. resistance of acid-habituated cells and their increased ability to bring about Weigle reactivation appear to involve RecA-independent processes and are presumably, therefore, independent of the SOS response. (author)

  20. Drastic anthocyanin increase in response to PAP1 overexpression in fls1 knockout mutant confers enhanced osmotic stress tolerance in Arabidopsis thaliana.

    Science.gov (United States)

    Lee, Won Je; Jeong, Chan Young; Kwon, Jaeyoung; Van Kien, Vu; Lee, Dongho; Hong, Suk-Whan; Lee, Hojoung

    2016-11-01

    KEY MESSAGE : pap1 - D/fls1ko double mutant plants that produce substantial amounts of anthocyanin show tolerance to abiotic stress. Anthocyanins are flavonoids that are abundant in various plants and have beneficial effects on both plants and humans. Many genes in flavonoid biosynthetic pathways have been identified, including those in the MYB-bHLH-WD40 (MBW) complex. The MYB gene Production of Anthocyanin Pigment 1 (PAP1) plays a particularly important role in anthocyanin accumulation. PAP1 expression in many plant systems strongly increases anthocyanin levels, resulting in a dark purple color in many plant organs. In this study, we generated double mutant plants that harbor fls1ko in the pap1-D background (i.e., pap1-D/fls1ko plants), to examine whether anthocyanins can be further enhanced by blocking flavonol biosynthesis under PAP1 overexpression. We also wanted to examine whether the increased anthocyanin levels contribute to defense against osmotic stresses. The pap1-D/fls1ko mutants accumulated higher anthocyanin levels than pap1-D plants in both control and sucrose-treated conditions. However, flavonoid biosynthesis genes were slightly down-regulated in the pap1-D/fls1ko seedlings as compared to their expression in pap1-D seedlings. We also report the performance of pap1-D/fls1ko seedlings in response to plant osmotic stresses.

  1. Detecting Sulfuric and Nitric Acid Rain Stresses on Quercus glauca through Hyperspectral Responses.

    Science.gov (United States)

    Wang, Shanqian; Zhang, Xiuying; Ma, Yuandan; Li, Xinhui; Cheng, Min; Zhang, Xiaomin; Liu, Lei

    2018-03-09

    Acid rain, which has become one of the most severe global environmental issues, is detrimental to plant growth. However, effective methods for monitoring plant responses to acid rain stress are currently lacking. The hyperspectral technique provides a cost-effective and nondestructive way to diagnose acid rain stresses. Taking a widely distributed species ( Quercus glauca ) in Southern China as an example, this study aims to monitor the hyperspectral responses of Q. glauca to simulated sulfuric acid rain (SAR) and nitric acid rain (NAR). A total of 15 periods of leaf hyperspectral data under four pH levels of SAR and NAR were obtained during the experiment. The results showed that hyperspectral information could be used to distinguish plant responses under acid rain stress. An index (green peak area index, GPAI) was proposed to indicate acid rain stresses, based on the significantly variations in the region of 500-660 nm. Light acid rain (pH 4.5 SAR and NAR) promoted Q. glauca growth relative to the control groups (pH 5.6 SAR and NAR); moderate acid rain (pH 3.0 SAR) firstly promoted and then inhibited plant growth, while pH 3.0 NAR showed mild inhibitory effects during the experiment; and heavy acid rain (pH 2.0) significantly inhibited plant growth. Compared with NAR, SAR induced more serious damages to Q. glauca . These results could help monitor acid rain stress on plants on a regional scale using remote sensing techniques.

  2. Increased Protein Stability and Decreased Protein Turnover in the Caenorhabditis elegans Ins/IGF-1 daf-2 Mutant.

    Science.gov (United States)

    Depuydt, Geert; Shanmugam, Nilesh; Rasulova, Madina; Dhondt, Ineke; Braeckman, Bart P

    2016-12-01

    In Caenorhabditis elegans, cellular proteostasis is likely essential for longevity. Autophagy has been shown to be essential for lifespan extension of daf-2 insulin/IGF mutants. Therefore, it can be hypothesized that daf-2 mutants achieve this phenotype by increasing protein turnover. However, such a mechanism would exert a substantial energy cost. By using classical 35 S pulse-chase labeling, we observed that protein synthesis and degradation rates are decreased in young adults of the daf-2 insulin/IGF mutants. Although reduction of protein turnover may be energetically favorable, it may lead to accumulation and aggregation of damaged proteins. As this has been shown not to be the case in daf-2 mutants, another mechanism must exist to maintain proteostasis in this strain. We observed that proteins isolated from daf-2 mutants are more soluble in acidic conditions due to increased levels of trehalose. This suggests that trehalose may decrease the potential for protein aggregation and increases proteostasis in the daf-2 mutants. We postulate that daf-2 mutants save energy by decreasing protein turnover rates and instead stabilize their proteome by trehalose. © The Author 2016. Published by Oxford University Press on behalf of The Gerontological Society of America.

  3. Transcriptional Response to Lactic Acid Stress in the Hybrid Yeast Zygosaccharomyces parabailii.

    Science.gov (United States)

    Ortiz-Merino, Raúl A; Kuanyshev, Nurzhan; Byrne, Kevin P; Varela, Javier A; Morrissey, John P; Porro, Danilo; Wolfe, Kenneth H; Branduardi, Paola

    2018-03-01

    Lactic acid has a wide range of applications starting from its undissociated form, and its production using cell factories requires stress-tolerant microbial hosts. The interspecies hybrid yeast Zygosaccharomyces parabailii has great potential to be exploited as a novel host for lactic acid production, due to high organic acid tolerance at low pH and a fermentative metabolism with a high growth rate. Here we used mRNA sequencing (RNA-seq) to analyze Z. parabailii 's transcriptional response to lactic acid added exogenously, and we explore the biological mechanisms involved in tolerance. Z. parabailii contains two homeologous copies of most genes. Under lactic acid stress, the two genes in each homeolog pair tend to diverge in expression to a significantly greater extent than under control conditions, indicating that stress tolerance is facilitated by interactions between the two gene sets in the hybrid. Lactic acid induces downregulation of genes related to cell wall and plasma membrane functions, possibly altering the rate of diffusion of lactic acid into cells. Genes related to iron transport and redox processes were upregulated, suggesting an important role for respiratory functions and oxidative stress defense. We found differences in the expression profiles of genes putatively regulated by Haa1 and Aft1/Aft2, previously described as lactic acid responsive in Saccharomyces cerevisiae Furthermore, formate dehydrogenase ( FDH ) genes form a lactic acid-responsive gene family that has been specifically amplified in Z. parabailii in comparison to other closely related species. Our study provides a useful starting point for the engineering of Z. parabailii as a host for lactic acid production. IMPORTANCE Hybrid yeasts are important in biotechnology because of their tolerance to harsh industrial conditions. The molecular mechanisms of tolerance can be studied by analyzing differential gene expression under conditions of interest and relating gene expression patterns

  4. Sustracted library obtained from mutant sugarcane variety B 4362 resistant to rust

    Directory of Open Access Journals (Sweden)

    María I. Oloriz

    2002-07-01

    Full Text Available The hypersensitive response is one of the most powerful mechanisms for which the plants resist pathogen attack. Mutations carried out previously on the variety B4362, of sugarcane, originated five mutants that express this mechanism towards the attack of rust (Puccinia melanocephala Syd.. By means of a subtractive hybridization among the cDNA obtained starting from the resistant clone inoculated with rust and a pool of cDNA of the susceptible variety (B4362 inoculated and of the resistant clone not inoculated, it was possible to reduce the number of genes expressed during the infection with the fungus. A subtractive library was carried out where we hope that most of the genes are involved in the hypersensitive response that present these mutants towards the infection of the pathogen. Key words: Subtractive hybridization, hypersensitive response, Puccinia melanocephala Syd.

  5. Rapid selection of escape mutants by the first CD8 T cell responses in acute HIV-1 infection

    Energy Technology Data Exchange (ETDEWEB)

    Korber, Bette Tina Marie [Los Alamos National Laboratory

    2008-01-01

    The recent failure of a vaccine that primes T cell responses to control primary HIV-1 infection has raised doubts about the role of CD8+ T cells in early HIV-1 infection. We studied four patients who were identified shortly after HIV-1 infection and before seroconversion. In each patient there was very rapid selection of multiple HIV-1 escape mutants in the transmitted virus by CD8 T cells, including examples of complete fixation of non-synonymous substitutions within 2 weeks. Sequencing by single genome amplification suggested that the high rate of virus replication in acute infection gave a selective advantage to virus molecules that contained simultaneous and gained sequential T cell escape mutations. These observations show that whilst early HIV-1 specific CD8 T cells can act against virus, rapid escape means that these T cell responses are unlikely to benefit the patient and may in part explain why current HIV-1 T cell vaccines may not be protective.

  6. Genetic cytological and biochemical study of a tomato chlorophyll mutant of the xanthic type, obtained by irradiation of the seeds; Etude genetique, cytologique et biochimique d'un mutant chlorophyllien de tomate du type xantha, obtenu par irradiation de graines

    Energy Technology Data Exchange (ETDEWEB)

    Lefort, M; Duranton, J; Galmiche, J M; Roux, E [Commissariat a l' Energie Atomique, Saclay (France). Centre d' Etudes Nucleaires

    1958-07-01

    Irradiation of Lycopersicum aesculantum seeds with increasing doses of X-rays and thermal neutrons leads to the appearance of chlorophyll mutations in the descendants of the irradiated seeds. A genetic study of one of these mutants of the xanthic type showed that it was a recessive mutant with typical mono-genetic separation, while the cytological study demonstrated that the differentiation of the plast stopped at the stage of elementary lamella. Finally it is shown that in the light, the mutation brings about a very large deviation of the carbon metabolism towards the synthesis of amino acids and proteins, at the expense of that of glucosides. (author) [French] L'irradiation de graines de Lycopersicum Aesculantum avec des doses croissantes de rayons X et de neutrons thermiques entraine l'apparition de mutations chlorophylliennes dans la descendance des graines irradiees. L'etude genetique d'un de ces mutants du type xantha a montre qu'il s'agissait d'un mutant recessif a disjonction monogenique typique, tandis que l'etude cytologique a revele que la differentiation du plaste s'arretait au stade de lamelles elementaires. Il est apparu enfin qu'a la lumiere la mutation entrainait une deviation tres importante du metabolisme du carbone vers la synthese des acides amines et des proteines, au detriment de celle des glucides. (auteur)

  7. Systems Analysis of Adaptive Responses to MAP Kinase Pathway Blockade in BRAF Mutant Melanoma.

    Directory of Open Access Journals (Sweden)

    Brian J Capaldo

    Full Text Available Fifty percent of cutaneous melanomas are driven by activated BRAFV600E, but tumors treated with RAF inhibitors, even when they respond dramatically, rapidly adapt and develop resistance. Thus, there is a pressing need to identify the major mechanisms of intrinsic and adaptive resistance and develop drug combinations that target these resistance mechanisms. In a combinatorial drug screen on a panel of 12 treatment-naïve BRAFV600E mutant melanoma cell lines of varying levels of resistance to mitogen-activated protein kinase (MAPK pathway inhibition, we identified the combination of PLX4720, a targeted inhibitor of mutated BRaf, and lapatinib, an inhibitor of the ErbB family of receptor tyrosine kinases, as synergistically cytotoxic in the subset of cell lines that displayed the most resistance to PLX4720. To identify potential mechanisms of resistance to PLX4720 treatment and synergy with lapatinib treatment, we performed a multi-platform functional genomics analysis to profile the genome as well as the transcriptional and proteomic responses of these cell lines to treatment with PLX4720. We found modest levels of resistance correlated with the zygosity of the BRAF V600E allele and receptor tyrosine kinase (RTK mutational status. Layered over base-line resistance was substantial upregulation of many ErbB pathway genes in response to BRaf inhibition, thus generating the vulnerability to combination with lapatinib. The transcriptional responses of ErbB pathway genes are associated with a number of transcription factors, including ETS2 and its associated cofactors that represent a convergent regulatory mechanism conferring synergistic drug susceptibility in the context of diverse mutational landscapes.

  8. The Swedish mutant barley collection

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1989-07-01

    Full text: The Swedish mutation research programme in barley began about 50 years ago and has mainly been carried out at Svaloev in co-operation with the institute of Genetics at the University of Lund. The collection has been produced from different Swedish high-yielding spring barley varieties, using the following mutagens: X-rays, neutrons, several organic chemical compounds such as ethyleneimine, several sulfonate derivatives and the inorganic chemical mutagen sodium azide. Nearly 10,000 barley mutants are stored in the Nordic Gene Bank and documented in databases developed by Udda Lundquist, Svaloev AB. The collection consists of the following nine categories with 94 different types of mutants: 1. Mutants with changes in the spike and spikelets; 2. Changes in culm length and culm composition; 3. Changes in growth types; 4. Physiological mutants; 5. Changes in awns; 6. Changes in seed size and shape; 7. Changes in leaf blades; 8. Changes in anthocyanin and colour; 9. Resistance to barley powdery mildew. Barley is one of the most thoroughly investigated crops in terms of induction of mutations and mutation genetics. So far, about half of the mutants stored at the Nordic Gene Bank, have been analysed genetically; They constitute, however, only a minority of the 94 different mutant types. The genetic analyses have given valuable insights into the mutation process but also into the genetic architecture of various characters. A number of mutants of two-row barley have been registered and commercially released. One of the earliest released, Mari, an early maturing, daylength neutral, straw stiff mutant, is still grown in Iceland. The Swedish mutation material has been used in Sweden, but also in other countries, such as Denmark, Germany, and USA, for various studies providing a better understanding of the barley genome. The collection will be immensely valuable for future molecular genetical analyses of clone mutant genes. (author)

  9. The Swedish mutant barley collection

    International Nuclear Information System (INIS)

    1989-01-01

    Full text: The Swedish mutation research programme in barley began about 50 years ago and has mainly been carried out at Svaloev in co-operation with the institute of Genetics at the University of Lund. The collection has been produced from different Swedish high-yielding spring barley varieties, using the following mutagens: X-rays, neutrons, several organic chemical compounds such as ethyleneimine, several sulfonate derivatives and the inorganic chemical mutagen sodium azide. Nearly 10,000 barley mutants are stored in the Nordic Gene Bank and documented in databases developed by Udda Lundquist, Svaloev AB. The collection consists of the following nine categories with 94 different types of mutants: 1. Mutants with changes in the spike and spikelets; 2. Changes in culm length and culm composition; 3. Changes in growth types; 4. Physiological mutants; 5. Changes in awns; 6. Changes in seed size and shape; 7. Changes in leaf blades; 8. Changes in anthocyanin and colour; 9. Resistance to barley powdery mildew. Barley is one of the most thoroughly investigated crops in terms of induction of mutations and mutation genetics. So far, about half of the mutants stored at the Nordic Gene Bank, have been analysed genetically; They constitute, however, only a minority of the 94 different mutant types. The genetic analyses have given valuable insights into the mutation process but also into the genetic architecture of various characters. A number of mutants of two-row barley have been registered and commercially released. One of the earliest released, Mari, an early maturing, daylength neutral, straw stiff mutant, is still grown in Iceland. The Swedish mutation material has been used in Sweden, but also in other countries, such as Denmark, Germany, and USA, for various studies providing a better understanding of the barley genome. The collection will be immensely valuable for future molecular genetical analyses of clone mutant genes. (author)

  10. The ABA receptor PYL8 promotes lateral root growth by enhancing MYB77-dependent transcription of auxin-responsive genes.

    Science.gov (United States)

    Zhao, Yang; Xing, Lu; Wang, Xingang; Hou, Yueh-Ju; Gao, Jinghui; Wang, Pengcheng; Duan, Cheng-Guo; Zhu, Xiaohong; Zhu, Jian-Kang

    2014-06-03

    The phytohormone abscisic acid (ABA) regulates plant growth, development, and abiotic stress responses. ABA signaling is mediated by a group of receptors known as the PYR1/PYL/RCAR family, which includes the pyrabactin resistance 1-like protein PYL8. Under stress conditions, ABA signaling activates SnRK2 protein kinases to inhibit lateral root growth after emergence from the primary root. However, even in the case of persistent stress, lateral root growth eventually recovers from inhibition. We showed that PYL8 is required for the recovery of lateral root growth, following inhibition by ABA. PYL8 directly interacted with the transcription factors MYB77, MYB44, and MYB73. The interaction of PYL8 and MYB77 increased the binding of MYB77 to its target MBSI motif in the promoters of multiple auxin-responsive genes. Compared to wild-type seedlings, the lateral root growth of pyl8 mutant seedlings and myb77 mutant seedlings was more sensitive to inhibition by ABA. The recovery of lateral root growth was delayed in pyl8 mutant seedlings in the presence of ABA, and the defect was rescued by exposing pyl8 mutant seedlings to the auxin IAA (3-indoleacetic acid). Thus, PYL8 promotes lateral root growth independently of the core ABA-SnRK2 signaling pathway by enhancing the activities of MYB77 and its paralogs, MYB44 and MYB73, to augment auxin signaling. Copyright © 2014, American Association for the Advancement of Science.

  11. Lack of negative charge in the E46Q mutant of photoactive yellow protein prevents partial unfolding of the blue shifted intermediate

    NARCIS (Netherlands)

    Derix, N.M.; Wechselberger, R.W.|info:eu-repo/dai/nl/304829005; van der Horst, M.A.; Hellingwerf, K.J.; Boelens, R.|info:eu-repo/dai/nl/070151407; Kaptein, R.|info:eu-repo/dai/nl/074334603; van Nuland, N.A.J.

    2003-01-01

    The long-lived light-induced intermediate (pB) of the E46Q mutant (glutamic acid is replaced by glutamine at position 46) of photoactive yellow protein (PYP) has been investigated by NMR spectroscopy. The ground state of this mutant is very similar to that of wild-type PYP (WT), whereas the pB

  12. Genetic requirements for high constitutive SOS expression in recA730 mutants of Escherichia coli.

    Science.gov (United States)

    Vlašić, Ignacija; Šimatović, Ana; Brčić-Kostić, Krunoslav

    2011-09-01

    The RecA protein in its functional state is in complex with single-stranded DNA, i.e., in the form of a RecA filament. In SOS induction, the RecA filament functions as a coprotease, enabling the autodigestion of the LexA repressor. The RecA filament can be formed by different mechanisms, but all of them require three enzymatic activities essential for the processing of DNA double-stranded ends. These are helicase, 5'-3' exonuclease, and RecA loading onto single-stranded DNA (ssDNA). In some mutants, the SOS response can be expressed constitutively during the process of normal DNA metabolism. The RecA730 mutant protein is able to form the RecA filament without the help of RecBCD and RecFOR mediators since it better competes with the single-strand binding (SSB) protein for ssDNA. As a consequence, the recA730 mutants show high constitutive SOS expression. In the study described in this paper, we studied the genetic requirements for constitutive SOS expression in recA730 mutants. Using a β-galactosidase assay, we showed that the constitutive SOS response in recA730 mutants exhibits different requirements in different backgrounds. In a wild-type background, the constitutive SOS response is partially dependent on RecBCD function. In a recB1080 background (the recB1080 mutation retains only helicase), constitutive SOS expression is partially dependent on RecBCD helicase function and is strongly dependent on RecJ nuclease. Finally, in a recB-null background, the constitutive SOS expression of the recA730 mutant is dependent on the RecJ nuclease. Our results emphasize the importance of the 5'-3' exonuclease for high constitutive SOS expression in recA730 mutants and show that RecBCD function can further enhance the excellent intrinsic abilities of the RecA730 protein in vivo. Copyright © 2011, American Society for Microbiology. All Rights Reserved.

  13. Gibberellin requirement for Arabidopsis seed germination is determined both by testa characteristics and embryonic abscisic acid

    NARCIS (Netherlands)

    Debeaujon, I.; Koornneef, M.

    2000-01-01

    The mechanisms imposing a gibberellin (GA) requirement to promote the germination of dormant and non-dormant Arabidopsis seeds were analyzed using the GA-deficient mutant ga1, several seed coat pigmentation and structure mutants, and the abscisic acid (ABA)-deficient mutant aba1. Testa mutants,

  14. Probing the mechanism of insulin fibril formation with insulin mutants.

    Science.gov (United States)

    Nielsen, L; Frokjaer, S; Brange, J; Uversky, V N; Fink, A L

    2001-07-27

    The molecular basis of insulin fibril formation was investigated by studying the structural properties and kinetics of fibril formation of 20 different human insulin mutants at both low pH (conditions favoring monomer/dimer) and at pH 7.4 (conditions favoring tetramer/hexamer). Small-angle X-ray scattering showed insulin to be monomeric in 20% acetic acid, 0.1 M NaCl, pH 2. The secondary structure of the mutants was assessed using far-UV circular dichroism, and the tertiary structure was determined using near-UV circular dichroism, quenching of intrinsic fluorescence by acrylamide and interactions with the hydrophobic probe 1-anilino-8-naphthalene-sulfonic acid (ANS). The kinetics of fibril formation were monitored with the fluorescent dye, Thioflavin T. The results indicate that the monomer is the state from which fibrils arise, thus under some conditions dissociation of hexamers may be rate limiting or partially rate limiting. The insulin mutants were found to retain substantial nativelike secondary and tertiary structure under all conditions studied. The results suggest that fibril formation of the insulin mutants is controlled by specific molecular interactions that are sensitive to variations in the primary structure. The observed effects of several mutations on the rate of fibril formation are inconsistent with a previously suggested model for fibrillation [Brange, J., Whittingham, J., Edwards, D., Youshang, Z., Wollmer, A., Brandenburg, D., Dodson, G., and Finch, J. (1997) Curr. Sci. 72, 470-476]. Two surfaces on the insulin monomer are identified as potential interacting sites in insulin fibrils, one consisting of the residues B10, B16, and B17 and the other consisting of at least the residues A8 and B25. The marked increase in the lag time for fibril formation with mutations to more polar residues, as well as mutations to charged residues, demonstrates the importance of both hydrophobic and electrostatic interactions in the initial stages of fibrillation

  15. Acid detergent lignin, lodging resistance index, and expression of the caffeic acid O-methyltransferase gene in brown midrib-12 sudangrass.

    Science.gov (United States)

    Li, Yuan; Liu, Guibo; Li, Jun; You, Yongliang; Zhao, Haiming; Liang, Huan; Mao, Peisheng

    2015-09-01

    Understanding the relationship between acid detergent lignin (ADL) and lodging resistance index (LRI) is essential for breeding new varieties of brown midrib (bmr) sudangrass (Sorghum sudanense (Piper) Stapf.). In this study, bmr-12 near isogenic lines and their wild-types obtained by back cross breeding were used to compare relevant forage yield and quality traits, and to analyze expression of the caffeic acid O-methyltransferase (COMT) gene using quantitative real time-PCR. The research showed that the mean ADL content of bmr-12 mutants (20.94 g kg(-1)) was significantly (P bmr-12 mutants (0.29) was significantly (P bmr-12 materials (r = -0.44, P > 0.05). Sequence comparison of the COMT gene revealed two point mutations present in bmr-12 but not in the wild-type, the second mutation changed amino acid 129 of the protein from Gln (CAG) to a stop codon (UAG). The relative expression level of COMT gene was significantly reduced, which likely led to the decreased ADL content observed in the bmr-12 mutant.

  16. Alternative Splicing Control of Abiotic Stress Responses.

    Science.gov (United States)

    Laloum, Tom; Martín, Guiomar; Duque, Paula

    2018-02-01

    Alternative splicing, which generates multiple transcripts from the same gene, is an important modulator of gene expression that can increase proteome diversity and regulate mRNA levels. In plants, this post-transcriptional mechanism is markedly induced in response to environmental stress, and recent studies have identified alternative splicing events that allow rapid adjustment of the abundance and function of key stress-response components. In agreement, plant mutants defective in splicing factors are severely impaired in their response to abiotic stress. Notably, mounting evidence indicates that alternative splicing regulates stress responses largely by targeting the abscisic acid (ABA) pathway. We review here current understanding of post-transcriptional control of plant stress tolerance via alternative splicing and discuss research challenges for the near future. Copyright © 2017 Elsevier Ltd. All rights reserved.

  17. Response of the pearly eye melon fly Bactrocera cucurbitae (Coquillett)(Diptera:Tephritidae) mutant to host-associated visual cues

    Science.gov (United States)

    We report on a pearly eye mutant (PEM) line generated from a single male Bactrocera cucurbitae collected in Kapoho, Hawaii. Crossing experiments with colony wild-type flies indicate that the locus controlling this trait is autosomal and the mutant allele is recessive. Experiments with females to ass...

  18. A buoyancy-based screen of Drosophila larvae for fat-storage mutants reveals a role for Sir2 in coupling fat storage to nutrient availability.

    Directory of Open Access Journals (Sweden)

    Tânia Reis

    2010-11-01

    Full Text Available Obesity has a strong genetic component, but few of the genes that predispose to obesity are known. Genetic screens in invertebrates have the potential to identify genes and pathways that regulate the levels of stored fat, many of which are likely to be conserved in humans. To facilitate such screens, we have developed a simple buoyancy-based screening method for identifying mutant Drosophila larvae with increased levels of stored fat. Using this approach, we have identified 66 genes that when mutated increase organismal fat levels. Among these was a sirtuin family member, Sir2. Sirtuins regulate the storage and metabolism of carbohydrates and lipids by deacetylating key regulatory proteins. However, since mammalian sirtuins function in many tissues in different ways, it has been difficult to define their role in energy homeostasis accurately under normal feeding conditions. We show that knockdown of Sir2 in the larval fat body results in increased fat levels. Moreover, using genetic mosaics, we demonstrate that Sir2 restricts fat accumulation in individual cells of the fat body in a cell-autonomous manner. Consistent with this function, changes in the expression of metabolic enzymes in Sir2 mutants point to a shift away from catabolism. Surprisingly, although Sir2 is typically upregulated under conditions of starvation, Sir2 mutant larvae survive better than wild type under conditions of amino-acid starvation as long as sugars are provided. Our findings point to a Sir2-mediated pathway that activates a catabolic response to amino-acid starvation irrespective of the sugar content of the diet.

  19. Good and bad protons: genetic aspects of acidity stress responses in plants.

    Science.gov (United States)

    Shavrukov, Yuri; Hirai, Yoshihiko

    2016-01-01

    Physiological aspects of acidity stress in plants (synonymous with H(+) rhizotoxicity or low-pH stress) have long been a focus of research, in particular with respect to acidic soils where aluminium and H(+) rhizotoxicities often co-occur. However, toxic H(+) and Al(3+) elicit different response mechanisms in plants, and it is important to consider their effects separately. The primary aim of this review was to provide the current state of knowledge regarding the genetics of the specific reactions to low-pH stress in growing plants. A comparison of the results gleaned from quantitative trait loci analysis and global transcriptome profiling of plants in response to high proton concentrations revealed a two-stage genetic response: (i) in the short-term, proton pump H(+)-ATPases present the first barrier in root cells, allocating an excess of H(+) into either the apoplast or vacuole; the ensuing defence signaling system involves auxin, salicylic acid, and methyl jasmonate, which subsequently initiate expression of STOP and DREB transcription factors as well as chaperone ROF; (2) the long-term response includes other genes, such as alternative oxidase and type II NAD(P)H dehydrogenase, which act to detoxify dangerous reactive oxygen species in mitochondria, and help plants better manage the stress. A range of transporter genes including those for nitrate (NTR1), malate (ALMT1), and heavy metals are often up-regulated by H(+) rhizotoxicity. Expansins, cell-wall-related genes, the γ-aminobutyric acid shunt and biochemical pH-stat genes also reflect changes in cell metabolism and biochemistry in acidic conditions. However, the genetics underlying the acidity stress response of plants is complicated and only fragmentally understood. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  20. Directed mutagenesis in Candida albicans: one-step gene disruption to isolate ura3 mutants

    International Nuclear Information System (INIS)

    Kelly, R.; Miller, S.M.; Kurtz, M.B.; Kirsch, D.R.

    1987-01-01

    A method for introducing specific mutations into the diploid Candida albicans by one-step gene disruption and subsequent UV-induced recombination was developed. The cloned C. albicans URA3 gene was disrupted with the C. albicans ADE2 gene, and the linearized DNA was used for transformation of two ade2 mutants, SGY-129 and A81-Pu. Both an insertional inactivation of the URA3 gene and a disruption which results in a 4.0-kilobase deletion were made. Southern hybridization analyses demonstrated that the URA3 gene was disrupted on one of the chromosomal homologs in 15 of the 18 transformants analyzed. These analyses also revealed restriction site dimorphism of EcoRI at the URA3 locus which provides a unique marker to distinguish between chromosomal homologs. This enabled us to show that either homolog could be disrupted and that disrupted transformants of SGY-129 contained more than two copies of the URA3 locus. The A81-Pu transformants heterozygous for the ura3 mutations were rendered homozygous and Ura- by UV-induced recombination. The homozygosity of a deletion mutant and an insertion mutant was confirmed by Southern hybridization. Both mutants were transformed to Ura+ with plasmids containing the URA3 gene and in addition, were resistant to 5-fluoro-orotic acid, a characteristic of Saccharomyces cerevisiae ura3 mutants as well as of orotidine-5'-phosphate decarboxylase mutants of other organisms

  1. Transcriptome profiling identifies genes and pathways deregulated upon floxuridine treatment in colorectal cancer cells harboring GOF mutant p53

    Directory of Open Access Journals (Sweden)

    Arindam Datta

    2016-06-01

    Full Text Available Mutation in TP53 is a common genetic alteration in human cancers. Certain tumor associated p53 missense mutants acquire gain-of-function (GOF properties and confer oncogenic phenotypes including enhanced chemoresistance. The colorectal cancers (CRC harboring mutant p53 are generally aggressive in nature and difficult to treat. To identify a potential gene expression signature of GOF mutant p53-driven acquired chemoresistance in CRC, we performed transcriptome profiling of floxuridine (FUdR treated SW480 cells expressing mutant p53R273H (GEO#: GSE77533. We obtained several genes differentially regulated between FUdR treated and untreated cells. Further, functional characterization and pathway analysis revealed significant enrichment of crucial biological processes and pathways upon FUdR treatment in SW480 cells. Our data suggest that in response to chemotherapeutics treatment, cancer cells with GOF mutant p53 can modulate key cellular pathways to withstand the cytotoxic effect of the drugs. The genes and pathways identified in the present study can be further validated and targeted for better chemotherapy response in colorectal cancer patients harboring mutant p53.

  2. Thc6 protein, isolated from Trichoderma harzianum, can induce maize defense response against Curvularia lunata.

    Science.gov (United States)

    Fan, Lili; Fu, Kehe; Yu, Chuanjin; Li, Yingying; Li, Yaqian; Chen, Jie

    2015-05-01

    Mutant T66 was isolated from 450 mutants (constructed with Agrobacterium tumefaciens-mediated transformation method) of Trichoderma harzianum. Maize seeds coated with T66 were more susceptible to Curvularia lunata when compared with those coated with wild-type (WT) strain. The disease index of maize treated with T66 and WT were 62.5 and 42.1%, respectively. Further research showed T-DNA has inserted into the ORF of one gene, which resulted in the functional difference between WT and T66. The gene was cloned and named Thc6, which encodes a novel 327 amino acid protein. To investigate its function, we obtained knockout, complementation, and overexpression mutants of Thc6. Challenge inoculation studies suggested that the Thc6 overexpression mutant can reduce the disease index of maize inbred line Huangzao 4 against the leaf spot pathogen (C. lunata). Meanwhile, The Thc6 mutants were found to affect the resistance of maize inbred line Huangzao 4 against C. lunata by enhancing the activation of jasmonate-responsive genes expression. Liquid chromatography-mass spectrometry (LC-MS) data further confirmed that the concentration of jasmonate in the induced maize exhibits a parallel change tendency with the expression level of defense-related genes. Hence, the Thc6 gene could be participated in the induced resistance of maize inbred line Huangzao 4 against C. lunata infection through a jasmonic acid-dependent pathway. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Purification, gene cloning, gene expression, and mutants of Dps from the obligate anaerobe Porphyromonas gingivalis.

    Science.gov (United States)

    Ueshima, Junichi; Shoji, Mikio; Ratnayake, Dinath B; Abe, Kihachiro; Yoshida, Shinichi; Yamamoto, Kenji; Nakayama, Koji

    2003-03-01

    The periodontopathogen Porphyromonas gingivalis is an obligate anaerobe that is devoid of catalase but exhibits a relatively high degree of resistance to peroxide stress. In the present study, we demonstrate that P. gingivalis contains a Dps homologue that plays an important role in the protection of cells from peroxide stress. The Dps protein isolated from P. gingivalis displayed a ferritin-like spherical polymer consisting of 19-kDa subunits. Molecular cloning and sequencing of the gene encoding this protein revealed that it had a high similarity in nucleotide and amino acid sequences to Dps proteins from other species. The expression of Dps was significantly increased by exposure of P. gingivalis to atmospheric oxygen in an OxyR-dependent manner, indicating that it is regulated by the reactive oxygen species-regulating gene oxyR. The Dps-deficient mutants, including the dps single mutant and the ftn dps double mutant, showed no viability loss upon exposure to atmospheric oxygen for 6 h. In contrast to the wild type, however, these mutants exhibited the high susceptibility to hydrogen peroxide, thereby disrupting the viability. On the other hand, no significant difference in sensitivity to mitomycin C and metronidazole was observed between the wild type and the mutants. Furthermore, the dps single mutant, compared with the wild type, showed a lower viability in infected human umbilical vein endothelial cells.

  4. Human duodenal motor activity in response to acid and different nutrients

    NARCIS (Netherlands)

    Schwartz, M. P.; Samsom, M.; Smout, A. J.

    2001-01-01

    Duodenal motor activity in response to intraduodenal infusion of small volumes of acid and nutrients of different chemical composition was studied in 10 healthy humans, using a water-perfused catheter incorporating 20 antropyloroduodenal sideholes. Saline and dextrose did not affect motility. Acid

  5. RAF Suppression Synergizes with MEK Inhibition in KRAS Mutant Cancer Cells

    Directory of Open Access Journals (Sweden)

    Simona Lamba

    2014-09-01

    Full Text Available KRAS is the most frequently mutated oncogene in human cancer, yet no therapies are available to treat KRAS mutant cancers. We used two independent reverse genetic approaches to identify components of the RAS-signaling pathways required for growth of KRAS mutant tumors. Small interfering RNA (siRNA screening of 37 KRAS mutant colorectal cancer cell lines showed that RAF1 suppression was synthetic lethal with MEK inhibition. An unbiased kinome short hairpin RNA (shRNA-based screen confirmed this synthetic lethal interaction in colorectal as well as in lung cancer cells bearing KRAS mutations. Compounds targeting RAF kinases can reverse resistance to the MEK inhibitor selumetinib. MEK inhibition induces RAS activation and BRAF-RAF1 dimerization and sustains MEK-ERK signaling, which is responsible for intrinsic resistance to selumetinib. Prolonged dual blockade of RAF and MEK leads to persistent ERK suppression and efficiently induces apoptosis. Our data underlie the relevance of developing combinatorial regimens of drugs targeting the RAF-MEK pathway in KRAS mutant tumors.

  6. The Walker A motif mutation recA4159 abolishes the SOS response and recombination in a recA730 mutant of Escherichia coli.

    Science.gov (United States)

    Šimatović, Ana; Mitrikeski, Petar T; Vlašić, Ignacija; Sopta, Mary; Brčić-Kostić, Krunoslav

    2016-01-01

    In bacteria, the RecA protein forms recombinogenic filaments required for the SOS response and DNA recombination. In order to form a recombinogenic filament, wild type RecA needs to bind ATP and to interact with mediator proteins. The RecA730 protein is a mutant version of RecA with superior catalytic abilities, allowing filament formation without the help of mediator proteins. The mechanism of RecA730 filament formation is not well understood, and the question remains as to whether the RecA730 protein requires ATP binding in order to become competent for filament formation. We examined two mutants, recA730,4159 (presumed to be defective for ATP binding) and recA730,2201 (defective for ATP hydrolysis), and show that they have different properties with respect to SOS induction, conjugational recombination and double-strand break repair. We show that ATP binding is essential for all RecA730 functions, while ATP hydrolysis is required only for double-strand break repair. Our results emphasize the similarity of the SOS response and conjugational recombination, neither of which requires ATP hydrolysis by RecA730. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

  7. [Physiological response to acetic acid stress of Acetobacter pasteuranus during vinegar fermentation].

    Science.gov (United States)

    Qi, Zhengliang; Yang, Hailin; Xia, Xiaole; Wang, Wu; Leng, Yunwei; Yu, Xiaobin; Quan, Wu

    2014-03-04

    The aim of the study is to propose a dynamic acetic acid resistance mechanism through analysis on response of cellular morphology, physiology and metabolism of A. pasteurianus CICIM B7003 during vinegar fermentation. Vinegar fermentation was carried out in a Frings 9 L acetator by strain B7003 and cultures were sampled at different cellular growth phases. Simultaneously, percentage of capsular polysaccharide versus dry cells weight, ratio of unsaturated fatty acids to saturated fatty acids, transcription of acetic acid resistance genes, activity of alcohol respiratory chain enzymes and ATPase were detected for these samples to assay the responses of bacterial morphology, physiology and metabolism. When acetic acid was existed, no obvious capsular polysaccharide was secreted by cells. As vinegar fermentation proceeding, percentage of capsular polysaccharide versus dry cells weight was reduced from 2.5% to 0.89%. Ratio of unsaturated fatty acids to saturated fatty acids was increased obviously which can improve membrane fluidity. Also transcription level of acetic acid resistance genes was promoted. Interestingly, activity of alcohol respiratory chain and ATPase was not inhibited but promoted obviously with acetic acid accumulation which could provide enough energy for acetic acid resistance mechanism. On the basis of the results obtained from the experiment, A. pasteurianus CICIM B7003 relies mainly on the cooperation of changes of extracellular capsular polysaccharide and membrane fatty acids, activation of acid resistance genes transcription, enhancement of activity of alcohol respiratory chain and rapid energy production to tolerate acidic environment.

  8. Activity of Lactobacillus casei and its gamma-radiation induced mutant in different types of milk

    International Nuclear Information System (INIS)

    Singh, J.; Ranganathan, B.

    1979-01-01

    Lactobacillus casei (RTS) and one of its gamma-radiation induced mutants, selected on the basis of increased proteolytic activity were individually inoculated in skimmed milk samples of different species. After incubation at 37 0 C for 24 hours, both the cultures produced higher titratable and volatile acidities and liberated more tyrosine in buffalo's milk as compared to either cow's or goat's milk. These cultures did not produce diacetyl or acetoin in different types of milk. It was further observed that the mutant was biochemically more active as compared to the parent culture. L. casei (RTS), irrespective of milk of different species. (orig.) [de

  9. Genetic fingerprinting of mutant rose cultivars

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, S; Prasad, K V; Singh, K P; Singh, A.P. [Division of Floriculture and Landscaping, Indian Agricultural Research Institute, Pusa, New Delhi (India)], E-mail: kvprasad66@gmail.com

    2008-07-01

    Six rose mutants evolved at the Indian Agricultural Research Institute, New Delhi from four parent cultivars were characterized based on RAPD markers. Contrary to the earlier findings our effort has conclusively proven that the RAPD markers are indeed robust tools to discern the mutants from their parents. Among 40 primers screened, 7 primers produced inconsistent banding pattern. The number of polymorphic bands varied between 4 (OPA 14) and 10 (OPA1) with an average of 6.5 bands per primer. The percentage polymorphism ranged from 62.5 (OPM 9) to 100 percent (OPA 1). Most of the primers produced monomorphic bands between parent and mutant rose cultivars. When primer OPA 2 was used a specific band of 2.5 kb was noticed in mutant cv. Pusa Urmil and cv. Pusa Abhishek but was absent in parent cv. Jantar Mantar. A polymorphic band of 750 bp was noticed in the parent Kiss of Fire and helped in differentiating the parent from its mutant when amplified with OPK 3. Primer OPS 16 produced discriminatory band of 800 bp in mutant cv. Pink Sport of Montezuma while it was absent in its parent cv. Montezuma. Another specific band of 650 bp was present in parent cv. Montezuma and absent in its mutant cv. Pink Sport of Montezuma signifying the uniqueness of the mutant. Primer OPM 5 brought out distinct polymorphism among the parent Jantar Mantar and its three mutants with absence of a specific band of 1.5 kb in the parent. The four parents and 6 mutants were divided into four distinct groups in the Dendogram constructed by UPGMA method. The most genetically similar cultivar among the 10 cultivars analyzed are Montezuma and its pink sport of Montezuma whereas Abhisarika a mutant of cv. Kiss of Fire was distinctly different and formed a separate cluster. (author)

  10. Pimelic acid, the first precursor of the Bacillus subtilis biotin synthesis pathway, exists as the free acid and is assembled by fatty acid synthesis.

    Science.gov (United States)

    Manandhar, Miglena; Cronan, John E

    2017-05-01

    Biotin synthetic pathways are readily separated into two stages, synthesis of the seven carbon α, ω-dicarboxylic acid pimelate moiety and assembly of the fused heterocyclic rings. The biotin pathway genes responsible for pimelate moiety synthesis vary widely among bacteria whereas the ring synthesis genes are highly conserved. Bacillus subtilis seems to have redundant genes, bioI and bioW, for generation of the pimelate intermediate. Largely consistent with previous genetic studies it was found that deletion of bioW caused a biotin auxotrophic phenotype whereas deletion of bioI did not. BioW is a pimeloyl-CoA synthetase that converts pimelic acid to pimeloyl-CoA. The essentiality of BioW for biotin synthesis indicates that the free form of pimelic acid is an intermediate in biotin synthesis although this is not the case in E. coli. Since the origin of pimelic acid in Bacillus subtilis is unknown, 13 C-NMR studies were carried out to decipher the pathway for its generation. The data provided evidence for the role of free pimelate in biotin synthesis and the involvement of fatty acid synthesis in pimelate production. Cerulenin, an inhibitor of the key fatty acid elongation enzyme, FabF, markedly decreased biotin production by B. subtilis resting cells whereas a strain having a cerulenin-resistant FabF mutant produced more biotin. In addition, supplementation with pimelic acid fully restored biotin production in cerulenin-treated cells. These results indicate that pimelic acid originating from fatty acid synthesis pathway is a bona fide precursor of biotin in B. subtilis. © 2017 John Wiley & Sons Ltd.

  11. The Arabidopsis thaliana RNA editing factor SLO2, which affects the mitochondrial electron transport chain, participates in multiple stress and hormone responses.

    Science.gov (United States)

    Zhu, Qiang; Dugardeyn, Jasper; Zhang, Chunyi; Mühlenbock, Per; Eastmond, Peter J; Valcke, Roland; De Coninck, Barbara; Oden, Sevgi; Karampelias, Michael; Cammue, Bruno P A; Prinsen, Els; Van Der Straeten, Dominique

    2014-02-01

    Recently, we reported that the novel mitochondrial RNA editing factor SLO2 is essential for mitochondrial electron transport, and vital for plant growth through regulation of carbon and energy metabolism. Here, we show that mutation in SLO2 causes hypersensitivity to ABA and insensitivity to ethylene, suggesting a link with stress responses. Indeed, slo2 mutants are hypersensitive to salt and osmotic stress during the germination stage, while adult plants show increased drought and salt tolerance. Moreover, slo2 mutants are more susceptible to Botrytis cinerea infection. An increased expression of nuclear-encoded stress-responsive genes, as well as mitochondrial-encoded NAD genes of complex I and genes of the alternative respiratory pathway, was observed in slo2 mutants, further enhanced by ABA treatment. In addition, H2O2 accumulation and altered amino acid levels were recorded in slo2 mutants. We conclude that SLO2 is required for plant sensitivity to ABA, ethylene, biotic, and abiotic stress. Although two stress-related RNA editing factors were reported very recently, this study demonstrates a unique role of SLO2, and further supports a link between mitochondrial RNA editing events and stress response.

  12. From one body mutant to one cell mutant. A progress of radiation breeding in crops

    International Nuclear Information System (INIS)

    Nagatomi, Shigeki

    1996-01-01

    An effective method was established to obtain non-chimeral mutants with wide spectrum of flower colors, regenerated from floral organs on which mutated sectors were come out on chronic irradiated plants. By this way, six mutant varieties of flower colors have been selected from one pink flower of chrysanthemum, and cultivated for cut-flower production. By the same method, 3 mutant varieties with small and spray type flowers were selected in Eustoma. Mutant varieties such as a rust disease resistant in sugarcane, 6 dwarfs in Cytisus and pure-white mushroom in velvet shank have been selected successively for short period. (J.P.N.)

  13. RodZ and PgsA Play Intertwined Roles in Membrane Homeostasis of Bacillus subtilis and Resistance to Weak Organic Acid Stress

    NARCIS (Netherlands)

    van Beilen, Johan; Blohmke, Christoph J.; Folkerts, Hendrik; de Boer, Richard; Zakrzewska, Anna; Kulik, Wim; Vaz, Fred M.; Brul, Stanley; Ter Beek, Alexander

    2016-01-01

    Weak organic acids like sorbic and acetic acid are widely used to prevent growth of spoilage organisms such as Bacilli. To identify genes involved in weak acid stress tolerance we screened a transposon mutant library of Bacillus subtilis for sorbic acid sensitivity. Mutants of the rodZ (ymfM) gene

  14. Changes in transcript levels of starch hydrolysis genes and raising citric acid production via carbon ion irradiation mutagenesis of Aspergillus niger.

    Directory of Open Access Journals (Sweden)

    Wei Hu

    Full Text Available The filamentous ascomycete Aspergillus niger is well known for its ability to accumulate citric acid for the hydrolysis of starchy materials. To improve citric acid productivity, heavy ion beam mutagenesis was utilized to produce mutant A.niger strains with enhanced production of citric acid in this work. It was demonstrated that a mutant HW2 with high concentration of citric acid was isolated after carbon ion irradiation with the energy of 80Mev/μ, which was obvious increase higher than the original strain from liquefied corn starch as a feedstock. More importantly, with the evidence from the expression profiles of key genes and enzyme activity involved in the starch hydrolysis process between original strain and various phenotype mutants, our results confirmed that different transcript levels of key genes involving in starch hydrolysis process between original strain and mutants could be a significant contributor to different citric acid concentration in A.niger, such as, amyR and glaA, which therefore opened a new avenue for constructing genetically engineered A.niger mutants for high-yield citric acid accumulation in the future. As such, this work demonstrated that heavy ion beam mutagenesis presented an efficient alternative strategy to be developed to generate various phenotype microbe species mutants for functional genes research.

  15. Changes in transcript levels of starch hydrolysis genes and raising citric acid production via carbon ion irradiation mutagenesis of Aspergillus niger.

    Science.gov (United States)

    Hu, Wei; Li, Wenjian; Chen, Hao; Liu, Jing; Wang, Shuyang; Chen, Jihong

    2017-01-01

    The filamentous ascomycete Aspergillus niger is well known for its ability to accumulate citric acid for the hydrolysis of starchy materials. To improve citric acid productivity, heavy ion beam mutagenesis was utilized to produce mutant A.niger strains with enhanced production of citric acid in this work. It was demonstrated that a mutant HW2 with high concentration of citric acid was isolated after carbon ion irradiation with the energy of 80Mev/μ, which was obvious increase higher than the original strain from liquefied corn starch as a feedstock. More importantly, with the evidence from the expression profiles of key genes and enzyme activity involved in the starch hydrolysis process between original strain and various phenotype mutants, our results confirmed that different transcript levels of key genes involving in starch hydrolysis process between original strain and mutants could be a significant contributor to different citric acid concentration in A.niger, such as, amyR and glaA, which therefore opened a new avenue for constructing genetically engineered A.niger mutants for high-yield citric acid accumulation in the future. As such, this work demonstrated that heavy ion beam mutagenesis presented an efficient alternative strategy to be developed to generate various phenotype microbe species mutants for functional genes research.

  16. Changes in transcript levels of starch hydrolysis genes and raising citric acid production via carbon ion irradiation mutagenesis of Aspergillus niger

    Science.gov (United States)

    Li, Wenjian; Chen, Hao; Liu, Jing; Wang, Shuyang; Chen, Jihong

    2017-01-01

    The filamentous ascomycete Aspergillus niger is well known for its ability to accumulate citric acid for the hydrolysis of starchy materials. To improve citric acid productivity, heavy ion beam mutagenesis was utilized to produce mutant A.niger strains with enhanced production of citric acid in this work. It was demonstrated that a mutant HW2 with high concentration of citric acid was isolated after carbon ion irradiation with the energy of 80Mev/μ, which was obvious increase higher than the original strain from liquefied corn starch as a feedstock. More importantly, with the evidence from the expression profiles of key genes and enzyme activity involved in the starch hydrolysis process between original strain and various phenotype mutants, our results confirmed that different transcript levels of key genes involving in starch hydrolysis process between original strain and mutants could be a significant contributor to different citric acid concentration in A.niger, such as, amyR and glaA, which therefore opened a new avenue for constructing genetically engineered A.niger mutants for high-yield citric acid accumulation in the future. As such, this work demonstrated that heavy ion beam mutagenesis presented an efficient alternative strategy to be developed to generate various phenotype microbe species mutants for functional genes research. PMID:28650980

  17. Multiple Acid Sensors Control Helicobacter pylori Colonization of the Stomach.

    Science.gov (United States)

    Huang, Julie Y; Goers Sweeney, Emily; Guillemin, Karen; Amieva, Manuel R

    2017-01-01

    Helicobacter pylori's ability to respond to environmental cues in the stomach is integral to its survival. By directly visualizing H. pylori swimming behavior when encountering a microscopic gradient consisting of the repellent acid and attractant urea, we found that H. pylori is able to simultaneously detect both signals, and its response depends on the magnitudes of the individual signals. By testing for the bacteria's response to a pure acid gradient, we discovered that the chemoreceptors TlpA and TlpD are each independent acid sensors. They enable H. pylori to respond to and escape from increases in hydrogen ion concentration near 100 nanomolar. TlpD also mediates attraction to basic pH, a response dampened by another chemoreceptor TlpB. H. pylori mutants lacking both TlpA and TlpD (ΔtlpAD) are unable to sense acid and are defective in establishing colonization in the murine stomach. However, blocking acid production in the stomach with omeprazole rescues ΔtlpAD's colonization defect. We used 3D confocal microscopy to determine how acid blockade affects the distribution of H. pylori in the stomach. We found that stomach acid controls not only the overall bacterial density, but also the microscopic distribution of bacteria that colonize the epithelium deep in the gastric glands. In omeprazole treated animals, bacterial abundance is increased in the antral glands, and gland colonization range is extended to the corpus. Our findings indicate that H. pylori has evolved at least two independent receptors capable of detecting acid gradients, allowing not only survival in the stomach, but also controlling the interaction of the bacteria with the epithelium.

  18. Decreased levels of free D-aspartic acid in the forebrain of serine racemase (Srr) knock-out mice.

    Science.gov (United States)

    Horio, Mao; Ishima, Tamaki; Fujita, Yuko; Inoue, Ran; Mori, Hisashi; Hashimoto, Kenji

    2013-05-01

    d-Serine, an endogenous co-agonist of the N-methyl-d-aspartate (NMDA) receptor is synthesized from l-serine by serine racemase (SRR). A previous study of Srr knockout (Srr-KO) mice showed that levels of d-serine in forebrain regions, such as frontal cortex, hippocampus, and striatum, but not cerebellum, of mutant mice are significantly lower than those of wild-type (WT) mice, suggesting that SRR is responsible for d-serine production in the forebrain. In this study, we attempted to determine whether SRR affects the level of other amino acids in brain tissue. We found that tissue levels of d-aspartic acid in the forebrains (frontal cortex, hippocampus and striatum) of Srr-KO mice were significantly lower than in WT mice, whereas levels of d-aspartic acid in the cerebellum were not altered. Levels of d-alanine, l-alanine, l-aspartic acid, taurine, asparagine, arginine, threonine, γ-amino butyric acid (GABA) and methionine, remained the same in frontal cortex, hippocampus, striatum and cerebellum of WT and mutant mice. Furthermore, no differences in d-aspartate oxidase (DDO) activity were detected in the forebrains of WT and Srr-KO mice. These results suggest that SRR and/or d-serine may be involved in the production of d-aspartic acid in mouse forebrains, although further detailed studies will be necessary to confirm this finding. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. Biocontrol potential of salinity tolerant mutants of Trichoderma harzianum against Fusarium oxysporum Potencial de biocontrole de mutantes sal-tolerantes de Trichoderma harzianum contra Fusarium oxysporum

    Directory of Open Access Journals (Sweden)

    Hassan Abdel-Latif A. Mohamed

    2006-06-01

    Full Text Available Exposing a wild-type culture of Trichoderma harzianum to gamma irradiation induced two stable salt-tolerant mutants (Th50M6 and Th50M11. Under saline conditions, both mutants greatly surpassed their wild type strain in growth rate, sporulation and biological proficiency against Fusarium oxysporum, the causal agent of tomato wilt disease. Tolerant T. harzianum mutants detained a capability to grow and convinced sporulation in growth media containing up to 69 mM NaCl. In comparison with their parent strain, characterization of both mutants confirmed that they have reinforced contents of proline and hydroxyproline, relatively higher sodium content compared to potassium, calcium or magnesium contents, higher level of total phenols. Electrophoretic analysis of total soluble proteins in the salt tolerance mutant Th50M6 showed different bands accumulated in response to 69 mM NaCl. Data also showed that mutants produce certain active metabolites, such as chitinases, cellulases, beta-galactosidases, as well as, some antibiotics i.e., trichodermin, gliotoxin and gliovirin. Trichoderma mutants significantly reduced wilt disease incidence and improved yield and mineral contents of tomato plants under both saline and non-saline soil conditions, as well as, under infested and natural conditions. T. harzianum mutants were also more efficient in dropping the F. oxysporum growth in rhizosphere compared to the wild type strain. Population density of both mutants in rhizosphere far exceeded that of T. harzianum wild type strain.A exposição de uma cepa selvagem de Trichoderma harzianum à irradiação gama induziu dois mutantes tolerantes a sal (Th50M6 e Th50M11. Em condições salinas, os dois mutantes foram muito superiores à cepa selvagem em relação à velocidade de multiplicação, esporulação e eficiência contra Fusarium oxysporum, o agente causador da doença wilt do tomate. Os mutantes tolerantes foram capazes de multiplicação e esporulação em

  20. The inactivation of RNase G reduces the Stenotrophomonas maltophilia susceptibility to quinolones by triggering the heat shock response.

    Directory of Open Access Journals (Sweden)

    Alejandra eBernardini

    2015-10-01

    Full Text Available Quinolone resistance is usually due to mutations in the genes encoding bacterial topoisomerases. However different reports have shown that neither clinical quinolone resistant isolates nor in vitro obtained S. maltophilia mutants present mutations in such genes. The mechanisms so far described consist on efflux pumps' overexpression. Our objective is to get information on novel mechanisms of S. maltophilia quinolone resistance. For this purpose, a transposon-insertion mutant library was obtained in S. maltophilia D457.. One mutant presenting reduced susceptibility to nalidixic acid was selected. Inverse PCR showed that the inactivated gene encodes RNase G. Complementation of the mutant with wild-type RNase G allele restored the susceptibility to quinolones. Transcriptomic and real-time RT-PCR analyses showed that several genes encoding heat-shock response proteins were expressed at higher levels in the RNase defective mutant than in the wild-type strain. In agreement with this situation, heat-shock reduces the S. maltophilia susceptibility to quinolone. We can then conclude that the inactivation of the RNase G reduces the susceptibility of S. maltophilia to quinolones, most likely by regulating the expression of heat-shock response genes. Heat-shock induces a transient phenotype of quinolone resistance in S. maltophilia.