Mutational analysis of GlnB residues critical for NifA activation in Azospirillum brasilense.

Science.gov (United States)

Inaba, Juliana; Thornton, Jeremy; Huergo, Luciano Fernandes; Monteiro, Rose Adele; Klassen, Giseli; Pedrosa, Fábio de Oliveira; Merrick, Mike; de Souza, Emanuel Maltempi

2015-02-01

PII proteins are signal transduction that sense cellular nitrogen status and relay this signals to other targets. Azospirillum brasilense is a nitrogen fixing bacterium, which associates with grasses and cereals promoting beneficial effects on plant growth and crop yields. A. brasilense contains two PII encoding genes, named glnB and glnZ. In this paper, glnB was mutagenised in order to identify amino acid residues involved in GlnB signaling. Two variants were obtained by random mutagenesis, GlnBL13P and GlnBV100A and a site directed mutant, GlnBY51F, was obtained. Their ability to complement nitrogenase activity of glnB mutant strains of A. brasilense were determined. The variant proteins were also overexpressed in Escherichia coli, purified and characterized biochemically. None of the GlnB variant forms was able to restore nitrogenase activity in glnB mutant strains of A. brasilense LFH3 and 7628. The purified GlnBY51F and GlnBL13P proteins could not be uridylylated by GlnD, whereas GlnBV100A was uridylylated but at only 20% of the rate for wild type GlnB. Biochemical and computational analyses suggest that residue Leu13, located in the α helix 1 of GlnB, is important to maintain GlnB trimeric structure and function. The substitution V100A led to a lower affinity for ATP binding. Together the results suggest that NifA activation requires uridylylated GlnB bound to ATP. Copyright © 2014 Elsevier GmbH. All rights reserved.

Double heterozygous mutations Gln100Leu and His348Gln of the F7 gene in a patient with factor VII deficiency.

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Li, Min; Zheng, Fangxiu; Jin, Yanhui; Wang, Mingshan; Zhu, Liqing; Yang, Lihong

2013-03-01

A 25-year-old Chinese woman who had a history of easy bruising was admitted to hospital due to uncontrolled epistaxis. She showed factor VII activity level of 2% and factor VII antigen level of 4% of the normal value. We detected a novel missense mutation g.8355 A>T (p.Gln100Leu) in the second epidermal growth factor-like (EGF) domain and a g.11482 T>G (p.His348Gln) in the catalytic domain. Although the Gln100 residue is close to the junction of EGF-2 domain with the serine protease domain, we infer that the substitution of polar negatively charged Gln residue at the position 100 with introduction of nonpolar Leu residue may be likely to perturb proper folding, resulting in decreasing factor VII activity.

The IL23R A/Gln381 allele promotes IL-23 unresponsiveness in human memory T-helper 17 cells and impairs Th17 responses in psoriasis patients.

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Di Meglio, Paola; Villanova, Federica; Napolitano, Luca; Tosi, Isabella; Terranova Barberio, Manuela; Mak, Rose K; Nutland, Sarah; Smith, Catherine H; Barker, Jonathan N W N; Todd, John A; Nestle, Frank O

2013-10-01

We and others have shown that the minor, nonconserved allele Gln381 of the Arg381Gln single-nucleotide polymorphism (rs11209026G>A) of the IL-23 receptor gene (IL23R) protects against psoriasis. Moreover, we have recently shown impaired IL-23-induced IL-17A production and STAT-3 phosphorylation in Th17 cells generated in vitro from healthy individuals heterozygous for the protective A allele (GA). However, the biological effect of this variant has not been determined in homozygous carriers of the protective A allele (AA), nor in psoriatic patients. Here we expand our functional investigation of the IL23R Arg381Gln gene variant to include AA homozygous individuals. By using isolated memory CD4+ T cells, we found attenuated IL-23-induced Th17 response in heterozygous individuals. Moreover, we found that AA homozygous individuals were strikingly unresponsive to IL-23, with minimal or no IL-17A and IL-17F production and failure of human memory Th17 cell survival/expansion. Finally, IL-23-induced Th17 response was also attenuated in age- and sex-matched GA versus GG psoriatic patients undergoing systemic treatment. Taken together, our data provide evidence for an allele-dosage effect for IL-23R Gln381 and indicate that common gene alleles associated with complex diseases might have biological effects of considerable magnitude in homozygous carriers.

The Transcriptional Repressor, MtrR, of the mtrCDE Efflux Pump Operon of Neisseria gonorrhoeae Can Also Serve as an Activator of “off Target” Gene (glnE Expression

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Paul J. T. Johnson

2015-06-01

Full Text Available MtrR is a well-characterized repressor of the Neisseria gonorrhoeae mtrCDE efflux pump operon. However, results from a previous transcriptional profiling study suggested that MtrR also represses or activates expression of at least sixty genes outside of the mtr locus. Evidence that MtrR can directly repress so-called “off target” genes has previously been reported; in particular, MtrR was shown to directly repress glnA, which encodes glutamine synthetase. In contrast, evidence for the ability of MtrR to directly activate expression of gonococcal genes has been lacking; herein, we provide such evidence. We now report that MtrR has the ability to directly activate expression of glnE, which encodes the dual functional adenyltransferase/deadenylase enzyme GlnE that modifies GlnA resulting in regulation of its role in glutamine biosynthesis. With its capacity to repress expression of glnA, the results presented herein emphasize the diverse and often opposing regulatory properties of MtrR that likely contributes to the overall physiology and metabolism of N. gonorrhoeae.

No association of the Arg51Gln and Leu72Met polymorphisms of the ghrelin gene and polycystic ovary syndrome.

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Wang, Kehua; Wang, Leiguang; Zhao, Yueran; Shi, Yuhua; Wang, Laicheng; Chen, Zi-Jiang

2009-02-01

Ghrelin plays a role in regulating glucose metabolism and energy balance. Polymorphisms in preproghrelin and ghrelin gene could be responsible for obesity, insulin resistance and low ghrelin levels observed in some individuals. The objective of this study was to evaluate the influence of two single-nucleotide polymorphisms (SNPs) of ghrelin gene on the clinical, the hormonal and metabolic features in women with polycystic ovary syndrome (PCOS) in a Chinese population. A large sample of Chinese PCOS (n = 271) women and a control group (n = 296) of healthy women matched for age were studied. Hormone and metabolic profiles were measured and blood samples were collected for genotype and allelic frequency analysis. Non-synonymous SNPs in the coding region (exon 2) of the preproghrelin gene (Arg51Gln (346 G>A) and Leu72Met (408 C>A) were studied using PCR and restriction fragment length polymorphism analysis. The polymorphism Arg51Gln was not found in the cohorts studied. The distribution of Leu72Met was similar in PCOS group and in healthy controls. There was no significant difference in age, BMI, waist-hip-ratio and levels of FSH, LH, estradiol, testosterone and prolactin between PCOS patients with different genotypes, and the level of plasma glucose and insulin was also similar. No association was found between Leu72Met and Arg51Gln polymorphisms in the ghrelin gene and PCOS in Chinese population.

The separation of Gln and Glu in STEAM: a comparison study using short and long TEs/TMs at 3 and 7 T.

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Dou, Weiqiang; Kaufmann, Jörn; Li, Meng; Zhong, Kai; Walter, Martin; Speck, Oliver

2015-08-01

This study aimed to determine the optimal echo time (TE) and mixing time (TM) for in vivo glutamine (Gln) and glutamate (Glu) separation in stimulated-echo acquisition mode at 3 and 7 T. We applied a short TE/TM (20/10 ms) for a high signal-to-noise-ratio and a field-specific long TE/TM (3 T: 72/6 ms; 7 T: 74/68 ms) for optimal Gln and Glu separation of the Carbon-4 proton resonances. Corresponding Gln and Glu spectra were simulated using VeSPA software, and measured in a phantom and human brains at 3 and 7 T. Higher spectral separation for Gln and Glu was achieved at 7 than 3 T. At 7 T, short TE/TM provided comparable spectral separation and in vitro Gln and Glu quantification compared to long TE/TM. Moreover, it showed greater reliability in in vivo Gln and Glu detection and separation than long TE/TM, with significantly lower Cramer-Rao lower bounds (Gln: 14.9 vs. 75.8; Glu: 3.8 vs. 6.5) and correlation between Gln and Glu (p = 0.004). Based on the optimal separation for Gln and Glu, a short TE/TM at 7 T is proposed for future in vivo Gln and Glu acquisition.

Analysis of Gln223Agr Polymorphism of Leptin Receptor Gene in Type II Diabetic Mellitus Subjects among Malaysians

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Chong Pei Pei

2013-09-01

Full Text Available Leptin is known as the adipose peptide hormone. It plays an important role in the regulation of body fat and inhibits food intake by its action. Moreover, it is believed that leptin level deductions might be the cause of obesity and may play an important role in the development of Type 2 Diabetes Mellitus (T2DM, as well as in cardiovascular diseases (CVD. The Leptin Receptor (LEPR gene and its polymorphisms have not been extensively studied in relation to the T2DM and its complications in various populations. In this study, we have determined the association of Gln223Agr loci of LEPR gene in three ethnic groups of Malaysia, namely: Malays, Chinese and Indians. A total of 284 T2DM subjects and 281 healthy individuals were recruited based on International Diabetes Federation (IDF criteria. Genomic DNA was extracted from the buccal specimens of the subjects. The commercial polymerase chain reaction (PCR method was carried out by proper restriction enzyme MSP I to both amplify and digest the Gln223Agr polymorphism. The p-value among the three studied races was 0.057, 0.011 and 0.095, respectively. The values such as age, WHR, FPG, HbA1C, LDL, HDL, Chol and Family History were significantly different among the subjects with Gln223Agr polymorphism of LEPR (p < 0.05.

Analysis of Gln223Agr polymorphism of Leptin Receptor Gene in type II diabetic mellitus subjects among Malaysians.

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Etemad, Ali; Ramachandran, Vasudevan; Pishva, Seyyed Reza; Heidari, Farzad; Aziz, Ahmad Fazli Abdul; Yusof, Ahmad Khairuddin Mohamed; Pei, Chong Pei; Ismail, Patimah

2013-09-18

Leptin is known as the adipose peptide hormone. It plays an important role in the regulation of body fat and inhibits food intake by its action. Moreover, it is believed that leptin level deductions might be the cause of obesity and may play an important role in the development of Type 2 Diabetes Mellitus (T2DM), as well as in cardiovascular diseases (CVD). The Leptin Receptor (LEPR) gene and its polymorphisms have not been extensively studied in relation to the T2DM and its complications in various populations. In this study, we have determined the association of Gln223Agr loci of LEPR gene in three ethnic groups of Malaysia, namely: Malays, Chinese and Indians. A total of 284 T2DM subjects and 281 healthy individuals were recruited based on International Diabetes Federation (IDF) criteria. Genomic DNA was extracted from the buccal specimens of the subjects. The commercial polymerase chain reaction (PCR) method was carried out by proper restriction enzyme MSP I to both amplify and digest the Gln223Agr polymorphism. The p-value among the three studied races was 0.057, 0.011 and 0.095, respectively. The values such as age, WHR, FPG, HbA1C, LDL, HDL, Chol and Family History were significantly different among the subjects with Gln223Agr polymorphism of LEPR (p < 0.05).

XRCC1 Arg194Trp and Arg399Gln polymorphisms and arsenic methylation capacity are associated with urothelial carcinoma

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Chiang, Chien-I [School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan (China); Huang, Ya-Li [Department of Public Health, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China); Chen, Wei-Jen [School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan (China); Shiue, Horng-Sheng [Department of Chinese Medicine, Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Taoyuan, Taiwan (China); Huang, Chao-Yuan; Pu, Yeong-Shiau [Department of Urology, National Taiwan University Hospital, College of Medicine National Taiwan University, Taipei, Taiwan (China); Lin, Ying-Chin [Department of Family Medicine, Shung Ho Hospital, Taipei Medical University, New Taipei, Taiwan (China); Department of Health Examination, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan (China); Division of Family Medicine, School of Medicine, Taipei Medical University, Taipei, Taiwan (China); Hsueh, Yu-Mei, E-mail: ymhsueh@tmu.edu.tw [School of Public Health, College of Public Health and Nutrition, Taipei Medical University, Taipei, Taiwan (China); Department of Public Health, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan (China)

2014-09-15

The association between DNA repair gene polymorphisms and bladder cancer has been widely studied. However, few studies have examined the correlation between urothelial carcinoma (UC) and arsenic or its metabolites. The aim of this study was to examine the association between polymorphisms of the DNA repair genes, XRCC1 Arg194Trp, XRCC1 Arg399Gln, XRCC3 Thr241Met, and XPD Lys751Gln, with urinary arsenic profiles and UC. To this end, we conducted a hospital-based case–control study with 324 UC patients and 647 age- and gender-matched non-cancer controls. Genomic DNA was used to examine the genotype of XRCC1 Arg194Trp, XRCC1 Arg399Gln, XRCC3 Thr241Met, and XPD Lys751Gln by PCR-restriction fragment length polymorphism analysis (PCR-RFLP). Urinary arsenic profiles were measured by high performance liquid chromatography (HPLC) linked with hydride generator and atomic absorption spectrometry. The XRCC1 399 Gln/Gln and 194 Arg/Trp and Trp/Trp genotypes were significantly related to UC, and the odds ratio (OR) and 95% confidence interval (95%CI) were 1.68 (1.03–2.75) and 0.66 (0.48–0.90), respectively. Participants with higher total urinary arsenic levels, a higher percentage of inorganic arsenic (InAs%) and a lower percentage of dimethylarsinic acid (DMA%) had a higher OR of UC. Participants carrying XRCC1 risk diplotypes G-C/G-C, A-C/A-C, and A-T/G-T, and who had higher total arsenic levels, higher InAs%, or lower DMA% compared to those with other XRCC1 diplotypes had a higher OR of UC. Our results suggest that the XRCC1 399 Gln/Gln and 194 Arg/Arg DNA repair genes play an important role in poor arsenic methylation capacity, thereby increasing the risk of UC in non-obvious arsenic exposure areas. - Highlights: • The XRCC1 399Gln/Gln genotype was significantly associated with increased OR of UC. • The XRCC1 194 Arg/Trp and Trp/Trp genotype had a significantly decreased OR of UC. • Combined effect of the XRCC1 genotypes and poor arsenic methylation capacity on

Reciprocal Regulation of GlnR and PhoP in Response to Nitrogen and Phosphate Limitations in Saccharopolyspora erythraea

Science.gov (United States)

Yao, Li-li

2015-01-01

Nitrogen and phosphate source sensing, uptake, and assimilation are essential for the growth and development of microorganisms. In this study, we demonstrated that SACE_6965 encodes the phosphate regulator PhoP, which controls the transcription of genes involved in phosphate metabolism in the erythromycin-producing Saccharopolyspora erythraea. We found that PhoP and the nitrogen regulator GlnR both regulate the transcription of glnR as well as other nitrogen metabolism-related genes. Interestingly, both GlnR- and PhoP-binding sites were identified in the phoP promoter region. Unlike the nonreciprocal regulation of GlnR and PhoP observed in Streptomyces coelicolor and Streptomyces lividans, GlnR negatively controls the transcription of the phoP gene in S. erythraea. This suggests that GlnR directly affects phosphate metabolism and demonstrates that the cross talk between GlnR and PhoP is reciprocal. Although GlnR and PhoP sites in the glnR and phoP promoter regions are located in close proximity to one another (separated by only 2 to 4 bp), the binding of both regulators to their respective region was independent and noninterfering. These results indicate that two regulators could separately bind to their respective binding sites and control nitrogen and phosphate metabolism in response to environmental changes. The reciprocal cross talk observed between GlnR and PhoP serves as a foundation for understanding the regulation of complex primary and secondary metabolism in antibiotic-producing actinomycetes. PMID:26519391

TOLL-LIKE RECEPTOR 7 GENE Gln11Leu MISSENSEMUTATION AND SUSCEPTIBILITY TO PSORIASIS

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E. S. Galimova

2017-01-01

Full Text Available Toll-like receptor (TLR are responsible for recognizing various molecular patterns associated with pathogens. Their expression have been detected in skin cells such as keratinocytes and melanocytes. Numerous experimental studies demonstrate the key role of TLRs in the pathogenesis of immune diseases, including psoriasis. The objective of this study is to analyze the associations of polymorphisms in TLR7 gene and the risk of psoriasis development. DNA samples were collected from 138 patients with psoriasis and 317 healthy controls. Genotyping of rs179003, rs179008, rs179020, rs850632, rs12013728 polymorphic loci in TLR7 gene was performed using the SNPlex™method (AB, USA. SNP in the TLR7 gene rs179008 (Gln11Leu was associated with psoriasis in entire psoriasis, late onset and sporadic subgroups (Рс = 0.0065, OR = 1.95; Рс = 0.0004, OR = 2.50; Рс = 0.0078, OR = 2.2, respectively. In conclusion, this study is the first to identify genetic variants of the TLR7 gene significantly associated with psoriasis. 

TOLL-LIKE RECEPTOR 7 GENE Gln11Leu MISSENSEMUTATION AND SUSCEPTIBILITY TO PSORIASIS

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E. S. Galimova

2017-01-01

Full Text Available Toll-like receptor (TLR are responsible for recognizing various molecular patterns associated with pathogens. Their expression have been detected in skin cells such as keratinocytes and melanocytes. Numerous experimental studies demonstrate the key role of TLRs in the pathogenesis of immune diseases, including psoriasis. The objective of this study is to analyze the associations of polymorphisms in TLR7 gene and the risk of psoriasis development. DNA samples were collected from 138 patients with psoriasis and 317 healthy controls. Genotyping of rs179003, rs179008, rs179020, rs850632, rs12013728 polymorphic loci in TLR7 gene was performed using the SNPlex™method (AB, USA. SNP in the TLR7 gene rs179008 (Gln11Leu was associated with psoriasis in entire psoriasis, late onset and sporadic subgroups (Рс = 0.0065, OR = 1.95; Рс = 0.0004, OR = 2.50; Рс = 0.0078, OR = 2.2, respectively. In conclusion, this study is the first to identify genetic variants of the TLR7 gene significantly associated with psoriasis.

Studies on the roles of GlnK and GlnB in regulating Klebsiella pneumoniae NifL-dependent nitrogen control.

NARCIS (Netherlands)

Arcondeguy, T.; van Heeswijk, W.C.; Merrick, M.

1999-01-01

In Klebsiella pneumoniae, nitrogen fixation (nif) genes are regulated in response to fixed nitrogen and oxygen. The activity of the nif-specific transcriptional activator NifA is modulated by NifL, which mediates both oxygen and nitrogen control. The signal transduction protein GlnK is required to

DNA Repair Gene (XRCC1 Polymorphism (Arg399Gln Associated with Schizophrenia in South Indian Population: A Genotypic and Molecular Dynamics Study.

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S P Sujitha

Full Text Available This paper depicts the first report from an Indian population on the association between the variant Arg399Gln of XRCC1 locus in the DNA repair system and schizophrenia, the debilitating disease that affects 1% of the world population. Genotypic analysis of a total of 523 subjects (260 patients and 263 controls revealed an overwhelming presence of Gln399Gln in the case subjects against the controls (P < 0.0068, indicating significant level of association of this nsSNP with schizophrenia; the Gln399 allele frequency was also perceptibly more in cases than in controls (p < 0.003; OR = 1.448. The results of the genotypic studies were further validated using pathogenicity and stability prediction analysis employing computational tools [I-Mutant Suite, iStable, PolyPhen2, SNAP, and PROVEAN], with a view toassess the magnitude of deleteriousness of the mutation. The pathogenicity analysis reveals that the nsSNP could be deleterious inasmuch as it could affect the functionality of the gene, and interfere with protein function. Molecular dynamics simulation of 60ns was performed using GROMACS to analyse structural change due to a mutation (Arg399Gln that was never examined before. RMSD, RMSF, hydrogen bonds, radius of gyration and SASA analysis showedthe existence of asignificant difference between the native and the mutant protein. The present study gives astrong indication that the XRCC1 locus deserves serious attention, as it could be a potential candidatecontributing to the etio-pathogenesis of the disease.

Uridylylation of Herbaspirillum seropedicae GlnB and GlnK proteins is differentially affected by ATP, ADP and 2-oxoglutarate in vitro.

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Bonatto, Ana C; Souza, Emanuel M; Oliveira, Marco A S; Monteiro, Rose A; Chubatsu, Leda S; Huergo, Luciano F; Pedrosa, Fábio O

2012-08-01

PII are signal-transducing proteins that integrate metabolic signals and transmit this information to a large number of proteins. In proteobacteria, PII are modified by GlnD (uridylyltransferase/uridylyl-removing enzyme) in response to the nitrogen status. The uridylylation/deuridylylation cycle of PII is also regulated by carbon and energy signals such as ATP, ADP and 2-oxoglutarate (2-OG). These molecules bind to PII proteins and alter their tridimensional structure/conformation and activity. In this work, we determined the effects of ATP, ADP and 2-OG levels on the in vitro uridylylation of Herbaspirillum seropedicae PII proteins, GlnB and GlnK. Both proteins were uridylylated by GlnD in the presence of ATP or ADP, although the uridylylation levels were higher in the presence of ATP and under high 2-OG levels. Under excess of 2-OG, the GlnB uridylylation level was higher in the presence of ATP than with ADP, while GlnK uridylylation was similar with ATP or ADP. Moreover, in the presence of ADP/ATP molar ratios varying from 10/1 to 1/10, GlnB uridylylation level decreased as ADP concentration increased, whereas GlnK uridylylation remained constant. The results suggest that uridylylation of both GlnB and GlnK responds to 2-OG levels, but only GlnB responds effectively to variation on ADP/ATP ratio.

Polynomial representations of GLn

CERN Document Server

Green, James A; Erdmann, Karin

2007-01-01

The first half of this book contains the text of the first edition of LNM volume 830, Polynomial Representations of GLn. This classic account of matrix representations, the Schur algebra, the modular representations of GLn, and connections with symmetric groups, has been the basis of much research in representation theory. The second half is an Appendix, and can be read independently of the first. It is an account of the Littelmann path model for the case gln. In this case, Littelmann's 'paths' become 'words', and so the Appendix works with the combinatorics on words. This leads to the repesentation theory of the 'Littelmann algebra', which is a close analogue of the Schur algebra. The treatment is self- contained; in particular complete proofs are given of classical theorems of Schensted and Knuth.

Polynomial representations of GLN

CERN Document Server

Green, James A

1980-01-01

The first half of this book contains the text of the first edition of LNM volume 830, Polynomial Representations of GLn. This classic account of matrix representations, the Schur algebra, the modular representations of GLn, and connections with symmetric groups, has been the basis of much research in representation theory. The second half is an Appendix, and can be read independently of the first. It is an account of the Littelmann path model for the case gln. In this case, Littelmann's 'paths' become 'words', and so the Appendix works with the combinatorics on words. This leads to the repesentation theory of the 'Littelmann algebra', which is a close analogue of the Schur algebra. The treatment is self- contained; in particular complete proofs are given of classical theorems of Schensted and Knuth.

Characterization of the orf1glnKamtB operon of Herbaspirillum seropedicae.

Science.gov (United States)

Noindorf, Lilian; Rego, Fabiane G M; Baura, Valter A; Monteiro, Rose A; Wassem, Roseli; Cruz, Leonardo M; Rigo, Liu U; Souza, Emanuel M; Steffens, Maria B R; Pedrosa, Fabio O; Chubatsu, Leda S

2006-03-01

Herbaspirillum seropedicae is an endophytic nitrogen-fixing bacterium that colonizes economically important grasses. In this organism, the amtB gene is co-transcribed with two other genes: glnK that codes for a PII-like protein and orf1 that codes for a probable periplasmatic protein of unknown function. The expression of the orf1glnKamtB operon is increased under nitrogen-limiting conditions and is dependent on NtrC. An amtB mutant failed to transport methylammonium. Post-translational control of nitrogenase was also partially impaired in this mutant, since a complete switch-off of nitrogenase after ammonium addition was not observed. This result suggests that the AmtB protein is involved in the signaling pathway for the reversible inactivation of nitrogenase in H. seropedicae.

Identification of BC005512 as a DNA damage responsive murine endogenous retrovirus of GLN family involved in cell growth regulation.

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Yuanfeng Wu

Full Text Available Genotoxicity assessment is of great significance in drug safety evaluation, and microarray is a useful tool widely used to identify genotoxic stress responsive genes. In the present work, by using oligonucleotide microarray in an in vivo model, we identified an unknown gene BC005512 (abbreviated as BC, official full name: cDNA sequence BC005512, whose expression in mouse liver was specifically induced by seven well-known genotoxins (GTXs, but not by non-genotoxins (NGTXs. Bioinformatics revealed that BC was a member of the GLN family of murine endogenous retrovirus (ERV. However, the relationship to genotoxicity and the cellular function of GLN are largely unknown. Using NIH/3T3 cells as an in vitro model system and quantitative real-time PCR, BC expression was specifically induced by another seven GTXs, covering diverse genotoxicity mechanisms. Additionally, dose-response and linear regression analysis showed that expression level of BC in NIH/3T3 cells strongly correlated with DNA damage, measured using the alkaline comet assay,. While in p53 deficient L5178Y cells, GTXs could not induce BC expression. Further functional studies using RNA interference revealed that down-regulation of BC expression induced G1/S phase arrest, inhibited cell proliferation and thus suppressed cell growth in NIH/3T3 cells. Together, our results provide the first evidence that BC005512, a member from GLN family of murine ERV, was responsive to DNA damage and involved in cell growth regulation. These findings could be of great value in genotoxicity predictions and contribute to a deeper understanding of GLN biological functions.

Identification and characterisation of the angiotensin converting enzyme-3 (ACE3 gene: a novel mammalian homologue of ACE

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Phelan Anne

2007-06-01

Full Text Available Abstract Background Mammalian angiotensin converting enzyme (ACE plays a key role in blood pressure regulation. Although multiple ACE-like proteins exist in non-mammalian organisms, to date only one other ACE homologue, ACE2, has been identified in mammals. Results Here we report the identification and characterisation of the gene encoding a third homologue of ACE, termed ACE3, in several mammalian genomes. The ACE3 gene is located on the same chromosome downstream of the ACE gene. Multiple sequence alignment and molecular modelling have been employed to characterise the predicted ACE3 protein. In mouse, rat, cow and dog, the predicted protein has mutations in some of the critical residues involved in catalysis, including the catalytic Glu in the HEXXH zinc binding motif which is Gln, and ESTs or reverse-transcription PCR indicate that the gene is expressed. In humans, the predicted ACE3 protein has an intact HEXXH motif, but there are other deletions and insertions in the gene and no ESTs have been identified. Conclusion In the genomes of several mammalian species there is a gene that encodes a novel, single domain ACE-like protein, ACE3. In mouse, rat, cow and dog ACE3, the catalytic Glu is replaced by Gln in the putative zinc binding motif, indicating that in these species ACE3 would lack catalytic activity as a zinc metalloprotease. In humans, no evidence was found that the ACE3 gene is expressed and the presence of deletions and insertions in the sequence indicate that ACE3 is a pseudogene.

Polymorphism of XRCC1, XRCC3, and XPD Genes and Risk of Chronic Myeloid Leukemia

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Claudia Bănescu

2014-01-01

Full Text Available The genetic polymorphisms of X-ray repair cross complementing group 1 (XRCC1, X-ray repair cross complementing group 3 (XRCC3, and xeroderma pigmentosum complementation group D (XPD repair genes may lead to genetic instability and leukemogenesis. The purpose of the study was to evaluate the association between XRCC1 Arg399Gln, Arg280His and Arg194Trp, XRCC3 Thr241Met, and XPD Lys751Gln polymorphisms and the risk of developing CML in Romanian patients. A total of 156 patients diagnosed with CML and 180 healthy controls were included in this study. We found no association between CML and XRCC1 or XRCC3 variant genotypes in any of the investigated cases. A significant difference was observed in the variant genotype frequencies of the XPD Lys751Gln polymorphism between the patients with CML and control group (for variant homozygous genotypes, OR=2.37; 95% CI=1.20–4.67; P value = 0.016 and for combined heterozygous and variant homozygous genotypes, OR=1.72; 95% CI=1.10–2.69; P value = 0.019. This was also observed when analyzing the variant 751Gln allele (OR=1.54; 95% CI=1.13–2.11; P value = 0.008. Our results suggest that the XPD Lys751Gln variant genotype increases the risk of CML.

Removal of the effect of ammonium on the regulation of nitrogenase enzyme in Rhodobacter capsulatus DSM1710 for improved hydrogen production

Energy Technology Data Exchange (ETDEWEB)

Pekgoez, Guelsah; Guenduez, Ufuk [Middle East Technical Univ. (Turkey). Dept. of Biology; Eroglu, Inci [Middle East Technical Univ. (Turkey). Dept. of Chemical Engineering; Rakhely, Gabor [Szeged Univ. (Hungary). Dept. of Biotechnology

2010-07-01

Photofermentative biohydrogen production by purple non-sulfur (PNS) bacteria is a renewable and clean way of producing hydrogen. Hydrogen production by PNS bacteria, Rhodobacter capsulatus, is mediated mainly by nitrogenases, which primarily fix molecular nitrogen to ammonium and produce hydrogen as byproduct. The reaction catalyzed by nitrogenases requires a lot of energy. Hence, there is a complex regulation on nitrogenase enzyme complex, consequently, on hydrogen production. Whenever ammonium, which is the end product of nitrogen fixation reaction, is found in the environment, hydrogen production stops. GlnB and GlnK proteins are the critical regulatory proteins in ammonium dependent regulation of the nitrogenase gene expression. In this study, the aim is to release the ammonium regulation on nitrogenase enzyme by inactivating glnB and glnK genes. For this purpose, relevant recombinant vectors were constructed; R.capsulatus glnB- strain was obtained. The double R.capsulatus glnB{sup -}glnK{sup -} strain, able to produce hydrogen independent of ammonium concentration of the environment is to be obtained. (orig.)

Role of the APOB Gene Polymorphism (c.12669G>A, p. Gln4154Lys) in Coronary Artery Disease in the Indian Punjabi Population.

Science.gov (United States)

Sharma, R; Mahajan, M; Singh, B; Singh, G; Singh, P

2011-12-01

High concentration of apolipoprotein B (apoB) is a risk factor for coronary artery disease (CAD). The association of the APOB gene polymorphism c.12669G>A, p.Gln4154Lys with the risk of CAD varies considerably in different populations. The present study represents the first investigation regarding the role of this APOB gene polymorphism with CAD in the Indian Punjabi population. We have studied the APOB gene polymorphism c.12669G>A, p.Gln4154Lys and its relationship with lipid, apoB, low-density lipoprotein (LDL) heterogeneity and oxidation in subjects suffering from CAD. The study was conducted on 87 patients with CAD; 75 healthy subjects served as controls. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to determine the DNA polymorphism in the APOB gene. Frequency of R- (mutant) allele was significantly high (p 0.05). However, serum apoB levels were significantly raised (p Punjabi population. Overall, it may be concluded that the R- allele might be associated with increased susceptibility towards CAD development in the Indian Punjabi population, and one of the linking factor is the elevation in serum apoB levels. However, this association needs further evaluation in a larger population. Secondly, the robust mechanism behind the positive association of the R- allele with raised serum apoB levels needs to be explored, which might be helpful in the strengthening the observed results.

Lack of association between the XPD Lys751Gln polymorphism and colorectal cancer risk: a meta-analysis

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Zhang T

2014-07-01

Full Text Available Tao Zhang,1,* Dong-ming Zhang,2,* Da Zhao,1 Xiao-ming Hou,1 Shou-cheng Ma,1 Xiao-jun Liu1 1Department of Oncology, The First Hospital of Lanzhou University, The Branch Hospital of Donggang, Lanzhou, 2Department of Oncology, The Second People's Hospital of Pingliang, Pingliang, People's Republic of China  *These authors contributed equally to this work Background: The xeroderma pigmentosum complementary group D (XPD gene has been linked to the development of colorectal cancer (CRC through disruption of DNA repair. Several studies have suggested that the XPD polymorphism Lys751Gln is associated with an increased risk of developing CRC. However, previous results remain inconclusive. Herein, we performed a meta-analysis to evaluate the potential for this relationship. Methods: Relevant studies were retrieved from the PubMed database. Strict selection and exclusion criteria were determined, and the odds ratio with a 95% confidence interval was used to assess the strength of associations. The fixed or random effects model was selected on the basis of heterogeneity tests among studies. Publication bias was estimated using funnel plots and Egger's regression test. Results: The meta-analysis included 2,961 cases and 4,539 controls from eleven studies. The results indicated that the XPD Lys751Gln polymorphism had no association with CRC risk for all genetic models (Gln-Gln versus Lys-Lys, P=0.477; Lys-Gln versus Lys-Lys, P=0.283; Lys-Gln + Gln-Gln versus Lys-Lys, P=0.562, even when compared within subgroups based on ethnicity and source of controls. Conclusion: Based on the results of our meta-analysis, there is no evidence of a link between the XPD Lys751Gln polymorphism and risk of CRC. Keywords: XPD Lys751Gln polymorphism, colorectal cancer risk, meta-analysis

Plasmodium Apicoplast Gln-tRNA Gln Biosynthesis Utilizes a Unique GatAB Amidotransferase Essential for Erythrocytic Stage Parasites

KAUST Repository

Mailu, Boniface M.; Li, Ling; Arthur, Jen; Nelson, Todd M.; Ramasamy, Gowthaman; Fritz-Wolf, Karin; Becker, Katja; Gardner, Malcolm J.

2015-01-01

© 2015 by The American Society for Biochemistry and Molecular Biology, Inc. The malaria parasite Plasmodium falciparum apicoplast indirect aminoacylation pathway utilizes a non-discriminating glutamyl-tRNA synthetase to synthesize Glu-tRNAGln and a glutaminyl-tRNA amidotransferase to convert Glu-tRNAGln to Gln-tRNAGln. Here, we show that Plasmodium falciparum and other apicomplexans possess a unique heterodimeric glutamyltRNA amidotransferase consisting of GatA and GatB subunits (GatAB). We localized the P. falciparum GatA and GatB subunits to the apicoplast in blood stage parasites and demonstrated that recombinant GatAB converts Glu-tRNAGln to Gln-tRNAGln in vitro. We demonstrate that the apicoplast GatAB-catalyzed reaction is essential to the parasite blood stages because we could not delete the Plasmodium berghei gene encoding GatA in blood stage parasites in vivo. A phylogenetic analysis placed the split between Plasmodium GatB, archaeal GatE, and bacterial GatB prior to the phylogenetic divide between bacteria and archaea. Moreover, Plasmodium GatA also appears to have emerged prior to the bacterial-archaeal phylogenetic divide. Thus, although GatAB is found in Plasmodium, it emerged prior to the phylogenetic separation of archaea and bacteria.

Plasmodium Apicoplast Gln-tRNA Gln Biosynthesis Utilizes a Unique GatAB Amidotransferase Essential for Erythrocytic Stage Parasites

KAUST Repository

Mailu, Boniface M.

2015-08-28

© 2015 by The American Society for Biochemistry and Molecular Biology, Inc. The malaria parasite Plasmodium falciparum apicoplast indirect aminoacylation pathway utilizes a non-discriminating glutamyl-tRNA synthetase to synthesize Glu-tRNAGln and a glutaminyl-tRNA amidotransferase to convert Glu-tRNAGln to Gln-tRNAGln. Here, we show that Plasmodium falciparum and other apicomplexans possess a unique heterodimeric glutamyltRNA amidotransferase consisting of GatA and GatB subunits (GatAB). We localized the P. falciparum GatA and GatB subunits to the apicoplast in blood stage parasites and demonstrated that recombinant GatAB converts Glu-tRNAGln to Gln-tRNAGln in vitro. We demonstrate that the apicoplast GatAB-catalyzed reaction is essential to the parasite blood stages because we could not delete the Plasmodium berghei gene encoding GatA in blood stage parasites in vivo. A phylogenetic analysis placed the split between Plasmodium GatB, archaeal GatE, and bacterial GatB prior to the phylogenetic divide between bacteria and archaea. Moreover, Plasmodium GatA also appears to have emerged prior to the bacterial-archaeal phylogenetic divide. Thus, although GatAB is found in Plasmodium, it emerged prior to the phylogenetic separation of archaea and bacteria.

XRCC1 codon 399Gln polymorphism is associated with radiotherapy-induced acute dermatitis and mucositis in nasopharyngeal carcinoma patients

International Nuclear Information System (INIS)

Li, Haijun; You, Yanjie; Lin, Canfeng; Zheng, Mingzhang; Hong, Chaoqun; Chen, Jiongyu; Li, Derui; Au, William W; Chen, Zhijian

2013-01-01

To evaluate the association between single nucleotide polymorphisms (SNPs) at the 194 and 399 codons of XRCC1, and the risk of severe acute skin and oral mucosa reactions in nasopharyngeal carcinoma patients in China. 114 patients with nasopharyngeal carcinoma were sequentially recruited in this study. Heparinized peripheral blood samples were taken for SNPs analysis before the start of radiation treatment. SNPs in XRCC1 (194Arg/Trp and 399Arg/Gln) gene were analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Dermatitis at upper neck and oral mucositis were clinically recorded according to the Common Terminology Criteria for Adverse Events v.3.0. The variant allele frequencies were 0.289 for XRCC1 194Trp and 0.263 for XRCC1 399Gln. Of the 114 patients, 24 experienced grade 3 acute dermatitis and 48 had grade 3 acute mucositis. The XRCC1 399Arg/Gln was significantly associated with the development of grade 3 dermatitis (Odds Ratio, 2.65; 95% CI, 1.04–6.73; p = 0.037, χ2 = 4.357). In addition, it was also associated with higher incidence of grade 3 mucositis with a borderline statistical significance (Odds Ratio, 2.11; 95% CI, 0.951–4.66; p = 0.065, χ2 = 3.411). The relationship between XRCC1 194Arg/Trp and acute dermatitis, and mucositis was not found. Our investigation shows, for the first time, that patients with the XRCC1 399Arg/Gln genotype were more likely to experience severe acute dermatitis and oral mucositis. With further validation, the information can be used to determine personalized radiotherapy strategy

Modulation of Colorectal Cancer Risk by Polymorphisms in 51Gln/His, 64Ile/Val, and 148Asp/Glu of APEX Gene; 23Gly/Ala of XPA Gene; and 689Ser/Arg of ERCC4 Gene

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L. Dziki

2017-01-01

Full Text Available Polymorphisms in DNA repair genes may affect the activity of the BER (base excision repair and NER (nucleotide excision repair systems. Using DNA isolated from blood taken from patients (n=312 and a control group (n=320 with CRC, we have analyzed the polymorphisms of selected DNA repair genes and we have demonstrated that genotypes 51Gln/His and 148Asp/Glu of APEX gene and 23Gly/Ala of XPA gene may increase the risk of colorectal cancer. At the same time analyzing the gene-gene interactions, we suggest the thesis that the main factor to be considered when analyzing the impact of polymorphisms on the risk of malignant transformation should be intergenic interactions. Moreover, we are suggesting that some polymorphisms may have impact not only on the malignant transformation but also on the stage of the tumor.

Genetic polymorphisms of xeroderma pigmentosum group D gene Asp312Asn and Lys751Gln and susceptibility to prostate cancer: a systematic review and meta-analysis.

Science.gov (United States)

Ma, Qingtong; Qi, Can; Tie, Chong; Guo, Zhanjun

2013-11-10

Many studies have reported the role of xeroderma pigmentosum group D (XPD) with prostate cancer risk, but the results remained controversial. To derive a more precise estimation of the relationship, a meta-analysis was performed. Odds ratios (ORs) with 95% confidence intervals (CIs) were estimated to assess the association between XPD Asp312Asn and Lys751Gln polymorphisms and prostate cancer risk. A total of 8 studies including 2620 cases and 3225 controls described Asp312Asn genotypes, among which 10 articles involving 3230 cases and 3582 controls described Lys751Gln genotypes and were also involved in this meta-analysis. When all the eligible studies were pooled into this meta-analysis, a significant association between prostate cancer risk and XPD Asp312Asn polymorphism was found. For Asp312Asn polymorphism, in the stratified analysis by ethnicity and source of controls, prostate cancer risk was observed in co-dominant, dominant and recessive models, while no evidence of any associations of XPD Lys751Gln polymorphism with prostate cancer was found in the overall or subgroup analyses. Our meta-analysis supports that the XPD Asp312Asn polymorphism contributed to the risk of prostate cancer from currently available evidence. However, a study with a larger sample size is needed to further evaluate gene-environment interaction on XPD Asp312Asn and Lys751Gln polymorphisms and prostate cancer risk. © 2013.

Modular representations of GL(n) distinguished by GL(n-1) over a p-adic field

OpenAIRE

Sécherre , Vincent; Venketasubramanian , C. G.

2015-01-01

Let $\\F$ be a non-Archimedean locally compact field, $q$ be the cardinality of its residue field, and $\\R$ be an algebraically closed field of characteristic $\\ell$ not dividing $q$.We classify all irredu\\-cible smooth $\\R$-representations of $\\GL\\_n(\\F)$ having a nonzero $\\GL\\_{n-1}(\\F)$-inva\\-riant linear form, when $q$ is not congruent to $1$ mod $\\ell$.Partial results in the case when $q$ is $1$ mod $\\ell$ show that, unlike the complex case, the space of $\\GL\\_{n-1}(\\F)$-invariant linear ...

DNA Repair Gene Polymorphism and the Risk of Mitral Chordae Tendineae Rupture

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Aysel Kalayci Yigin

2015-01-01

Full Text Available Polymorphisms in Lys939Gln XPC gene may diminish DNA repair capacity, eventually increasing the risk of carcinogenesis. The aim of the present study was to evaluate the significance of polymorphism Lys939Gln in XPC gene in patients with mitral chordae tendinea rupture (MCTR. Twenty-one patients with MCTR and thirty-seven age and sex matched controls were enrolled in the study. Genotyping of XPC gene Lys939Gln polymorphism was carried out using polymerase chain reaction- (PCR- restriction fragment length polymorphism (RFLP. The frequencies of the heterozygote genotype (Lys/Gln-AC and homozygote genotype (Gln/Gln-CC were significantly different in MCTR as compared to control group, respectively (52.4% versus 43.2%, p=0.049; 38.15% versus 16.2%, p=0.018. Homozygote variant (Gln/Gln genotype was significantly associated with increased risk of MCTR (OR = 2.059; 95% CI: 1.097–3.863; p=0.018. Heterozygote variant (Lys/Gln genotype was also highly significantly associated with increased risk of MCTR (OR = 1.489; 95% CI: 1.041–2.129; p=0.049. The variant allele C was found to be significantly associated with MCTR (OR = 1.481; 95% CI: 1.101–1.992; p=0.011. This study has demonstrated the association of XPC gene Lys939Gln polymorphism with MCTR, which is significantly associated with increased risk of MCTR.

Comparative genome analysis of central nitrogen metabolism and its control by GlnR in the class Bacilli

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Kormelink Tom

2012-05-01

Full Text Available Abstract Background The assimilation of nitrogen in bacteria is achieved through only a few metabolic conversions between alpha-ketoglutarate, glutamate and glutamine. The enzymes that catalyze these conversions are glutamine synthetase, glutaminase, glutamate dehydrogenase and glutamine alpha-ketoglutarate aminotransferase. In low-GC Gram-positive bacteria the transcriptional control over the levels of the related enzymes is mediated by four regulators: GlnR, TnrA, GltC and CodY. We have analyzed the genomes of all species belonging to the taxonomic families Bacillaceae, Listeriaceae, Staphylococcaceae, Lactobacillaceae, Leuconostocaceae and Streptococcaceae to determine the diversity in central nitrogen metabolism and reconstructed the regulation by GlnR. Results Although we observed a substantial difference in the extent of central nitrogen metabolism in the various species, the basic GlnR regulon was remarkably constant and appeared not affected by the presence or absence of the other three main regulators. We found a conserved regulatory association of GlnR with glutamine synthetase (glnRA operon, and the transport of ammonium (amtB-glnK and glutamine/glutamate (i.e. via glnQHMP, glnPHQ, gltT, alsT. In addition less-conserved associations were found with, for instance, glutamate dehydrogenase in Streptococcaceae, purine catabolism and the reduction of nitrite in Bacillaceae, and aspartate/asparagine deamination in Lactobacillaceae. Conclusions Our analyses imply GlnR-mediated regulation in constraining the import of ammonia/amino-containing compounds and the production of intracellular ammonia under conditions of high nitrogen availability. Such a role fits with the intrinsic need for tight control of ammonia levels to limit futile cycling.

Leptin receptor Gln223Arg polymorphism and breast cancer risk in Nigerian women: A case control study

International Nuclear Information System (INIS)

Okobia, Michael N; Taioli, Emanuela; Bunker, Clareann H; Garte, Seymour J; Zmuda, Joseph M; Ezeome, Emmanuel R; Anyanwu, Stanley N; Uche, Emmanuel E; Kuller, Lewis H; Ferrell, Robert E

2008-01-01

Leptin, a 16 kDa polypeptide hormone, implicated in various physiological processes, exerts its action through the leptin receptor, a member of the class I cytokine receptor family. Both leptin and leptin receptor have recently been implicated in processes leading to breast cancer initiation and progression in animal models and humans. An A to G transition mutation in codon 223 in exon 6 of the leptin receptor gene, resulting in glutamine to arginine substitution (Gln223Arg), lies within the first of two putative leptin-binding regions and may be associated with impaired signaling capacity of the leptin receptor. This study was designed to assess the role of this polymorphism in breast cancer susceptibility in Nigerian women. We utilized a polymerase chain reaction (PCR)-based restriction fragment length polymorphism (RFLP) assay to evaluate the association between the Gln223Arg polymorphism of the leptin receptor gene and breast risk in Nigeria in a case control study involving 209 women with breast cancer and 209 controls without the disease. Study participants were recruited from surgical outpatient clinics and surgical wards of four University Teaching Hospitals located in Midwestern and southeastern Nigeria between September 2002 and April 2004. Premenopausal women carrying at least one LEPR 223Arg allele were at a modestly increased risk of breast cancer after adjusting for confounders (OR = 1.8, 95% confidence interval [CI] 1.0–3.2, p = 0.07). There was no association with postmenopausal breast cancer risk (OR = 0.9, 95% CI 0.4–1.8, p = 0.68). Our results suggest that the LEPR Gln223Arg polymorphism in the extracellular domain of the LEPR receptor gene is associated with a modestly increased risk of premenopausal breast cancer in Nigerian women

Deletion of Type I glutamine synthetase deregulates nitrogen metabolism and increases ethanol production in Clostridium thermocellum

Energy Technology Data Exchange (ETDEWEB)

Rydzak, Thomas [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Garcia, David [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Stevenson, David M. [Univ. of Wisconsin, Madison, WI (United States). Dept. of Bacteriology; Sladek, Margaret [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Klingeman, Dawn M. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Holwerda, Evert K. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division; Dartmouth College, Hanover, NH (United States). Thayer School of Engineering; Amador-Noguez, Daniel [Univ. of Wisconsin, Madison, WI (United States). Dept. of Bacteriology; Brown, Steven D. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Guss, Adam M. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center

2017-05-01

Clostridium thermocellum rapidly deconstructs cellulose and ferments resulting hydrolysis products into ethanol and other products, and is thus a promising platform organism for the development of cellulosic biofuel production via consolidated bioprocessing. And while recent metabolic engineering strategies have targeted eliminating canonical fermentation products (acetate, lactate, formate, and H₂), C. thermocellum also secretes amino acids, which has limited ethanol yields in engineered strains to approximately 70% of the theoretical maximum. To decrease amino acid secretion, we attempted to reduce ammonium assimilation by deleting the Type I glutamine synthetase (glnA) in C. thermocellum. Deletion of glnA reduced levels of secreted valine and total amino acids by 53% and 44% respectively, and increased ethanol yields by 53%. RNA-seq analysis revealed that genes encoding the RNF-complex were more highly expressed in ΔglnA and may have a role in improving NADH-availability for ethanol production. While a significant up-regulation of genes involved in nitrogen assimilation and urea uptake suggested that deletion of glnA induces a nitrogen starvation response, metabolomic analysis showed an increase in intracellular glutamine and α-ketoglutarate levels indicative of nitrogen-rich conditions. Here, we propose that deletion of glnA causes deregulation of nitrogen metabolism, leading to overexpression of nitrogen metabolism genes and, in turn, elevated glutamine/α-ketoglutarate levels. Here we demonstrate that perturbation of nitrogen assimilation is a promising strategy to redirect flux from the production of nitrogenous compounds toward biofuels in C. thermocellum.

Polymorphisms of the XRCC1, XRCC3, & XPD genes, and colorectal cancer risk: a case-control study in Taiwan

International Nuclear Information System (INIS)

Yeh, Chih-Ching; Sung, Fung-Chang; Tang, Reiping; Chang-Chieh, Chung Rong; Hsieh, Ling-Ling

2005-01-01

Recent studies relating to the association between DNA repair-gene polymorphisms and colorectal cancer risk would, to the best of our knowledge, appear to be very limited. This study was designed to examine the polymorphisms associated with three DNA repair genes, namely: XRCC1 Arg399Gln, XRCC3 Thr241Met and XPD Lys751Gln, and investigate their role as susceptibility markers for colorectal cancer. We conducted a case-control study including 727 cases of cancer and 736 hospital-based age- and sex-matched healthy controls to examine the role of genetic polymorphisms of three DNA-repair genes (XRCC1, XRCC3 and XPD) in the context of colorectal cancer risk for the Taiwanese population. Genomic DNA isolated from 10 ml whole blood was used to genotype XRCC1 Arg399Gln, XRCC3 Thr241Met and XPD Lys751Gln by means of polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis. The risk for colorectal cancer did not appear to differ significantly amongst individuals featuring the XRCC1 399Arg/Arg genotype (OR = 1.18; 95% CI, 0.96–1.45), the XRCC3 241Thr/Thr genotype (OR = 1.25; 95% CI, 0.88–1.79) or the XPD 751Gln allele (OR = 1.20; 95% CI, 0.90–1.61), although individuals featuring a greater number of risk genotypes (genotype with OR greater than 1) did experience a higher risk for colorectal cancer when compared to those who didn't feature any risk genotypes (Trend test P = 0.03). Compared with those individuals who didn't express any putative risk genotypes, individuals featuring all of the putative risk genotypes did experience a significantly greater cancer risk (OR = 2.43, 95% CI = 1.21–4.90), particularly for individuals suffering tumors located in the rectum (OR = 3.18, 95% CI = 1.29–7.82) and diagnosed prior to the age of 60 years (OR = 4.90, 95% CI = 1.72–14.0). Our results suggest that DNA-repair pathways may simultaneously modulate the risk of colorectal cancer for the Taiwanese population, and, particularly

Arg16Gly and Gln27Glu β2 adrenergic polymorphisms influence cardiac autonomic modulation and baroreflex sensitivity in healthy young Brazilians

Science.gov (United States)

Atala, Magda M; Goulart, Alessandra; Guerra, Grazia M; Mostarda, Cristiano; Rodrigues, Bruno; Mello, Priscila R; Casarine, Dulce E; Irigoyen, Maria-Claudia; Pereira, Alexandre C; Consolim-Colombo, Fernanda M

2015-01-01

The association between functional β2 adrenergic receptor (β2-AR) polymorphisms and cardiac autonomic modulation is still unclear. Thus, two common polymorphisms in the β2-AR gene (Gln27Glu β2 and Arg16Gly β2) were studied to determine whether they might affect tonic and reflex cardiac sympathetic activity in healthy young subjects. A total of 213 healthy young white subjects of both genders (53% female), aged 18-30 years (23.5±3.4 y), had their continuous blood pressure curves noninvasively recorded by Finometer at baseline, and other hemodynamic parameters, as cardiac autonomic modulation, baroreflex sensitivity, and allele, genotype, and diplotype frequencies calculated. Associations were made between Arg16Gly β2 and Gln27Glu β2 polymorphisms and between β2-AR diplotypes and all variables. The heart rate was significantly lower (P<0.001) in the presence of homozygous Arg/Arg alleles (60.9±1.5 bpm) than in that of Arg/Gly heterozygotes (65.9±1.0 bpm) or Gly/Gly homozygotes (66.3±1.2 bpm). Homozygous carriers of Arg16 allele had an alpha index (19.2±1.3) significantly higher (P<0.001) than that of the subjects with the Gly allele Gly/Gly (14.5±0.7) or Arg/Gly (14.6±0.7). Furthermore, the recessive Glu27Glu and the heterozygous Gln27Glu genotypes had a higher percentage of low-frequency components (LF%) than the homozygous Gln27Gln (15.1% vs. 16.0% vs. 8.2%, P=0.03, respectively). In healthy young subjects, the presence of β2-AR Arg16 allele in a recessive model was associated with higher baroreflex sensitivity, and increased parasympathetic modulation in studied individuals. PMID:25755837

Deletion of Type I glutamine synthetase deregulates nitrogen metabolism and increases ethanol production in Clostridium thermocellum.

Science.gov (United States)

Rydzak, Thomas; Garcia, David; Stevenson, David M; Sladek, Margaret; Klingeman, Dawn M; Holwerda, Evert K; Amador-Noguez, Daniel; Brown, Steven D; Guss, Adam M

2017-05-01

Clostridium thermocellum rapidly deconstructs cellulose and ferments resulting hydrolysis products into ethanol and other products, and is thus a promising platform organism for the development of cellulosic biofuel production via consolidated bioprocessing. While recent metabolic engineering strategies have targeted eliminating canonical fermentation products (acetate, lactate, formate, and H 2 ), C. thermocellum also secretes amino acids, which has limited ethanol yields in engineered strains to approximately 70% of the theoretical maximum. To investigate approaches to decrease amino acid secretion, we attempted to reduce ammonium assimilation by deleting the Type I glutamine synthetase (glnA) in an essentially wild type strain of C. thermocellum. Deletion of glnA reduced levels of secreted valine and total amino acids by 53% and 44% respectively, and increased ethanol yields by 53%. RNA-seq analysis revealed that genes encoding the RNF-complex were more highly expressed in ΔglnA and may have a role in improving NADH-availability for ethanol production. While a significant up-regulation of genes involved in nitrogen assimilation and urea uptake suggested that deletion of glnA induces a nitrogen starvation response, metabolomic analysis showed an increase in intracellular glutamine levels indicative of nitrogen-rich conditions. We propose that deletion of glnA causes deregulation of nitrogen metabolism, leading to overexpression of nitrogen metabolism genes and, in turn, elevated glutamine levels. Here we demonstrate that perturbation of nitrogen assimilation is a promising strategy to redirect flux from the production of nitrogenous compounds toward biofuels in C. thermocellum. Copyright © 2017. Published by Elsevier Inc.

Herbaspirillum seropedicae signal transduction protein PII is structurally similar to the enteric GlnK.

Science.gov (United States)

Machado Benelli, Elaine; Buck, Martin; Polikarpov, Igor; Maltempi de Souza, Emanuel; Cruz, Leonardo M; Pedrosa, Fábio O

2002-07-01

PII-like proteins are signal transduction proteins found in bacteria, archaea and eukaryotes. They mediate a variety of cellular responses. A second PII-like protein, called GlnK, has been found in several organisms. In the diazotroph Herbaspirillum seropedicae, PII protein is involved in sensing nitrogen levels and controlling nitrogen fixation genes. In this work, the crystal structure of the unliganded H. seropedicae PII was solved by X-ray diffraction. H. seropedicae PII has a Gly residue, Gly108 preceding Pro109 and the main-chain forms a beta turn. The glycine at position 108 allows a bend in the C-terminal main-chain, thereby modifying the surface of the cleft between monomers and potentially changing function. The structure suggests that the C-terminal region of PII proteins may be involved in specificity of function, and nonenteric diazotrophs are found to have the C-terminal consensus XGXDAX(107-112). We are also proposing binding sites for ATP and 2-oxoglutarate based on the structural alignment of PII with PII-ATP/GlnK-ATP, 5-carboxymethyl-2-hydroxymuconate isomerase and 4-oxalocrotonate tautomerase bound to the inhibitor 2-oxo-3-pentynoate.

Effect of T- and C-loop mutations on the Herbaspirillum seropedicae GlnB protein in nitrogen signalling.

Science.gov (United States)

Bonatto, Ana C; Souza, Emanuel M; Pedrosa, Fábio O; Yates, M Geoffrey; Benelli, Elaine M

2005-01-01

Proteins of the PII family are found in species of all kingdoms. Although these proteins usually share high identity, their functions are specific to the different organisms. Comparison of structural data from Escherichia coli GlnB and GlnK and Herbaspirillum seropedicae GlnB showed that the T-loop and C-terminus were variable regions. To evaluate the role of these regions in signal transduction by the H. seropedicae GlnB protein, four mutants were constructed: Y51F, G108A/P109a, G108W and Q3R/T5A. The activities of the native and mutated proteins were assayed in an E. coli background constitutively expressing the Klebsiella pneumoniae nifLA operon. The results suggested that the T-loop and C-terminus regions of H. seropedicae GlnB are involved in nitrogen signal transduction.

Conformational Dynamics of apo-GlnBP Revealed by Experimental and Computational Analysis

KAUST Repository

Feng, Yitao

2016-10-13

The glutamine binding protein (GlnBP) binds l-glutamine and cooperates with its cognate transporters during glutamine uptake. Crystal structure analysis has revealed an open and a closed conformation for apo- and holo-GlnBP, respectively. However, the detailed conformational dynamics have remained unclear. Herein, we combined NMR spectroscopy, MD simulations, and single-molecule FRET techniques to decipher the conformational dynamics of apo-GlnBP. The NMR residual dipolar couplings of apo-GlnBP were in good agreement with a MD-derived structure ensemble consisting of four metastable states. The open and closed conformations are the two major states. This four-state model was further validated by smFRET experiments and suggests the conformational selection mechanism in ligand recognition of GlnBP. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Conformational Dynamics of apo-GlnBP Revealed by Experimental and Computational Analysis

KAUST Repository

Feng, Yitao; Zhang, Lu; Wu, Shaowen; Liu, Zhijun; Gao, Xin; Zhang, Xu; Liu, Maili; Liu, Jianwei; Huang, Xuhui; Wang, Wenning

2016-01-01

The glutamine binding protein (GlnBP) binds l-glutamine and cooperates with its cognate transporters during glutamine uptake. Crystal structure analysis has revealed an open and a closed conformation for apo- and holo-GlnBP, respectively. However, the detailed conformational dynamics have remained unclear. Herein, we combined NMR spectroscopy, MD simulations, and single-molecule FRET techniques to decipher the conformational dynamics of apo-GlnBP. The NMR residual dipolar couplings of apo-GlnBP were in good agreement with a MD-derived structure ensemble consisting of four metastable states. The open and closed conformations are the two major states. This four-state model was further validated by smFRET experiments and suggests the conformational selection mechanism in ligand recognition of GlnBP. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim

Glutamine Synthetases GLN1;2 and GLN2 in Relation to Arabidopsis Growth Response to Elevated Atmospheric Carbon Dioxide and Varying Nitrogen Forms

DEFF Research Database (Denmark)

Vurrakula, Swathi

content while diluting nitrogen concentrations. Such a reduction in nitrogen concentration will affect plant response to stress and seed/grain yield. Glutamine synthetase (GS) is the central nitrogen-assimilatory enzyme, performing primary and secondary nitrogen assimilation, in response to environmental....... Plants grown under elevated CO2 absorbed ammonia from the atmosphere, except with a high ammonium supply. GLN1;2 had a non-redundant role in determining vegetative growth and ammonium tolerance in response to elevated CO2. Under elevated CO2, GLN1;2 was compensable by GLN2 in assimilating nitrate...

Rhizobia with 16S rRNA and nifH similar to Mesorhizobium huakuii but Novel recA, glnII, nodA and nodC genes are symbionts of New Zealand Carmichaelinae.

Directory of Open Access Journals (Sweden)

Heng Wee Tan

Full Text Available New Zealand became geographically isolated about 80 million years ago and this separation gave rise to a unique native flora including four genera of legume, Carmichaelia, Clianthus and Montigena in the Carmichaelinae clade, tribe Galegeae, and Sophora, tribe Sophoreae, sub-family Papilionoideae. Ten bacterial strains isolated from NZ Carmichaelinae growing in natural ecosystems grouped close to the Mesorhizobium huakuii type strain in relation to their 16S rRNA and nifH gene sequences. However, the ten strains separated into four groups on the basis of their recA and glnII sequences: all groups were clearly distinct from all Mesorhizobium type strains. The ten strains separated into two groups on the basis of their nodA sequences but grouped closely together in relation to nodC sequences; all nodA and nodC sequences were novel. Seven strains selected and the M. huakuii type strain (isolated from Astragalus sinicus produced functional nodules on Carmichaelia spp., Clianthus puniceus and A. sinicus but did not nodulate two Sophora species. We conclude that rhizobia closely related to M. huakuii on the basis of 16S rRNA and nifH gene sequences, but with variable recA and glnII genes and novel nodA and nodC genes, are common symbionts of NZ Carmichaelinae.

Constitutive and nitrogen catabolite repression-sensitive production of Gat1 isoforms.

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Rai, Rajendra; Tate, Jennifer J; Georis, Isabelle; Dubois, Evelyne; Cooper, Terrance G

2014-01-31

Nitrogen catabolite repression (NCR)-sensitive transcription is activated by Gln3 and Gat1. In nitrogen excess, Gln3 and Gat1 are cytoplasmic, and transcription is minimal. In poor nitrogen, Gln3 and Gat1 become nuclear and activate transcription. A long standing paradox has surrounded Gat1 production. Gat1 was first reported as an NCR-regulated activity mediating NCR-sensitive transcription in gln3 deletion strains. Upon cloning, GAT1 transcription was, as predicted, NCR-sensitive and Gln3- and Gat1-activated. In contrast, Western blots of Gat1-Myc(13) exhibited two constitutively produced species. Investigating this paradox, we demonstrate that wild type Gat1 isoforms (IsoA and IsoB) are initiated at Gat1 methionines 40, 95, and/or 102, but not at methionine 1. Their low level production is the same in rich and poor nitrogen conditions. When the Myc(13) tag is placed after Gat1 Ser-233, four N-terminal Gat1 isoforms (IsoC-F) are also initiated at methionines 40, 95, and/or 102. However, their production is highly NCR-sensitive, being greater in proline than glutamine medium. Surprisingly, all Gat1 isoforms produced in sufficient quantities to be confidently analyzed (IsoA, IsoC, and IsoD) require Gln3 and UASGATA promoter elements, both requirements typical of NCR-sensitive transcription. These data demonstrate that regulated Gat1 production is more complex than previously recognized, with wild type versus truncated Gat1 proteins failing to be regulated in parallel. This is the first reported instance of Gln3 UASGATA-dependent protein production failing to derepress in nitrogen poor conditions. A Gat1-lacZ ORF swap experiment indicated sequence(s) responsible for the nonparallel production are downstream of Gat1 leucine 61.

The relationship between XRCC1 and XRCC3 gene polymorphisms and lung cancer risk in northeastern Chinese.

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Shujie Guo

Full Text Available BACKGROUND: The prevalence of lung cancer in China will be the world's highest if allowed to proceed uncurbed. To unravel its genetic underpinnings, we sought to investigate the association of three well-characterized nonsynonymous polymorphisms in XRCC1 (Arg194Trp and Arg399Gln and XRCC3 (Thr241Met genes with lung cancer risk in northeastern Chinese. METHODOLOGY/PRINCIPAL FINDINGS: This study was hospital-based in design, encompassing 684 patients with lung cancer and 604 cancer-free controls. Genotyping was performed using the PCR-LDR (ligase detection reactions method. Data were analyzed by R language and multifactor dimensionality reduction (MDR software. Single-locus analysis identified significance in genotype distributions of polymorphism Arg194Trp (P = 0.002 and Arg399Gln (P = 0.017, and in allele distributions of Thr241Met (P = 0.005. Carriers of 399Gln/Gln genotype conferred a 147% increased risk relative to the non-carriers (odds ratio (OR: 2.47; 95% confidence interval (95% CI: 1.48-4.13; P<0.001. For Thr241Met, significance persisted under allelic (OR = 1.63; 95% CI: 1.14-2.33; P = 0.005, additive (OR = 1.64; 95% CI: 1.16-2.32; P = 0.005 and dominant (OR = 1.67; 95% CI: 1.17-2.38; P = 0.004 models. However, common allele combinations were comparable in frequency between patients and controls. In interaction analysis, the overall best MDR model included Arg399Gln and Thr241Met polymorphisms, with a maximal testing accuracy of 63.18% and a maximal cross-validation consistency of 10 out of 10 (P = 0.0175. CONCLUSIONS: Our study significantly demonstrated an independent and synergistic contribution of XRCC1 Arg399Gln and XRCC3 Thr241Met polymorphisms to lung cancer susceptibility in northeastern Chinese.

Role of B13 Glu in insulin assembly. The hexamer structure of recombinant mutant (B13 Glu-->Gln) insulin.

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Bentley, G A; Brange, J; Derewenda, Z; Dodson, E J; Dodson, G G; Markussen, J; Wilkinson, A J; Wollmer, A; Xiao, B

1992-12-20

The assembly of the insulin hexamer brings the six B13 glutamate side-chains at the centre into close proximity. Their mutual repulsion is unfavourable and zinc co-ordination to B10 histidine is necessary to stabilize the well known zinc-containing hexamers. Since B13 is always a carboxylic acid in all known sequences of hexamer forming insulins, it is likely to be important in the hormone's biology. The mutation of B13 Glu-->Gln leads to a stable zinc-free hexamer with somewhat reduced potency. The structures of the zinc-free B13 Gln hexamer and the 2Zn B13 insulin hexamer have been determined by X-ray analysis and refined with 2.5 A and 2.0 A diffraction data, respectively. Comparisons show that in 2Zn B13 Gln insulin, the hexamer structure (T6) is very like that of the native hormone. On the other hand, the zinc-free hexamer assumes a quaternary structure (T3/R3) seen in the native 4Zn insulin hexamer, and normally associated only with high chloride ion concentrations in the medium. The crystal structures show the B13 Gln side-chains only contact water in contrast to the B13 glutamate in 2Zn insulin. The solvation of the B13 Gln may be associated with this residue favouring helix at B1 to B8. The low potency of the B13 Gln insulin also suggests the residue influences the hormone's conformation.

Structural and functional study of the GlnB22-insulin mutant responsible for maturity-onset diabetes of the young.

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Květoslava Křížková

Full Text Available The insulin gene mutation c.137G>A (R46Q, which changes an arginine at the B22 position of the mature hormone to glutamine, causes the monogenic diabetes variant maturity-onset diabetes of the young (MODY. In MODY patients, this mutation is heterozygous, and both mutant and wild-type (WT human insulin are produced simultaneously. However, the patients often depend on administration of exogenous insulin. In this study, we chemically synthesized the MODY mutant [GlnB22]-insulin and characterized its biological and structural properties. The chemical synthesis of this insulin analogue revealed that its folding ability is severely impaired. In vitro and in vivo tests showed that its binding affinity and biological activity are reduced (both approximately 20% that of human insulin. Comparison of the solution structure of [GlnB22]-insulin with the solution structure of native human insulin revealed that the most significant structural effect of the mutation is distortion of the B20-B23 β-turn, leading to liberation of the B chain C-terminus from the protein core. The distortion of the B20-B23 β-turn is caused by the extended conformational freedom of the GlnB22 side chain, which is no longer anchored in a hydrogen bonding network like the native ArgB22. The partially disordered [GlnB22]-insulin structure appears to be one reason for the reduced binding potency of this mutant and may also be responsible for its low folding efficiency in vivo. The altered orientation and flexibility of the B20-B23 β-turn may interfere with the formation of disulfide bonds in proinsulin bearing the R46Q (GlnB22 mutation. This may also have a negative effect on the WT proinsulin simultaneously biosynthesized in β-cells and therefore play a major role in the development of MODY in patients producing [GlnB22]-insulin.

Arg753gln and Arg677 Trp Polymorphisms of Toll-Like Receptor 2 In Acute Apical Abscess.

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Miri-Moghaddam, Ebrahim; Farhad Mollashahi, Narges; Naghibi, Nava; Garme, Yasaman; Bazi, Ali

2018-06-01

Genetic polymorphisms can alter immunity response against pathogens, which in turn influence individuals' susceptibility to certain infections. Our aim was to determine the association of Arg753Gln (rs5743708) and Arg677Trp (rs12191786) polymorphisms of toll like receptor-2 gene with the two clinical forms of apical periodontitis: acute apical abscess (AAA) and asymptomatic apical periodontitis (AAP). There were 50 patients with AAA as case group and 50 with AAP as control group. Genotyping was done using Tetra-ARMS (amplification refractory mutation system) PCR. Heterozygous genotype of Arg677Trp polymorphism was associated with risk of AAA (OR=1.9, 95% CI: 0.7-5.5, p = 0.05). Although statistically insignificant, Arg677Trp polymorphism promoted the risk of AAA in dominant model (OR=2.1, 95% CI: 0.7-5.9, p > 0.05). The frequency of mutant allele (T) of Arg677Trp polymorphism was higher in AAA (14%) than AAP (7%) subjects (OR=1.7, 95% CI: 0.6-4.7). For Arg753Gln polymorphism, wild homozygous (GG) represented the dominant genotype in both cases (96%) and controls (100%). Variant allele (A) of Arg753Gln polymorphism was identified in 2% of AAA, while no individual represented with this allele in AAP subjects. Individuals with Arg753Gln; Arg677Trp (GG; TC) combination showed an elevated risk of AAA (OR=1.6, 95% CI: 0.5- 4.2, p > 0.05). Arg677Trp polymorphism of TLR-2 rendered a higher risk for the development of abscesses in apical periodontitis. It is recommended to explore role of this polymorphism in other populations.

Partial characterization of nif genes from the bacterium Azospirillum amazonense

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D.P. Potrich

2001-09-01

Full Text Available Azospirillum amazonense revealed genomic organization patterns of the nitrogen fixation genes similar to those of the distantly related species A. brasilense. Our work suggests that A. brasilense nifHDK, nifENX, fixABC operons and nifA and glnB genes may be structurally homologous to the counterpart genes of A. amazonense. This is the first analysis revealing homology between A. brasilense nif genes and the A. amazonense genome. Sequence analysis of PCR amplification products revealed similarities between the amino acid sequences of the highly conserved nifD and glnB genes of A. amazonense and related genes of A. brasilense and other bacteria. However, the A. amazonense non-coding regions (the upstream activator sequence region and the region between the nifH and nifD genes differed from related regions of A. brasilense even in nitrogenase structural genes which are highly conserved among diazotrophic bacteria. The feasibility of the 16S ribosomal RNA gene-based PCR system for specific detection of A. amazonense was shown. Our results indicate that the PCR primers for 16S rDNA defined in this article are highly specific to A. amazonense and can distinguish this species from A. brasilense.

Human-specific SNP in obesity genes, adrenergic receptor beta2 (ADRB2, Beta3 (ADRB3, and PPAR γ2 (PPARG, during primate evolution.

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Akiko Takenaka

Full Text Available UNLABELLED: Adrenergic-receptor beta2 (ADRB2 and beta3 (ADRB3 are obesity genes that play a key role in the regulation of energy balance by increasing lipolysis and thermogenesis. The Glu27 allele in ADRB2 and the Arg64 allele in ADRB3 are associated with abdominal obesity and early onset of non-insulin-dependent diabetes mellitus (NIDDM in many ethnic groups. Peroxisome proliferator-activated receptor γ (PPARG is required for adipocyte differentiation. Pro12Ala mutation decreases PPARG activity and resistance to NIDDM. In humans, energy-expense alleles, Gln27 in ADRB2 and Trp64 in ADRB3, are at higher frequencies than Glu27 and Arg64, respectively, but Ala12 in PPARG is at lower frequency than Pro12. Adaptation of humans for lipolysis, thermogenesis, and reduction of fat accumulation could be considered by examining which alleles in these genes are dominant in non-human primates (NHP. All NHP (P. troglodytes, G. gorilla, P. pygmaeus, H. agilis and macaques had energy-thrifty alleles, Gly16 and Glu27 in ADRB2, and Arg64 in ADRB3, but did not have energy-expense alleles, Arg16, Gln27 and Trp64 alleles. In PPARG gene, all NHP had large adipocyte accumulating type, the Pro12 allele. CONCLUSIONS: These results indicate that a tendency to produce much more heat through the energy-expense alleles developed only in humans, who left tropical rainforests for savanna and developed new features in their heat-regulation systems, such as reduction of body hair and increased evaporation of water, and might have helped the protection of entrails from cold at night, especially in glacial periods.

Effect of ATP and 2-oxoglutarate on the in vitro interaction between the NifA GAF domain and the GlnB protein of Azospirillum brasilense

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Sotomaior, P.; Araújo, L.M.; Nishikawa, C.Y.; Huergo, L.F.; Monteiro, R.A.; Pedrosa, F.O.; Chubatsu, L.S.; Souza, E.M.

2012-01-01

Azospirillum brasilense is a diazotroph that associates with important agricultural crops and thus has potential to be a nitrogen biofertilizer. The A. brasilense transcription regulator NifA, which seems to be constitutively expressed, activates the transcription of nitrogen fixation genes. It has been suggested that the nitrogen status-signaling protein GlnB regulates NifA activity by direct interaction with the NifA N-terminal GAF domain, preventing the inhibitory effect of this domain under conditions of nitrogen fixation. In the present study, we show that an N-terminal truncated form of NifA no longer required GlnB for activity and lost regulation by ammonium. On the other hand, in trans co-expression of the N-terminal GAF domain inhibited the N-truncated protein in response to fixed nitrogen levels. We also used pull-down assays to show in vitro interaction between the purified N-terminal GAF domain of NifA and the GlnB protein. The results showed that A. brasilense GlnB interacts directly with the NifA N-terminal domain and this interaction is dependent on the presence of ATP and 2-oxoglutarate

Effect of ATP and 2-oxoglutarate on the in vitro interaction between the NifA GAF domain and the GlnB protein of Azospirillum brasilense.

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Sotomaior, P; Araújo, L M; Nishikawa, C Y; Huergo, L F; Monteiro, R A; Pedrosa, F O; Chubatsu, L S; Souza, E M

2012-12-01

Azospirillum brasilense is a diazotroph that associates with important agricultural crops and thus has potential to be a nitrogen biofertilizer. The A. brasilense transcription regulator NifA, which seems to be constitutively expressed, activates the transcription of nitrogen fixation genes. It has been suggested that the nitrogen status-signaling protein GlnB regulates NifA activity by direct interaction with the NifA N-terminal GAF domain, preventing the inhibitory effect of this domain under conditions of nitrogen fixation. In the present study, we show that an N-terminal truncated form of NifA no longer required GlnB for activity and lost regulation by ammonium. On the other hand, in trans co-expression of the N-terminal GAF domain inhibited the N-truncated protein in response to fixed nitrogen levels. We also used pull-down assays to show in vitro interaction between the purified N-terminal GAF domain of NifA and the GlnB protein. The results showed that A. brasilense GlnB interacts directly with the NifA N-terminal domain and this interaction is dependent on the presence of ATP and 2-oxoglutarate.

Effect of ATP and 2-oxoglutarate on the in vitro interaction between the NifA GAF domain and the GlnB protein of Azospirillum brasilense

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Sotomaior, P.; Araújo, L.M.; Nishikawa, C.Y.; Huergo, L.F.; Monteiro, R.A.; Pedrosa, F.O.; Chubatsu, L.S.; Souza, E.M. [Departamento de Bioquímica e Biologia Molecular, Universidade Federal do Paraná, Curitiba, PR (Brazil)

2012-09-21

Azospirillum brasilense is a diazotroph that associates with important agricultural crops and thus has potential to be a nitrogen biofertilizer. The A. brasilense transcription regulator NifA, which seems to be constitutively expressed, activates the transcription of nitrogen fixation genes. It has been suggested that the nitrogen status-signaling protein GlnB regulates NifA activity by direct interaction with the NifA N-terminal GAF domain, preventing the inhibitory effect of this domain under conditions of nitrogen fixation. In the present study, we show that an N-terminal truncated form of NifA no longer required GlnB for activity and lost regulation by ammonium. On the other hand, in trans co-expression of the N-terminal GAF domain inhibited the N-truncated protein in response to fixed nitrogen levels. We also used pull-down assays to show in vitro interaction between the purified N-terminal GAF domain of NifA and the GlnB protein. The results showed that A. brasilense GlnB interacts directly with the NifA N-terminal domain and this interaction is dependent on the presence of ATP and 2-oxoglutarate.

Towards understanding the nitrogen signal transduction for nif gene expression in Klebsiella pneumoniae.

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Glöer, Jens; Thummer, Robert; Ullrich, Heike; Schmitz, Ruth A

2008-12-01

In the diazotroph Klebsiella pneumoniae, the nitrogen sensory protein GlnK mediates the cellular nitrogen status towards the NifL/NifA system that regulates transcription of the nitrogen fixation genes in response to ammonium and molecular oxygen. To identify amino acids of GlnK essential for this signal transduction by protein-protein interaction, we performed random point mutagenesis by PCR amplification under conditions of reduced Taq polymerase fidelity. Three thousand two hundred mutated glnK genes were screened to identify those that would no longer complement a K. pneumoniaeDeltaglnK strain for growth under nitrogen fixing conditions. Twenty-four candidates resulting in a Nif(-) phenotype were identified, carrying 1-11 amino acid changes in GlnK. Based on these findings, as well as structural data, several single mutations were introduced into glnK by site-directed mutagenesis, and the Nif phenotype and the respective effects on NifA-mediated nif gene induction was monitored in K. pneumoniae using a chromosomal nifK'-'lacZ fusion. Single amino acid changes resulting in significant nif gene inhibition under nitrogen limiting conditions were located within the highly conserved T-loop (A43G, A49T and N54D), the body of the protein (G87V and K79E) and in the C-terminal region (I100M, R103S, E106Q and D108G). Complex formation analyses between GlnK (wild-type or derivatives) and NifL or NifA in response to 2-oxoglutarate indicated that: (a) besides the T-loop, the C-terminal region of GlnK is essential for the interaction with NifL and NifA and (b) GlnK binds both proteins in the absence of 2-oxoglutarate, whereas, in the presence of 2-oxoglutarate, NifA is released but NifL remains bound to GlnK.

The nif Gene Operon of the Methanogenic Archaeon Methanococcus maripaludis

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Kessler, Peter S.; Blank, Carrine; Leigh, John A.

1998-01-01

Nitrogen fixation occurs in two domains, Archaea and Bacteria. We have characterized a nif (nitrogen fixation) gene cluster in the methanogenic archaeon Methanococcus maripaludis. Sequence analysis revealed eight genes, six with sequence similarity to known nif genes and two with sequence similarity to glnB. The gene order, nifH, ORF105 (similar to glnB), ORF121 (similar to glnB), nifD, nifK, nifE, nifN, and nifX, was the same as that found in part in other diazotrophic methanogens and except for the presence of the glnB-like genes, also resembled the order found in many members of the Bacteria. Using transposon insertion mutagenesis, we determined that an 8-kb region required for nitrogen fixation corresponded to the nif gene cluster. Northern analysis revealed the presence of either a single 7.6-kb nif mRNA transcript or 10 smaller mRNA species containing portions of the large transcript. Polar effects of transposon insertions demonstrated that all of these mRNAs arose from a single promoter region, where transcription initiated 80 bp 5′ to nifH. Distinctive features of the nif gene cluster include the presence of the six primary nif genes in a single operon, the placement of the two glnB-like genes within the cluster, the apparent physical separation of the cluster from any other nif genes that might be in the genome, the fragmentation pattern of the mRNA, and the regulation of expression by a repression mechanism described previously. Our study and others with methanogenic archaea reporting multiple mRNAs arising from gene clusters with only a single putative promoter sequence suggest that mRNA processing following transcription may be a common occurrence in methanogens. PMID:9515920

Genetic investigation of ocular developmental genes in 52 patients with anophthalmia/microphthalmia.

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Vidya, Nair Gopinathan; Rajkumar, Sankaranarayanan; Vasavada, Abhay R

2018-06-01

Mutation in eye developmental genes has been reported to cause anophthalmia and microphthalmia. However, in India, especially in the Western Indian population, such reports are scarce. Hence, the present study aims to investigate mutations in 15 ocular developmental genes in patients with anophthalmia and microphthalmia in the western region of India. Genomic DNA was isolated from the blood of 52 individuals affected with microphthalmia and anophthalmia, and 50 healthy normal controls. Polymerase chain reaction (PCR) was carried out for 15 genes including BMP4, CRYBA4, FOXE3, GDF6, GJA3, GJA8, MITF, OTX2, PAX6, PITX3, RAX, SIX3, SIX6, SOX2, and VSX2 using gene-specific primers spanning the exon-intron boundaries and part of a promoter region. The amplified PCR products were purified and then subjected to Sanger's bi-directional sequencing. Nucleotide variations were examined using a basic local alignment search tool (BLAST). Bi-directional sequencing identified 8 novel and 14 known variations. Out of this, the variations GJA3-c.92T>A; p.Ile31Asn, SOX2-c.542C>A; p.Pro181Gln and SOX2-c.541_542delinsGA; p.Pro181Glu were found to be deleterious by in silico analysis. The GJA3-p.Ile31Asn mutation was identified in a patient with bilateral microphthalmia, microcornea, and membranous cataract. The SOX2-p.Pro181Gln and SOX2-p.Pro181Glu mutations were identified in patients with isolated bilateral microphthalmia and microphthalmia with microcornea, respectively. A novel nondeleterious missense variation was identified in the GJA8 gene in a patient with anophthalmia. These results support the crucial role of GJA3 and SOX2 in eye development and indicate a detailed functional study to understand the molecular mechanisms underlying the disease pathology.

A phase II study of HMB/Arg/Gln against oral mucositis induced by chemoradiotherapy for patients with head and neck cancer.

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Yokota, Tomoya; Hamauchi, Satoshi; Yoshida, Yukio; Yurikusa, Takashi; Suzuki, Miho; Yamashita, Aiko; Ogawa, Hirofumi; Onoe, Tsuyoshi; Mori, Keita; Onitsuka, Tetsuro

2018-04-07

This phase II trial assessed the clinical benefit of beta-hydroxy-beta-methylbutyrate, arginine, and glutamine (HMB/Arg/Gln) for preventing chemoradiotherapy (CRT)-induced oral mucositis (OM) in patients with head and neck cancer (HNC). Patients with HNC receiving definitive or postoperative cisplatin-based CRT were enrolled. HMB/Arg/Gln was administered orally or per percutaneous endoscopic gastrostomy from the first day of CRT up to its completion. All patients received opioid-based pain control and oral care programs that we previously reported. The primary endpoint was the incidence of grade ≥ 3 OM (functional/symptomatic) according to the Common Terminology Criteria of Adverse Events version 3.0. Quality of life (EORTC QLQ-C30/PROMS) at baseline and upon radiotherapy at a dosage of 50 Gy were assessed. Thirty-five patients with HNC were enrolled. Sixteen of them (45.7%) developed grade ≥ 3 OM (i.e., functional/symptomatic). The incidence of grade ≤ 1 OM (functional/symptomatic) was 51.5% at 2 weeks and 82.9% at 4 weeks after radiotherapy completion. Clinical examination revealed that 10 patients (28.6%) developed grade ≥ 3 OM. The incidence of grade ≤ 1 OM (clinical exam) was 80.0% at 2 weeks and 100% at 4 weeks after radiotherapy completion. Adverse events related to HMB/Arg/Gln were an increase in blood urea nitrogen and diarrhea, but were easily managed. The addition of HMB/Arg/Gln to opioid-based pain control and oral care programs was feasible but still insufficient at reducing the incidence of CRT-induced severe OM. However, the benefit of HMB/Arg/Gln should not be neglected given the findings of clinical examinations and the rapid recovery from severe OM. UMIN000016453.

Obesity-related gene ADRB2, ADRB3 and GHRL polymorphisms and the response to a weight loss diet intervention in adult women.

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Saliba, Louise F; Reis, Rodrigo S; Brownson, Ross C; Hino, Adriano A; Tureck, Luciane V; Valko, Cheryl; de Souza, Ricardo L R; Furtado-Alle, Lupe

2014-03-01

The individual response to diet may be influenced by gene polymorphisms. This study hypothesized that ADRB2 (Gln27Glu, rs1042714 and Arg16Gly, rs1042713), ADRB3 (Trp64Arg, rs4994) and GHRL (Leu72Met, rs696217) polymorphisms moderate weight loss. The study was a seven weeks dietary weight loss intervention with Brazilian adult obese women (n = 109). The body mass index (BMI) was calculated and polymorphisms in these genes were assessed by real-time PCR assays. Two-way repeated-measures ANOVA (2 × 2) were used to analyze the intervention effect between polymorphisms and BMI over the period and after stratification for age and socioeconomic status (SES). The weight loss intervention resulted in decreased BMI over the seven-week period (p polymorphism carriers and non-carriers, and although, the ADRB2, ADRB3 and GHRL polymorphisms did not moderate weight loss, the Gln27Glu polymorphism carriers showed a lower BMI compared to non-carriers in the low SES (p = 0.018) and the 30-39 y (p = 0.036) groups, suggesting a role for this polymorphism related to BMI control.

Obesity-related gene ADRB2, ADRB3 and GHRL polymorphisms and the response to a weight loss diet intervention in adult women

OpenAIRE

Saliba,Louise F.; Reis,Rodrigo S.; Brownson,Ross C.; Hino,Adriano A.; Tureck,Luciane V.; Valko,Cheryl; Souza,Ricardo L.R. de; Furtado-Alle,Lupe

2014-01-01

The individual response to diet may be influenced by gene polymorphisms. This study hypothesized that ADRB2 (Gln27Glu, rs1042714 and Arg16Gly, rs1042713), ADRB3 (Trp64Arg, rs4994) and GHRL (Leu72Met, rs696217) polymorphisms moderate weight loss. The study was a seven weeks dietary weight loss intervention with Brazilian adult obese women (n = 109). The body mass index (BMI) was calculated and polymorphisms in these genes were assessed by real-time PCR assays. Two-way repeated-measures ANOVA (...

Polymorphisms in DNA Repair Genes (APEX1, XPD, XRCC1 and XRCC3 and Risk of Preeclampsia in a Mexican Mestizo Population

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Ada Sandoval-Carrillo

2014-03-01

Full Text Available Variations in genes involved in DNA repair systems have been proposed as risk factors for the development of preeclampsia (PE. We conducted a case-control study to investigate the association of Human apurinic/apyrimidinic (AP endonuclease (APEX1 Asp148Glu (rs1130409, Xeroderma Pigmentosum group D (XPD Lys751Gln (rs13181, X-ray repair cross-complementing group 1 (XRCC Arg399Gln (rs25487 and X-ray repair cross-complementing group 3 (XRCC3 Thr241Met (rs861539 polymorphisms with PE in a Mexican population. Samples of 202 cases and 350 controls were genotyped using RTPCR. Association analyses based on a χ2 test and binary logistic regression were performed to determine the odds ratio (OR and a 95% confidence interval (95% CI for each polymorphism. The allelic frequencies of APEX1 Asp148Glu polymorphism showed statistical significant differences between preeclamptic and normal women (p = 0.036. Although neither of the polymorphisms proved to be a risk factor for the disease, the APEX1 Asp148Glu polymorphism showed a tendency of association (OR: 1.74, 95% CI = 0.96–3.14 and a significant trend (p for trend = 0.048. A subgroup analyses revealed differences in the allelic frequencies of APEX1 Asp148Glu polymorphism between women with mild preeclampsia and severe preeclampsia (p = 0.035. In conclusion, our results reveal no association between XPD Lys751Gln, XRCC Arg399Gln and XRCC3 Thr241Met polymorphisms and the risk of PE in a Mexican mestizo population; however, the results in the APEX1 Asp148Glu polymorphism suggest the need for future studies using a larger sample size.

Effect of protein solution components in the adsorption of Herbaspirillum seropedicae GlnB protein on mica.

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Ferreira, Cecília F G; Benelli, Elaine M; Klein, Jorge J; Schreiner, Wido; Camargo, Paulo C

2009-10-15

The adsorption of proteins and its buffer solution on mica surfaces was investigated by atomic force microscopy (AFM). Different salt concentration of the Herbaspirillum seropedicae GlnB protein (GlnB-Hs) solution deposited on mica was investigated. This protein is a globular, soluble homotrimer (36kDa), member of PII-like proteins family involved in signal transducing in prokaryote. Supramolecular structures were formed when this protein was deposited onto bare mica surface. The topographic AFM images of the GlnB-Hs films showed that at high salt concentration the supramolecular structures are spherical-like, instead of the typical doughnut-like shape for low salt concentration. AFM images of NaCl and Tris from the buffer solution showed structures with the same pattern as those observed for high salt protein solution, misleading the image interpretation. XPS experiments showed that GlnB protein film covers the mica surface without chemical reaction.

Roles of beta2-adrenergic receptor gene polymorphisms in a Turkish population with obstructive sleep apnea syndrome or obesity.

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Gök, I; Celebi, I; Hüseyinoğlu, N; Ozic, C

2014-10-20

We determined the distribution of the Arg16Gly and Gln27Glu polymorphisms of the beta-2 adrenergic receptor gene (ADRB2) in patients with obstructive sleep apnea syndrome as well as a control group in Northeastern Turkey. A total of 52 patients diagnosed with obstructive sleep apnea in a sleep laboratory and 78 control subjects were examined. Peripheral blood samples were taken from patients diagnosed with obstructive sleep apnea by polysomnography. DNA was extracted from blood samples and amplified using polymerase chain reaction. Amplification products were digested with restriction enzymes to investigate gene polymorphisms. Restriction products were extracted from agarose gel electrophoresis and polymorphisms were analyzed using gel images. The Arg16Gly polymorphism was observed in 18 of 52 patients and in 23 of 78 controls. The Gln27Glu polymorphism was observed in 21 of 52 patients and in 28 of 78 controls. In conclusion, there was no correlation among polymorphic frequencies between patient and control groups. Based on the results, these polymorphisms do not contribute to the clinical diagnosis of this syndrome. However, the distribution of Arg16Gly vs Gln27Glu polymorphisms may contribute to obesity in patients with a body mass index greater than 30 (P sleep apnea disease are changed.

Novel heterozygous nonsense mutation of the OPTN gene segregating in a Danish family with ALS

DEFF Research Database (Denmark)

Tümer, Zeynep; Bertelsen, Birgitte; Gredal, Ole

2012-01-01

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder. About 10% of ALS cases are familial (FALS) and the genetic defect is known only in approximately 20%-30% of these cases. The most common genetic cause of ALS is SOD1 (superoxide dismutase 1) mutation. Very recently......, mutations of the optineurin gene (OPTN), which is involved in open-angle glaucoma, were identified in 3 Japanese patients/families with ALS, and subsequently in a few FALS patients of European descent. We found a heterozygous nonsense mutation (c.493C>T, p.Gln165X, exon 6) in the OPTN gene in a Danish...... patient with ALS, and the mutation segregated from his affected father. The p.Gln165X mutation could not be detected in 1070 healthy Danish controls, in 1000 Danish individuals with metabolic phenotypes or in 64 sporadic ALS (SALS) cases. The p.Gln165X mutation described in this study is the first...

Associations between Dietary Patterns, ADRβ2 Gln27Glu and ADRβ3 Trp64Arg with Regard to Serum Triglyceride Levels: J-MICC Study

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Hinako Nanri

2016-09-01

Full Text Available Interactions between dietary patterns and 2 β-adrenergic receptor (ADRβ gene polymorphisms (ADRβ2 Gln27Glu and ADRβ3 Trp64Arg were examined with regard to the effects on serum triglyceride levels. The cross-sectional study comprised 1720 men and women (aged 35–69 years enrolled in the Japan Multi-Institutional Collaborative Cohort (J-MICC Study. Genotyping was conducted using a multiplex polymerase chain reaction-based invader assay. We used 46 items from a validated short food frequency questionnaire and examined major dietary patterns by factor analysis. We identified four dietary patterns: healthy, Western, seafood and bread patterns. There was no significant association between any dietary pattern and serum triglyceride levels. After a separate genotype-based analysis, significant interactions between ADRβ3 Trp64Arg genotype and the bread pattern (p for interaction = 0.01 were associated with serum triglyceride levels; specifically, after adjusting for confounding factors, Arg allele carriers with the bread pattern had lower serum triglycerides (p for trend = 0.01. However, the Trp/Trp homozygous subjects with the bread pattern showed no association with serum triglycerides (p for trend = 0.55. Interactions between other dietary patterns and ADRβ polymorphisms were not significant for serum triglyceride levels. Our findings suggest that ADRβ3 polymorphism modifies the effects of the bread pattern on triglyceride levels.

Association Between Genetic Polymorphisms in the XRCC1, XRCC3, XPD, GSTM1, GSTT1, MSH2, MLH1, MSH3, and MGMT Genes and Radiosensitivity in Breast Cancer Patients

International Nuclear Information System (INIS)

Mangoni, Monica; Bisanzi, Simonetta; Carozzi, Francesca; Sani, Cristina; Biti, Giampaolo; Livi, Lorenzo; Barletta, Emanuela; Costantini, Adele Seniori; Gorini, Giuseppe

2011-01-01

Purpose: Clinical radiosensitivity varies considerably among patients, and radiation-induced side effects developing in normal tissue can be therapy limiting. Some single nucleotide polymorphisms (SNPs) have been shown to correlate with hypersensitivity to radiotherapy. We conducted a prospective study of 87 female patients with breast cancer who received radiotherapy after breast surgery. We evaluated the association between acute skin reaction following radiotherapy and 11 genetic polymorphisms in DNA repair genes: XRCC1 (Arg399Gln and Arg194Trp), XRCC3 (Thr241Met), XPD (Asp312Asn and Lys751Gln), MSH2 (gIVS12-6T>C), MLH1 (Ile219Val), MSH3 (Ala1045Thr), MGMT (Leu84Phe), and in damage-detoxification GSTM1 and GSTT1 genes (allele deletion). Methods and Materials: Individual genetic polymorphisms were determined by polymerase chain reaction and single nucleotide primer extension for single nucleotide polymorphisms or by a multiplex polymerase chain reaction assay for deletion polymorphisms. The development of severe acute skin reaction (moist desquamation or interruption of radiotherapy due to toxicity) associated with genetic polymorphisms was modeled using Cox proportional hazards, accounting for cumulative biologically effective radiation dose. Results: Radiosensitivity developed in eight patients and was increased in carriers of variants XRCC3-241Met allele (hazard ratio [HR] unquantifiably high), MSH2 gIVS12-6nt-C allele (HR = 53.36; 95% confidence intervals [95% CI], 3.56-798.98), and MSH3-1045Ala allele (HR unquantifiably high). Carriers of XRCC1-Arg194Trp variant allele in combination with XRCC1-Arg399Gln wild-type allele had a significant risk of radiosensitivity (HR = 38.26; 95% CI, 1.19-1232.52). Conclusions: To our knowledge, this is the first report to find an association between MSH2 and MSH3 genetic variants and the development of radiosensitivity in breast cancer patients. Our findings suggest the hypothesis that mismatch repair mechanisms may be

Role of PII proteins in nitrogen fixation control of Herbaspirillum seropedicae strain SmR1.

Science.gov (United States)

Noindorf, Lilian; Bonatto, Ana C; Monteiro, Rose A; Souza, Emanuel M; Rigo, Liu U; Pedrosa, Fabio O; Steffens, Maria B R; Chubatsu, Leda S

2011-01-11

The PII protein family comprises homotrimeric proteins which act as transducers of the cellular nitrogen and carbon status in prokaryotes and plants. In Herbaspirillum seropedicae, two PII-like proteins (GlnB and GlnK), encoded by the genes glnB and glnK, were identified. The glnB gene is monocistronic and its expression is constitutive, while glnK is located in the nlmAglnKamtB operon and is expressed under nitrogen-limiting conditions. In order to determine the involvement of the H. seropedicae glnB and glnK gene products in nitrogen fixation, a series of mutant strains were constructed and characterized. The glnK- mutants were deficient in nitrogen fixation and they were complemented by plasmids expressing the GlnK protein or an N-truncated form of NifA. The nitrogenase post-translational control by ammonium was studied and the results showed that the glnK mutant is partially defective in nitrogenase inactivation upon addition of ammonium while the glnB mutant has a wild-type phenotype. Our results indicate that GlnK is mainly responsible for NifA activity regulation and ammonium-dependent post-translational regulation of nitrogenase in H. seropedicae.

Systems approach identifies an organic nitrogen-responsive gene network that is regulated by the master clock control gene CCA1.

Science.gov (United States)

Gutiérrez, Rodrigo A; Stokes, Trevor L; Thum, Karen; Xu, Xiaodong; Obertello, Mariana; Katari, Manpreet S; Tanurdzic, Milos; Dean, Alexis; Nero, Damion C; McClung, C Robertson; Coruzzi, Gloria M

2008-03-25

Understanding how nutrients affect gene expression will help us to understand the mechanisms controlling plant growth and development as a function of nutrient availability. Nitrate has been shown to serve as a signal for the control of gene expression in Arabidopsis. There is also evidence, on a gene-by-gene basis, that downstream products of nitrogen (N) assimilation such as glutamate (Glu) or glutamine (Gln) might serve as signals of organic N status that in turn regulate gene expression. To identify genome-wide responses to such organic N signals, Arabidopsis seedlings were transiently treated with ammonium nitrate in the presence or absence of MSX, an inhibitor of glutamine synthetase, resulting in a block of Glu/Gln synthesis. Genes that responded to organic N were identified as those whose response to ammonium nitrate treatment was blocked in the presence of MSX. We showed that some genes previously identified to be regulated by nitrate are under the control of an organic N-metabolite. Using an integrated network model of molecular interactions, we uncovered a subnetwork regulated by organic N that included CCA1 and target genes involved in N-assimilation. We validated some of the predicted interactions and showed that regulation of the master clock control gene CCA1 by Glu or a Glu-derived metabolite in turn regulates the expression of key N-assimilatory genes. Phase response curve analysis shows that distinct N-metabolites can advance or delay the CCA1 phase. Regulation of CCA1 by organic N signals may represent a novel input mechanism for N-nutrients to affect plant circadian clock function.

The Canonical Immediate Early 3 Gene Product pIE611 of Mouse Cytomegalovirus Is Dispensable for Viral Replication but Mediates Transcriptional and Posttranscriptional Regulation of Viral Gene Products.

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Rattay, Stephanie; Trilling, Mirko; Megger, Dominik A; Sitek, Barbara; Meyer, Helmut E; Hengel, Hartmut; Le-Trilling, Vu Thuy Khanh

2015-08-01

Transcription of mouse cytomegalovirus (MCMV) immediate early ie1 and ie3 is controlled by the major immediate early promoter/enhancer (MIEP) and requires differential splicing. Based on complete loss of genome replication of an MCMV mutant carrying a deletion of the ie3-specific exon 5, the multifunctional IE3 protein (611 amino acids; pIE611) is considered essential for viral replication. Our analysis of ie3 transcription resulted in the identification of novel ie3 isoforms derived from alternatively spliced ie3 transcripts. Construction of an IE3-hemagglutinin (IE3-HA) virus by insertion of an in-frame HA epitope sequence allowed detection of the IE3 isoforms in infected cells, verifying that the newly identified transcripts code for proteins. This prompted the construction of an MCMV mutant lacking ie611 but retaining the coding capacity for the newly identified isoforms ie453 and ie310. Using Δie611 MCMV, we demonstrated the dispensability of the canonical ie3 gene product pIE611 for viral replication. To determine the role of pIE611 for viral gene expression during MCMV infection in an unbiased global approach, we used label-free quantitative mass spectrometry to delineate pIE611-dependent changes of the MCMV proteome. Interestingly, further analysis revealed transcriptional as well as posttranscriptional regulation of MCMV gene products by pIE611. Cytomegaloviruses are pathogenic betaherpesviruses persisting in a lifelong latency from which reactivation can occur under conditions of immunosuppression, immunoimmaturity, or inflammation. The switch from latency to reactivation requires expression of immediate early genes. Therefore, understanding of immediate early gene regulation might add insights into viral pathogenesis. The mouse cytomegalovirus (MCMV) immediate early 3 protein (611 amino acids; pIE611) is considered essential for viral replication. The identification of novel protein isoforms derived from alternatively spliced ie3 transcripts prompted

Effects of Ala-Gln feeding strategies on growth, metabolism, and crowding stress resistance of juvenile Cyprinus carpio var. Jian.

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Chen, Xiu-Mei; Guo, Gui-Liang; Sun, Li; Yang, Qiu-Shi; Wang, Gui-Qin; Qin, Gui-Xin; Zhang, Dong-Ming

2016-04-01

The present study was conducted to evaluate the effects of different L-alanyl-l-glutamine (Ala-Gln) feeding strategies on the growth performance, metabolism and crowding stress resistance related parameters in juvenile Jian carp (Cyprinus carpio var. Jian) under crowded condition (80 g/L). Juvenile Jian carp (initial weight 26.1 ± 0.6 g) were distributed into five groups which fed with graded concentrations (0% or 1.0%) of Ala-Gln for eight weeks. Control group (I, 0/0) fed with control diet (0% Ala-Gln) throughout the feeding trial. The other four groups employed different control and experimental diet feeding strategies ranging from two weeks control diet fed and two weeks experimental diet (1% Ala-Gln) fed (II, 0/2) to eight weeks experimental diet fed (V, 4/4). Results revealed that Mean weight gain (MEG) under all different feeding strategies of Ala-Gln were significantly higher than that of the control group (p < 0.05), and MEG of group II (201.90%) was even higher than that of group IV (184.70%). Liver glycogen and blood total protein of groups II, III and V were significantly higher than that in groups I and IV (p < 0.05). The highest level of serum thyroxine (10.07 ng/ml), insulin-like growth factor-I (52.40 ng/ml) and insulin (9.73 μ IU/mL) were observed in group V. However, diet supplemented with Ala-Gln did not affect the levels of serum glucose, cortisol and catecholamine in fish. The mRNA expression of GR1a, GR1b and GR2 were also significantly changed in Ala-Gln supplementation groups compared with control group (p < 0.05). After fish intraperitoneally injected with virulent Aeromonas hydrophila, the fish survival rates were significantly increased in all Ala-Gln supplementation groups compared with control group (p < 0.05). Results from the present experiment showed the importance of dietary supplementation of Ala-Gln in benefaction of the growth performance, metabolism and crowding stress resistance in Jian carp breeding. The

Obesity-related gene ADRB2, ADRB3 and GHRL polymorphisms and the response to a weight loss diet intervention in adult women

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Louise F. Saliba

2014-01-01

Full Text Available The individual response to diet may be influenced by gene polymorphisms. This study hypothesized that ADRB2 (Gln27Glu, rs1042714 and Arg16Gly, rs1042713, ADRB3 (Trp64Arg, rs4994 and GHRL (Leu72Met, rs696217 polymorphisms moderate weight loss. The study was a seven weeks dietary weight loss intervention with Brazilian adult obese women (n = 109. The body mass index (BMI was calculated and polymorphisms in these genes were assessed by real-time PCR assays. Two-way repeated-measures ANOVA (2 x 2 were used to analyze the intervention effect between polymorphisms and BMI over the period and after stratification for age and socioeconomic status (SES. The weight loss intervention resulted in decreased BMI over the seven-week period (p < 0.001, for high and low SES (p < 0.05 and mainly for participants with 30-49 y. The intervention did not result in a statistically significant difference in weight loss between polymorphism carriers and non-carriers, and although, the ADRB2, ADRB3 and GHRL polymorphisms did not moderate weight loss, the Gln27Glu polymorphism carriers showed a lower BMI compared to non-carriers in the low SES (p = 0.018 and the 30-39 y (p = 0.036 groups, suggesting a role for this polymorphism related to BMI control.

Molecular dynamics characterization of the SAMHD1 Aicardi-Goutières Arg145Gln mutant: structural determinants for the impaired tetramerization

Science.gov (United States)

Cardamone, Francesca; Falconi, Mattia; Desideri, Alessandro

2018-05-01

Aicardi-Goutières syndrome, a rare genetic disorder characterized by calcification of basal ganglia, results in psychomotor delays and epilepsy states from the early months of children life. This disease is caused by mutations in seven different genes encoding proteins implicated in the metabolism of nucleic acids, including SAMHD1. Twenty SAMHD1 gene variants have been discovered and in this work, a structural characterization of the SAMHD1 Aicardi-Goutières Arg145Gln mutant is reported by classical molecular dynamics simulation. Four simulations have been carried out and compared. Two concerning the wild-type SAMHD1 form in presence and absence of cofactors, in order to explain the role of cofactors in the SAMHD1 assembly/disassembly process and, two concerning the Arg145Gln mutant, also in presence and absence of cofactors, in order to have an accurate comparison with the corresponding native forms. Results show the importance of native residue Arg145 in maintaining the tetramer, interacting with GTP cofactor inside allosteric sites. Replacement of arginine in glutamine gives rise to a loosening of GTP-protein interactions, when cofactors are present in allosteric sites, whilst in absence of cofactors, the occurrence of intra and inter-chain interactions is observed in the mutant, not seen in the native enzyme, making energetically unfavourable the tetramerization process.

Role of PII proteins in nitrogen fixation control of Herbaspirillum seropedicae strain SmR1

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Steffens Maria BR

2011-01-01

Full Text Available Abstract Background The PII protein family comprises homotrimeric proteins which act as transducers of the cellular nitrogen and carbon status in prokaryotes and plants. In Herbaspirillum seropedicae, two PII-like proteins (GlnB and GlnK, encoded by the genes glnB and glnK, were identified. The glnB gene is monocistronic and its expression is constitutive, while glnK is located in the nlmAglnKamtB operon and is expressed under nitrogen-limiting conditions. Results In order to determine the involvement of the H. seropedicae glnB and glnK gene products in nitrogen fixation, a series of mutant strains were constructed and characterized. The glnK- mutants were deficient in nitrogen fixation and they were complemented by plasmids expressing the GlnK protein or an N-truncated form of NifA. The nitrogenase post-translational control by ammonium was studied and the results showed that the glnK mutant is partially defective in nitrogenase inactivation upon addition of ammonium while the glnB mutant has a wild-type phenotype. Conclusions Our results indicate that GlnK is mainly responsible for NifA activity regulation and ammonium-dependent post-translational regulation of nitrogenase in H. seropedicae.

GLN standard as a facilitator of physical location identification within process of distribution

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Davor Dujak

2017-09-01

Full Text Available Background: Distribution, from the business point of view, is a set of decisions and actions that will provide the right products at the right time and place, in line with customer expectations. It is a process that generates significant cost, but also effectively implemented, significantly affects the positive perception of the company. Institute of Logistics and Warehousing (IliM, based on the research results related to the optimization of the distribution network and consulting projects for companies, indicates the high importance of the correct description of the physical location within the supply chains in order to make transport processes more effective. Individual companies work on their own geocoding of warehouse locations and location of their business partners (suppliers, customers, but the lack of standardization in this area causes delays related to delivery problems with reaching the right destination. Furthermore, the cooperating companies do not have a precise indication of the operating conditions of each location, e.g. Time windows of the plant, logistic units accepted at parties, supported transport etc. Lack of this information generates additional costs associated with re-operation and the costs of lost benefits for the lack of goods on time. The solution to this problem seems to be a wide-scale implementation of GS1 standard which is the Global Location Number (GLN, that, thanks to a broad base of information will assist the distribution processes. Material and methods: The results of survey conducted among Polish companies in the second half of 2016 indicate an unsatisfactory degree of implementation of the transport processes, resulting from incorrect or inaccurate description of the location, and thus, a significant number of errors in deliveries. Accordingly, authors studied literature and examined case studies indicating the possibility of using GLN standard to identify the physical location and to show the

Role of blood ribosomal protein S19 in coagulum resorption: a study using Gln137Glu-ribosomal protein S19 gene knock-in mouse.

Science.gov (United States)

Chen, Jun; Fujino, Rika; Zhao, Rui; Semba, Umeko; Araki, Kimi; Yamamoto, Tetsuro

2014-11-01

Sera of human, guinea pig or mouse contain a strong monocyte chemoattractant capacity that is attributed to the ribosomal protein S19 (RP S19) oligomers generated during blood coagulation. In contrast, sera prepared from Gln137Glu-RP S19 gene knock-in mice contained negligible chemoattractant capacity. When coagula that had been pre-formed from the blood of both the wild type and knock-in mice were intraperitoneally inserted into host mice, after 3 days of recovery, the knock-in mouse coagula remained larger than the wild type mouse coagula. The wild type mouse coagula were covered by multiple macrophage layers at the surface and were infiltrated inside by macrophages. Knock-in mouse coagula exhibited less macrophage involvement. When coagula of knock-in mice and coagula of knock-in mice containing C5a/RP S19, an artificial substitute of the RP S19 oligomers, were intraperitoneally inserted as pairs, the C5a/RP S19 containing coagulum was more rapidly absorbed, concomitant with increased macrophage involvement. Finally, when the knock-in mouse and wild type mouse coagula pairs were inserted into mice in which macrophages had been depleted using clodronate liposome, the size difference of recovered coagula was reversed. These results indicate the importance of the RP S19 oligomer-induced macrophage recruitment in coagulum resorption. © 2014 Japanese Society of Pathology and Wiley Publishing Asia Pty Ltd.

Enhancement of antibiotic productions by engineered nitrate utilization in actinomycetes.

Science.gov (United States)

Meng, Sitong; Wu, Hang; Wang, Lei; Zhang, Buchang; Bai, Linquan

2017-07-01

Nitrate is necessary for primary and secondary metabolism of actinomycetes and stimulates the production of a few antibiotics, such as lincomycin and rifamycin. However, the mechanism of this nitrate-stimulating effect was not fully understood. Two putative ABC-type nitrate transporters were identified in Streptomyces lincolnensis NRRL2936 and verified to be involved in lincomycin biosynthesis. With nitrate supplementation, the transcription of nitrogen assimilation genes, nitrate-specific ABC1 transporter genes, and lincomycin exporter gene lmrA was found to be enhanced and positively regulated by the global regulator GlnR, whose expression was also improved. Moreover, heterologous expression of ABC2 transporter genes in Streptomyces coelicolor M145 resulted in an increased actinorhodin production. Further incorporation of a nitrite-specific transporter gene nirC, as in nirC-ABC2 cassette, led to an even higher actinorhodin production. Similarly, the titers of salinomycin, ansamitocin, lincomycin, and geldanamycin were increased with the integration of this cassette to Streptomyces albus BK3-25, Actinosynnema pretiosum ATCC31280, S. lincolnensis LC-G, and Streptomyces hygroscopicus XM201, respectively. Our work expanded the nitrate-stimulating effect to many antibiotic producers by utilizing the nirC-ABC2 cassette for enhanced nitrate utilization, which could become a general tool for titer increase of antibiotics in actinomycetes.

Deletion of meso-2,3-butanediol dehydrogenase gene budC for enhanced D-2,3-butanediol production in Bacillus licheniformis

Science.gov (United States)

2014-01-01

Background D-2,3-butanediol has many industrial applications such as chiral reagents, solvents, anti-freeze agents, and low freezing point fuels. Traditional D-2,3-butanediol producing microorganisms, such as Klebsiella pneumonia and K. xoytoca, are pathogenic and not capable of producing D-2,3-butanediol at high optical purity. Bacillus licheniformis is a potential 2,3-butanediol producer but the wild type strain (WX-02) produces a mix of D- and meso-type isomers. BudC in B. licheniformis is annotated as 2,3-butanediol dehydrogenase or acetoin reductase, but no pervious experiment was performed to verify this hypothesis. Results We developed a genetically modified strain of B. licheniformis (WX-02 ΔbudC) as a D-2,3-butanediol producer with high optimal purity. A marker-less gene deletion protocol based on a temperature sensitive knock-out plasmid T2-Ori was used to knock out the budC gene in B. licheniformis WX-02. The budC knock-out strain successfully abolished meso-2,3-butanediol production with enhanced D-2,3-butanediol production. No meso-BDH activity was detectable in cells of this strain. On the other hand, the complementary strain restored the characteristics of wild strain, and produced meso-2,3-butanediol and possessed meso-BDH activity. All of these data suggested that budC encoded the major meso-BDH catalyzing the reversible reaction from acetoin to meso-2,3-butanediol in B. licheniformis. The budC knock-out strain produced D-2,3-butanediol isomer only with a high yield of 30.76 g/L and a productivity of 1.28 g/L-h. Conclusions We confirmed the hypothesis that budC gene is responsible to reversibly transfer acetoin to meso-2,3-butanediol in B. licheniformis. A mutant strain of B. licheniformis with depleted budC gene was successfully developed and produced high level of the D-2,3-butanediol with high optimal purity. PMID:24475980

Frequency of distribution of leptin receptor gene polymorphism in obstructive sleep apnea patients.

Science.gov (United States)

Popko, K; Gorska, E; Wasik, M; Stoklosa, A; Pływaczewski, R; Winiarska, M; Gorecka, D; Sliwinski, P; Demkow, U

2007-11-01

Leptin is an adipocyte-derived hormone regulating energy homeostasis and body weight. Leptin concentration is increased in patients with the obstructive sleep apnea syndrome (OSAS). Leptin receptor (LEPR) is a single transmembrane protein belonging to the superfamily of cytokine receptors related by a structure to the hemopoietin receptor family. The aim of the present study was to evaluate the frequency of distribution of leptin receptor gene polymorphism GLN223ARG in OSAS patients compared with healthy controls. The examined group included 179 subjects: 102 OSAS patients (74 men and 28 women) and 77 non-apneic controls (39 men and 38 women). Genomic DNA was isolated with the use of a column method and genotyping of DNA sequence variation was carried out by restriction enzyme analysis of PCR-amplified DNA. The results revealed a significant correlation between the polymorphism of LEPR and OSAS. Carriers of Arg allele in homozygotic genotype Arg/Arg and heterozygotic genotype Gln/Arg were more often obese and developed OSAS than the group of carriers of homozygotic Gln/Gln genotype. This tendency was observed in the whole examined population and in the group of obese women. We also found the highest levels of total cholesterol, LDL, HDL, and triglycerides in the group of homozygotic Arg/Arg genotype carriers, lower in heterozygotic Gln/Arg genotype carriers, and the lowest in the group of persons carring homozygotic Gln/Gln genotype. The presence of Arg allel seems linked to a higher risk of obesity and higher lipid levels in OSAS patients. OSAS may have a strong genetic basis due to the effects from a variety of genes including those for leptin receptor.

Synthesis of bacteriophage-coded gene products during infection of Escherichia coli with amber mutants of T3 and T7 defective in gene 1

DEFF Research Database (Denmark)

Issinger, O G; Hausmann, R

1973-01-01

During nonpermissive infection by a T7 amber mutant in gene 1 (phage RNA polymerase-deficient), synthesis of the products of the phage genes 3 (endonuclease), 3, 5 (lysozyme), 5 (DNA polymerase), and 17 (serum blocking power) was shown to occur at about half the rate as during wild-type infection...

Polymorphisms in base excision repair genes as colorectal cancer risk factors and modifiers of the effect of diets high in red meat.

Science.gov (United States)

Brevik, Asgeir; Joshi, Amit D; Corral, Román; Onland-Moret, N Charlotte; Siegmund, Kimberly D; Le Marchand, Loïc; Baron, John A; Martinez, Maria Elena; Haile, Robert W; Ahnen, Dennis J; Sandler, Robert S; Lance, Peter; Stern, Mariana C

2010-12-01

A diet high in red meat is an established colorectal cancer (CRC) risk factor. Carcinogens generated during meat cooking have been implicated as causal agents and can induce oxidative DNA damage, which elicits repair by the base excision repair (BER) pathway. Using a family-based study, we investigated the role of polymorphisms in 4 BER genes (APEX1 Gln51His, Asp148Glu; OGG1 Ser236Cys; PARP Val742Ala; and XRCC1 Arg194Trp, Arg280His, Arg399Gln) as potential CRC risk factors and modifiers of the association between diets high in red meat or poultry and CRC risk. We tested for gene-environment interactions using case-only analyses (n = 577) and compared statistically significant results with those obtained using case-unaffected sibling comparisons (n = 307 sibships). Carriers of the APEX1 codon 51 Gln/His genotype had a reduced CRC risk compared with carriers of the Gln/Gln genotype (odds ratio (OR) = 0.15, 95% CI = 0.03-0.69, P = 0.015). The association between higher red meat intake (>3 servings per week) and CRC was modified by the PARP Val762Ala single-nucleotide polymorphisms (SNP; case-only interaction P = 0.026). This SNP also modified the association between higher intake of high-temperature cooked red meat (case-only interaction P = 0.0009). We report evidence that the BER pathway PARP gene modifies the association of diets high in red meat cooked at high temperatures with risk of CRC. Our findings suggest a contribution to colorectal carcinogenesis of free radical damage as one of the possible harmful effects of a diet high in red meat. ©2010 AACR.

Mathematical model of the binding of allosteric effectors to the Escherichia coli PII signal transduction protein GlnB.

Science.gov (United States)

da Rocha, Ricardo Alves; Weschenfelder, Thiago André; de Castilhos, Fernanda; de Souza, Emanuel Maltempi; Huergo, Luciano Fernandes; Mitchell, David Alexander

2013-04-16

PII proteins are important regulators of nitrogen metabolism in a wide variety of organisms: the binding of the allosteric effectors ATP, ADP, and 2-oxoglutarate (2-OG) to PII proteins affects their ability to interact with target proteins. We modeled the simultaneous binding of ATP, ADP, and 2-OG to one PII protein, namely GlnB of Escherichia coli, using a modeling approach that allows the prediction of the proportions of individual binding states. Four models with different binding rules were compared. We selected one of these models (that assumes that the binding of the first nucleotide to GlnB makes it harder for subsequent nucleotides to bind) and used it to explore how physiological concentrations of ATP, ADP, and 2-OG would affect the proportions of those states of GlnB that interact with the target proteins ATase and NtrB. Our simulations indicate that GlnB can, as suggested by previous researchers, act as a sensor of both 2-OG and the ATP:ADP ratio. We conclude that our modeling approach will be an important tool in future studies concerning the PII binding states and their interactions with target proteins.

An Intelligent Method of Product Scheme Design Based on Product Gene

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Qing Song Ai

2013-01-01

Full Text Available Nowadays, in order to have some featured products, many customers tend to buy customized products instead of buying common ones in supermarket. The manufacturing enterprises, with the purpose of improving their competitiveness, are focusing on providing customized products with high quality and low cost as well. At present, how to produce customized products rapidly and cheaply has been the key challenge to manufacturing enterprises. In this paper, an intelligent modeling approach applied to supporting the modeling of customized products is proposed, which may improve the efficiency during the product design process. Specifically, the product gene (PG method, which is an analogy of biological evolution in engineering area, is employed to model products in a new way. Based on product gene, we focus on the intelligent modeling method to generate product schemes rapidly and automatically. The process of our research includes three steps: (1 develop a product gene model for customized products; (2 find the obtainment and storage method for product gene; and (3 propose a specific genetic algorithm used for calculating the solution of customized product and generating new product schemes. Finally, a case study is applied to test the usefulness of our study.

Association of Gln27Glu polymorphism of the β-2- adrenergic ...

African Journals Online (AJOL)

A group of 66 patients with preeclampsia and 72 control subjects were analyzed for the Arg16Gly and Gln27Glu polymorphisms of the ADRB2 by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Comparisons of the ADRB2 genotypes or alleles between different groups were performed ...

High Production of 2,3-Butanediol (2,3-BD by Raoultella ornithinolytica B6 via Optimizing Fermentation Conditions and Overexpressing 2,3-BD Synthesis Genes.

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Taeyeon Kim

Full Text Available Biological production of 2,3-butandiol (2,3-BD has received great attention as an alternative to the petroleum-based 2,3-BD production. In this study, a high production of 2,3-BD in fed-batch fermentation was investigated with a newly isolated bacterium designated as Raoultella ornithinolytica B6. The isolate produced 2,3-BD as the main product using hexoses (glucose, galactose, and fructose, pentose (xylose and disaccharide (sucrose. The effects of temperature, pH-control schemes, and agitation speeds on 2,3-BD production were explored to optimize the fermentation conditions. Notably, cell growth and 2,3-BD production by R. ornithinolytica B6 were higher at 25°C than at 30°C. When three pH control schemes (no pH control, pH control at 7, and pH control at 5.5 after the pH was decreased to 5.5 during fermentation were tested, the best 2,3-BD titer and productivity along with reduced by-product formation were achieved with pH control at 5.5. Among different agitation speeds (300, 400, and 500 rpm, the optimum agitation speed was 400 rpm with 2,3-BD titer of 68.27 g/L, but acetic acid was accumulated up to 23.32 g/L. Further enhancement of the 2,3-BD titer (112.19 g/L, yield (0.38 g/g, and productivity (1.35 g/L/h as well as a significant reduction of acetic acid accumulation (9.71 g/L was achieved by the overexpression of homologous budABC genes, the 2,3-BD-synthesis genes involved in the conversion of pyruvate to 2,3-BD. This is the first report presenting a high 2,3-BD production by R.ornithinolytica which has attracted little attention with respect to 2,3-BD production, extending the microbial spectrum of 2,3-BD producers.

Overexpression of the Squalene Epoxidase Gene Alone and in Combination with the 3-Hydroxy-3-methylglutaryl Coenzyme A Gene Increases Ganoderic Acid Production in Ganoderma lingzhi.

Science.gov (United States)

Zhang, De-Huai; Jiang, Lu-Xi; Li, Na; Yu, Xuya; Zhao, Peng; Li, Tao; Xu, Jun-Wei

2017-06-14

The squalene epoxidase (SE) gene from the biosynthetic pathway of ganoderic acid (GA) was cloned and overexpressed in Ganoderma lingzhi. The strain that overexpressed the SE produced approximately 2 times more GA molecules than the wild-type (WT) strain. Moreover, SE overexpression upregulated lanosterol synthase gene expression in the biosynthetic pathway. These results indicated that SE stimulates GA accumulation. Then, the SE and 3-hydroxy-3-methylglutaryl coenzyme A (HMGR) genes were simultaneously overexpressed in G. lingzhi. Compared with the individual overexpression of SE or HMGR, the combined overexpression of the two genes further enhanced individual GA production. The overexpressing strain produced maximum GA-T, GA-S, GA-Mk, and GA-Me contents of 90.4 ± 7.5, 35.9 ± 5.4, 6.2 ± 0.5, and 61.8 ± 5.8 μg/100 mg dry weight, respectively. These values were 5.9, 4.5, 2.4, and 5.8 times higher than those produced by the WT strain. This is the first example of the successful manipulation of multiple biosynthetic genes to improve GA content in G. lingzhi.

Arithmetic differential equations on $GL_n$, I: differential cocycles

OpenAIRE

Buium, Alexandru; Dupuy, Taylor

2013-01-01

The theory of differential equations has an arithmetic analogue in which derivatives are replaced by Fermat quotients. One can then ask what is the arithmetic analogue of a linear differential equation. The study of usual linear differential equations is the same as the study of the differential cocycle from $GL_n$ into its Lie algebra given by the logarithmic derivative. However we prove here that there are no such cocycles in the context of arithmetic differential equations. In sequels of t...

A novel syndrome of autosomal-dominant hyperinsulinemic hypoglycemia linked to a mutation in the human insulin receptor gene

DEFF Research Database (Denmark)

Højlund, Kurt; Hansen, Torben; Lajer, Maria

2004-01-01

a missense mutation (Arg1174Gln) in the tyrosine kinase domain of the insulin receptor gene that cosegregated with the disease phenotype (logarithm of odds [LOD] score 3.21). In conclusion, we report a novel syndrome of autosomal-dominant hyperinsulinemic hypoglycemia. The findings demonstrate...

Single nucleotide polymorphisms in the SEPTIN12 gene may be associated with azoospermia by meiotic arrest in Japanese men.

Science.gov (United States)

Miyamoto, Toshinobu; Tsujimura, Akira; Miyagawa, Yasushi; Koh, Eitetsu; Namiki, Mikio; Horikawa, Michiharu; Saijo, Yasuaki; Sengoku, Kazuo

2012-01-01

To investigate the association between SEPTIN12 gene variants and the risk of azoospermia caused by meiotic arrest. Mutational analysis of the SEPTIN12 gene was performed using DNA from 30 Japanese patients with azoospermia by meiotic arrest and 140 fertile male controls. The frequencies of the c.204G>C (Gln38His) allele and the CC genotype were significantly higher in patients than in fertile controls (p C (Gln38His) variant in the SEPTIN12 gene was associated with increased susceptibility to azoospermia caused by meiotic arrest.

ERCC1 Cys8092Ala and XRCC1 Arg399Gln polymorphisms predict progression-free survival after curative radiotherapy for nasopharyngeal carcinoma.

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Hekun Jin

Full Text Available BACKGROUND: Single nucleotide polymorphisms (SNPs in DNA repair genes can alter gene expression and activity and affect response to cancer treatment and, correspondingly, survival. The present study was designed to evaluate the utility of the XRCC1 Arg399Gln and ERCC1 Cys8092Ala SNPs, measured in pretreatment biopsy samples, as predictors of response to radiotherapy in patients with non-metastatic nasopharyngeal carcinoma (NPC. MATERIALS AND METHODS: The study included 75 consecutive patients with stage II-IVA-B NPC. XRCC1 Arg399Glu and ERCC1 Cys8092Ala SNPs were identified from paraffin-embedded biopsy specimens via Sanger sequencing. Expression of p53 and pAkt protein was analyzed by immunohistochemical staining. Potential relationships between genetic polymorphisms and progression-free survival (PFS were analyzed by using a Cox proportional hazards model, the Kaplan-Meier method, and the log-rank test. RESULTS: Multivariate analysis showed that carriers of the ERCC1 8092 Ala/Ala genotype [hazard ratio (HR 1.882; 95% confidence interval (CI 1.031-3.438; P = 0.039] and heavy smokers (≥20 pack-years carrying the XRCC1 Arg/Arg genotype (HR 2.019; 95% CI 1.010-4.036; P = 0.047 had significantly lower PFS rates. Moreover, combined positive expression of p53 and pAkt led to significantly increased PFS in subgroups carrying the XRCC1 Gln allele (HR 7.057; 95% CI 2.073-24.021; P = 0.002 or the ERCC1 Cys allele (HR 2.568; 95% CI 1.056-6.248; P = 0.038. CONCLUSIONS: The ERCC1 Cys8092Ala polymorphism is an independent predictor of response to radiotherapy for NPC, and the XRCC1 Arg399Glu mutation combined with smoking status seems to predict PFS as well. Our results further suggest a possible correlation between these genetic polymorphisms and p53 protein status on survival.

Adsorption of protein GlnB of Herbaspirillum seropedicae on Si(111) investigated by AFM and XPS.

Science.gov (United States)

Lubambo, A F; Benelli, E M; Klein, J; Schreiner, W; Camargo, P C

2006-01-01

The protein GlnB-Hs (GlnB of Herbaspirillum seropedicae) in diazotroph micro-organisms signalizes levels of nitrogen, carbon, and energy for a series of proteins involved in the regulation of expression and control of the activity of nitrogenase complex that converts atmospheric nitrogen in ammonia, resulting in biological nitrogen fixation. Its structure has already been determined by X-ray diffraction, revealing a trimer of (36 kDa) with lateral cavities having hydrophilic boundaries. The interactions of GlnB-Hs with the well-known Si(111) surface were investigated for different incubation times, protein concentrations in initial solution, deposition conditions, and substrate initial state. The protein solution was deposited on Si(111) and dried under controlled conditions. An atomic force microscope operating in dynamic mode shows images of circular, linear, and more complex donut-shaped protein arrangement, and also filament types of organization, which vary from a few nanometers to micrometers. Apparently, the filament formation was favored because of protein surface polarity when in contact with the silicon surface, following some specific orientation. The spin-coating technique was successfully used to obtain more uniform surface covering.

Effect of deletion of 2,3-butanediol dehydrogenase gene (bdhA) on acetoin production of Bacillus subtilis.

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Zhang, Junjiao; Zhao, Xiangying; Zhang, Jiaxiang; Zhao, Chen; Liu, Jianjun; Tian, Yanjun; Yang, Liping

2017-09-14

The present work aims to block 2,3-butanediol synthesis in acetoin fermentation of Bacillus subtilis. First, we constructed a recombinant strain BS168D by deleting the 2,3-butanediol dehydrogenase gene bdhA of the B. subtilis168, and there was almost no 2,3-butanediol production in 20 g/L of glucose media. The acetoin yield of BS168D reached 6.61 g/L, which was about 1.5 times higher than that of the control B. subtilis168 (4.47 g/L). Then, when the glucose concentration was increased to 100 g/L, the acetoin yield reached 24.6 g/L, but 2.4 g/L of 2,3-butanediol was detected at the end of fermentation. The analysis of 2,3-butanediol chiral structure indicated that the main 2,3-butanediol production of BS168D was meso-2,3-butanediol, and the bdhA gene was only responsible for (2R,3R)-2,3-butanediol synthesis. Therefore, we speculated that there may exit another pathway relating to the meso-2,3-butanediol synthesis in the B. subtilis. In addition, the results of low oxygen condition fermentation showed that deletion of bdhA gene successfully blocked the reversible transformation between acetoin and 2,3-butanediol and eliminated the effect of dissolved oxygen on the transformation.

Heterozygosity for the Mood Disorder-Associated Variant Gln460Arg Alters P2X7 Receptor Function and Sleep Quality.

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Metzger, Michael W; Walser, Sandra M; Dedic, Nina; Aprile-Garcia, Fernando; Jakubcakova, Vladimira; Adamczyk, Marek; Webb, Katharine J; Uhr, Manfred; Refojo, Damian; Schmidt, Mathias V; Friess, Elisabeth; Steiger, Axel; Kimura, Mayumi; Chen, Alon; Holsboer, Florian; Arzt, Eduardo; Wurst, Wolfgang; Deussing, Jan M

2017-11-29

A single nucleotide polymorphism substitution from glutamine (Gln, Q) to arginine (Arg, R) at codon 460 of the purinergic P2X7 receptor (P2X7R) has repeatedly been associated with mood disorders. The P2X7R-Gln460Arg variant per se is not compromised in its function. However, heterologous expression of P2X7R-Gln460Arg together with wild-type P2X7R has recently been demonstrated to impair receptor function. Here we show that this also applies to humanized mice coexpressing both human P2X7R variants. Primary hippocampal cells derived from heterozygous mice showed an attenuated calcium uptake upon agonist stimulation. While humanized mice were unaffected in their behavioral repertoire under basal housing conditions, mice that harbor both P2X7R variants showed alterations in their sleep quality resembling signs of a prodromal disease stage. Also healthy heterozygous human subjects showed mild changes in sleep parameters. These results indicate that heterozygosity for the wild-type P2X7R and its mood disorder-associated variant P2X7R-Gln460Arg represents a genetic risk factor, which is potentially able to convey susceptibility to mood disorders. SIGNIFICANCE STATEMENT Depression and bipolar disorder are the most common mood disorders. The P2X7 receptor (P2X7R) regulates many cellular functions. Its polymorphic variant Gln460Arg has repeatedly been associated with mood disorders. Genetically engineered mice, with human P2X7R, revealed that heterozygous mice (i.e., they coexpress the disease-associated Gln460Arg variant together with its normal version) have impaired receptor function and showed sleep disturbances. Human participants with the heterozygote genotype also had subtle alterations in their sleep profile. Our findings suggest that altered P2X7R function in heterozygote individuals disturbs sleep and might increase the risk for developing mood disorders. Copyright © 2017 the authors 0270-6474/17/3711688-13$15.00/0.

Gene analogue finder: a GRID solution for finding functionally analogous gene products

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Licciulli Flavio

2007-09-01

Full Text Available Abstract Background To date more than 2,1 million gene products from more than 100000 different species have been described specifying their function, the processes they are involved in and their cellular localization using a very well defined and structured vocabulary, the gene ontology (GO. Such vast, well defined knowledge opens the possibility of compare gene products at the level of functionality, finding gene products which have a similar function or are involved in similar biological processes without relying on the conventional sequence similarity approach. Comparisons within such a large space of knowledge are highly data and computing intensive. For this reason this project was based upon the use of the computational GRID, a technology offering large computing and storage resources. Results We have developed a tool, GENe AnaloGue FINdEr (ENGINE that parallelizes the search process and distributes the calculation and data over the computational GRID, splitting the process into many sub-processes and joining the calculation and the data on the same machine and therefore completing the whole search in about 3 days instead of occupying one single machine for more than 5 CPU years. The results of the functional comparison contain potential functional analogues for more than 79000 gene products from the most important species. 46% of the analyzed gene products are well enough described for such an analysis to individuate functional analogues, such as well-known members of the same gene family, or gene products with similar functions which would never have been associated by standard methods. Conclusion ENGINE has produced a list of potential functionally analogous relations between gene products within and between species using, in place of the sequence, the gene description of the GO, thus demonstrating the potential of the GO. However, the current limiting factor is the quality of the associations of many gene products from non

The presence of c-erbB-2 gene product-related protein in culture medium conditioned by breast cancer cell line SK-BR-3

International Nuclear Information System (INIS)

Alper, O.; Yamaguchi, K.; Hitomi, J.; Honda, S.; Matsushima, T.; Abe, K.

1990-01-01

The Mr 185,000 glycoprotein encoded by human c-erbB-2/neu/HER2 gene, termed c-erbB-2 gene product, shows a close structural similarity with epidermal growth factor receptor and is now regarded to be a growth factor receptor for an as yet unidentified ligand. Abundant c-erbB-2 mRNA was demonstrated by Northern blot studies in the human breast cancer cell line SK-BR-3. Cellular radiolabeling experiments followed by immunoprecipitation with three different anti-c-erbB-2 gene product antibodies, recognizing extracellular domain, kinase domain, and carboxyl-terminal portion, respectively, demonstrated the production of a large amount of c-erbB-2 gene product which had the capacity to be phosphorylated. Immunization of mice with concentrated culture medium conditioned by SK-BR-3 cells always generated antibodies against c-erbB-2 gene product, demonstrating that this culture medium contained substance(s) immunologically indistinguishable from c-erbB-2 gene product. This observation was supported by the successful development of a monoclonal antibody against c-erbB-2 gene product, GFD-OA-p185-1, by immunizing mice with this culture medium. The biochemical nature of the substance(s) present in the culture medium was further characterized. When the culture medium conditioned by [35S]cysteine-labeled SK-BR-3 cells was immunoprecipitated by three different anti-c-erbB-2 gene product antibodies, only the antibody recognizing extracellular domain precipitated the [35S]-labeled protein with a molecular weight of 110,000, namely p110. The newly developed monoclonal antibody also immunoprecipitated this protein

Ghrelin gene polymorphisms in rheumatoid arthritis.

Science.gov (United States)

Ozgen, Metin; Koca, Suleyman Serdar; Etem, Ebru Onalan; Yuce, Huseyin; Aydin, Suleyman; Isik, Ahmet

2011-07-01

Ghrelin, an endogenous orexigenic peptide, has anti-inflammatory effects, down-regulates pro-inflammatory cytokines, and its altered levels are reported in various inflammatory diseases. The human preproghrelin (ghrelin/obestatin) gene shows several single nucleotide polymorphisms (SNPs) including Arg51Gln, Leu72Met, Gln90Leu, and A-501C. The aim of this study was to investigate the frequency, and clinical significance, of these four SNPs in a small cohort of Turkish patients with rheumatoid arthritis (RA). The study included 103 patients with RA and 103 healthy controls. In the RA group, disease activity and disease-related damage were assessed using the Disease Activity Score-28 (DAS-28), and the modified Larsen scoring (MLS) methods. In all the participants, genomic DNA was isolated and genotyped by polymerase chain reaction and restriction fragment length polymorphism analysis. The frequencies of ghrelin gene SNPs were 82.5 and 79.6% in the RA and control groups, respectively, and there were no significant differences in terms of genotype distributions and allele frequencies for these four SNPs between the groups. However, the A-501C SNP was found to be associated with early disease onset, and Gln90Leu SNP with less frequent rheumatoid factor positivity, in the RA group. A-501C SNP is associated with earlier onset of RA suggesting that genetic variations in the ghrelin gene may have an impact on RA. Copyright © 2010 Société française de rhumatologie. Published by Elsevier SAS. All rights reserved.

Leptin responsiveness to energy restriction: genetic variation in the leptin receptor gene.

Science.gov (United States)

Mars, Monica; van Rossum, Caroline T M; de Graaf, Cees; Hoebee, Barbara; De Groot, Lisette C P G M; Kok, Frans J

2004-03-01

Serum leptin concentrations are an important afferent signal in energy balance homeostasis. It has been speculated that the leptin responsiveness to energy restriction is affected by the functionality of the leptin receptor. The purpose of this analysis was to explore the effect of polymorphisms in the LEPR gene on the acute decline in leptin after 4 days of 65% energy restriction. Leptin concentrations of the study group (n = 44; all men) declined by 2.3 +/- 1.5 micro g/L [-39.4% (95% confidence interval: -43.6 to -34.9)]. Leptin responses did not statistically differ between noncarriers and carriers of three mutant variants of the polymorphisms: Lys109/Lys109 (-41.4%) vs. Arg109/+ (-37.0%) (p = 0.33); Gln223/Gln223 (-41.5%) vs. Arg223/+ (-37.8%) (p = 0.40); Lys656/Lys656 (-39.5%) vs. Asn656/+ (-39.3%) (p = 0.96). No effect of the assessed polymorphisms in the LEPR gene on the acute decline in leptin after energy restriction was observed. Power calculations are provided for future studies on the leptin responsiveness to energy restriction.

Omission and Resupply of Nitrogen Affect Physiological and Enzymatic Activities and the Gene Expression of Eucalypt Clones

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Loane Vaz Fernandes

Full Text Available ABSTRACT: The mineral nutrient uptake of plants in the field occurs in pulses, due to variations in the substance concentrations at the root surface. The fluctuations in nutrient supply probably induce changes in the plant, which are to date unknown for Eucalyptus. This study evaluated these changes in plant growth, nutritional status, photosynthesis, and gene expression, which can serve as biomarkers of the nitrogen status, of four eucalypt clones exposed to N omission and resupply. A greenhouse experiment with four Eucalyptus clones was installed, and after initial growth exposed to N omission for 21 d, followed by N resupply in nutrient solution for 14 d. Nitrogen omission decreased the total N and photosynthetic pigments, net photosynthesis and photochemical dissipation, and increased enzyme activity especially in leaves and the gene expression in leaves and roots. Nitrogen resupply decreased these variations, indicating recovery. The total N concentration was highly and significantly correlated with net photosynthesis, enzyme activity, expression of genes GS2;1 and Gln1;3 in the leaves and AMT1;2 in the roots, contents of chlorophyll a and b, and photochemical energy dissipation. The enzymes GS and NR in the leaves and the genes AMT1;2, GS2;1 and Gln1;3 proved to be sensitive N indicators.

Transforming growth factor-β-induced gene product-h3 inhibits odontoblastic differentiation of dental pulp cells.

Science.gov (United States)

Serita, Suguru; Tomokiyo, Atsushi; Hasegawa, Daigaku; Hamano, Sayuri; Sugii, Hideki; Yoshida, Shinichiro; Mizumachi, Hiroyuki; Mitarai, Hiromi; Monnouchi, Satoshi; Wada, Naohisa; Maeda, Hidefumi

2017-06-01

The aim of this study was to investigate transforming growth factor-β-induced gene product-h3 (βig-h3) expression in dental pulp tissue and its effects on odontoblastic differentiation of dental pulp cells (DPCs). A rat direct pulp capping model was prepared using perforated rat upper first molars capped with mineral trioxide aggregate cement. Human DPCs (HDPCs) were isolated from extracted teeth. βig-h3 expression in rat dental pulp tissue and HDPCs was assessed by immunostaining. Mineralization of HDPCs was assessed by Alizarin red-S staining. Odontoblast-related gene expression in HDPCs was analyzed by quantitative RT-PCR. Expression of βig-h3 was detected in rat dental pulp tissue, and attenuated by direct pulp capping, while expression of interleukin-1β and tumor necrosis factor-α was increased in exposed pulp tissue. βig-h3 expression was also detected in HDPCs, with reduced expression during odontoblastic differentiation. The above cytokines reduced βig-h3 expression in HDPCs, and promoted their mineralization. Recombinant βig-h3 inhibited the expression of odontoblast-related genes and mineralization of HDPCs, while knockdown of βig-h3 gene expression promoted the expression of odontoblast-related genes in HDPCs. The present findings suggest that βig-h3 in DPCs may be involved in reparative dentin formation and that its expression is likely to negatively regulate this process. Copyright © 2017 Elsevier Ltd. All rights reserved.

Structure and thermodynamics of effector molecule binding to the nitrogen signal transduction PII protein GlnZ from Azospirillum brasilense.

Science.gov (United States)

Truan, Daphné; Bjelić, Saša; Li, Xiao-Dan; Winkler, Fritz K

2014-07-29

The trimeric PII signal transduction proteins regulate the function of a variety of target proteins predominantly involved in nitrogen metabolism. ATP, ADP and 2-oxoglutarate (2-OG) are key effector molecules influencing PII binding to targets. Studies of PII proteins have established that the 20-residue T-loop plays a central role in effector sensing and target binding. However, the specific effects of effector binding on T-loop conformation have remained poorly documented. We present eight crystal structures of the Azospirillum brasilense PII protein GlnZ, six of which are cocrystallized and liganded with ADP or ATP. We find that interaction with the diphosphate moiety of bound ADP constrains the N-terminal part of the T-loop in a characteristic way that is maintained in ADP-promoted complexes with target proteins. In contrast, the interactions with the triphosphate moiety in ATP complexes are much more variable and no single predominant interaction mode is apparent except for the ternary MgATP/2-OG complex. These conclusions can be extended to most investigated PII proteins of the GlnB/GlnK subfamily. Unlike reported for other PII proteins, microcalorimetry reveals no cooperativity between the three binding sites of GlnZ trimers for any of the three effectors under carefully controlled experimental conditions. Copyright © 2014 Elsevier Ltd. All rights reserved.

Loss of wobble uridine modification in tRNA anticodons interferes with TOR pathway signaling

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Viktor Scheidt

2014-11-01

Full Text Available Previous work in yeast has suggested that modification of tRNAs, in particular uridine bases in the anticodon wobble position (U34, is linked to TOR (target of rapamycin signaling. Hence, U34 modification mutants were found to be hypersensitive to TOR inhibition by rapamycin. To study whether this involves inappropriate TOR signaling, we examined interaction between mutations in TOR pathway genes (tip41∆, sap190∆, ppm1∆, rrd1∆ and U34 modification defects (elp3∆, kti12∆, urm1∆, ncs2∆ and found the rapamycin hypersensitivity in the latter is epistatic to drug resistance of the former. Epistasis, however, is abolished in tandem with a gln3∆ deletion, which inactivates transcription factor Gln3 required for TOR-sensitive activation of NCR (nitrogen catabolite repression genes. In line with nuclear import of Gln3 being under control of TOR and dephosphorylation by the Sit4 phosphatase, we identify novel TOR-sensitive sit4 mutations that confer rapamycin resistance and importantly, mislocalise Gln3 when TOR is inhibited. This is similar to gln3∆ cells, which abolish the rapamycin hypersensitivity of U34 modification mutants, and suggests TOR deregulation due to tRNA undermodification operates through Gln3. In line with this, loss of U34 modifications (elp3∆, urm1∆ enhances nuclear import of and NCR gene activation (MEP2, GAP1 by Gln3 when TOR activity is low. Strikingly, this stimulatory effect onto Gln3 is suppressed by overexpression of tRNAs that usually carry the U34 modifications. Collectively, our data suggest that proper TOR signaling requires intact tRNA modifications and that loss of U34 modifications impinges on the TOR-sensitive NCR branch via Gln3 misregulation.

Loss of wobble uridine modification in tRNA anticodons interferes with TOR pathway signaling.

Science.gov (United States)

Scheidt, Viktor; Jüdes, André; Bär, Christian; Klassen, Roland; Schaffrath, Raffael

2014-11-29

Previous work in yeast has suggested that modification of tRNAs, in particular uridine bases in the anticodon wobble position (U34), is linked to TOR (target of rapamycin) signaling. Hence, U34 modification mutants were found to be hypersensitive to TOR inhibition by rapamycin. To study whether this involves inappropriate TOR signaling, we examined interaction between mutations in TOR pathway genes ( tip41 ∆, sap190 ∆, ppm1 ∆, rrd1 ∆) and U34 modification defects ( elp3 ∆, kti 12∆, urm1 ∆, ncs2 ∆) and found the rapamycin hypersensitivity in the latter is epistatic to drug resistance of the former. Epistasis, however, is abolished in tandem with a gln3 ∆ deletion, which inactivates transcription factor Gln3 required for TOR-sensitive activation of NCR (nitrogen catabolite repression) genes. In line with nuclear import of Gln3 being under control of TOR and dephosphorylation by the Sit4 phosphatase, we identify novel TOR-sensitive sit4 mutations that confer rapamycin resistance and importantly, mislocalise Gln3 when TOR is inhibited. This is similar to gln3 ∆ cells, which abolish the rapamycin hypersensitivity of U34 modification mutants, and suggests TOR deregulation due to tRNA undermodification operates through Gln3. In line with this, loss of U34 modifications ( elp3 ∆, urm1 ∆) enhances nuclear import of and NCR gene activation ( MEP2 , GAP1 ) by Gln3 when TOR activity is low. Strikingly, this stimulatory effect onto Gln3 is suppressed by overexpression of tRNAs that usually carry the U34 modifications. Collectively, our data suggest that proper TOR signaling requires intact tRNA modifications and that loss of U34 modifications impinges on the TOR-sensitive NCR branch via Gln3 misregulation.

Metabolic engineering of the regulators in nitrogen catabolite repression to reduce the production of ethyl carbamate in a model rice wine system.

Science.gov (United States)

Zhao, Xinrui; Zou, Huijun; Fu, Jianwei; Zhou, Jingwen; Du, Guocheng; Chen, Jian

2014-01-01

Rice wine has been one of the most popular traditional alcoholic drinks in China. However, the presence of potentially carcinogenic ethyl carbamate (EC) in rice wine has raised a series of food safety issues. During rice wine production, the key reason for EC formation is urea accumulation, which occurs because of nitrogen catabolite repression (NCR) in Saccharomyces cerevisiae. NCR represses urea utilization by retaining Gln3p in the cytoplasm when preferred nitrogen sources are present. In order to increase the nuclear localization of Gln3p, some possible phosphorylation sites on the nuclear localization signal were mutated and the nuclear localization regulation signal was truncated, and the disruption of URE2 provided an additional method of reducing urea accumulation. By combining these strategies, the genes involved in urea utilization (DUR1,2 and DUR3) could be significantly activated in the presence of glutamine. During shake flask fermentations of the genetically modified strains, very little urea accumulated in the medium. Furthermore, the concentrations of urea and EC were reduced by 63% and 72%, respectively, in a model rice wine system. Examination of the normal nutrients in rice wine indicated that there were few differences in fermentation characteristics between the wild-type strain and the genetically modified strain. These results show that metabolic engineering of the NCR regulators has great potential as a method for eliminating EC during rice wine production.

Recombinant Brucella abortus gene expressing immunogenic protein

Energy Technology Data Exchange (ETDEWEB)

Mayfield, J.E.; Tabatabai, L.B.

1991-06-11

This patent describes a synthetic recombinant DNA molecule containing a DNA sequence. It comprises a gene of Brucella abortus encoding an immunogenic protein having a molecular weight of approximately 31,000 daltons as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis under denaturing conditions, the protein having an isoelectric point around 4.9, and containing a twenty-five amino acid sequence from its amino terminal end consisting of Gln-Ala-Pro-Thr-Phe-Phe-Arg-Ile-Gly-Thr-Gly-Gly-Thr-Ala-Gly-Thr-Tyr-Tyr-Pro-Ile-Gly-Gly-Leu-Ile-Ala, wherein Gln, Ala, Pro, Thr, Phe, Arg, Ile, Gly, Tyr, and Leu, respectively, represent glutamine, alanine, proline, threonine, phenylalanine, arginine, isolecuine, glycine, tyrosine, and leucine.

Toeplitz quantization of Kaehler manifolds and gl(N), N [yields] [infinity] limits

Energy Technology Data Exchange (ETDEWEB)

Bordemann, M. (Dept. of Physics, Univ. of Freiburg (Germany)); Meinrenken, E. (Dept. of Mathematics, M.I.T., Cambridge, MA (United States)); Schlichenmaier, M. (Dept. of Mathematics and Computer Science, Univ. of Mannheim (Germany))

1994-10-01

For general compact Kaehler manifolds it is shown that both Toeplitz quantization and geometric quantization lead to a well-defined (by operator norm estimates) classical limit. This generalizes earlier results of the authors and Klimek and Lesniewski obtained for the torus and higher genus Riemann surfaces, respectively. We thereby arrive at an approximation of the Poisson algebra by a sequence of finite-dimensional matrix algebras gl(N), N [yields] [infinity]. (orig.)

hPEPT1 Affinity and Translocation of Selected Gln-Sar and Glu-Sar Dipeptide Derivatives

DEFF Research Database (Denmark)

Eriksson, A. H.; Elm, Peter L.; Begtrup, Mikael

2005-01-01

using 14C-labeled Gly-Sar. Translocation was measured as fluorescence ratios induced by the substrates using the fluorescent probe BCECF and an epifluorescence microscope setup. All compounds showed high affinity to hPEPT1, but only the amides l-Gln(N,N-dimethyl)-Sar and l-Gln(N-piperidinyl)-Sar were...... been suggested. However, these are not necessarily predictive of compounds that are actually translocated by hPEPT1. More information on affinity to and translocation via hPEPT1 of side-chain-modified dipeptides may be gained by conducting a study of selected dipeptide derivatives with variety in size...... translocated by hPEPT1. hPEPT1 is very susceptible to modifications of the N-terminal amino acid side chain of dipeptidomimetic substrates, in terms of achieving compounds with high affinity for the transporter. However, as affinity is not predictive of translocation, derivatization in this position must...

Clinical and Prognostic Profiles of Cardiomyopathies Caused by Mutations in the Troponin T Gene.

Science.gov (United States)

Ripoll-Vera, Tomás; Gámez, José María; Govea, Nancy; Gómez, Yolanda; Núñez, Juana; Socías, Lorenzo; Escandell, Ángela; Rosell, Jorge

2016-02-01

Mutations in the troponin T gene (TTNT2) have been associated in small studies with the development of hypertrophic cardiomyopathy characterized by a high risk of sudden death and mild hypertrophy. We describe the clinical course of patients carrying mutations in this gene. We analyzed the clinical characteristics and prognosis of patients with mutations in the TNNT2 gene who were seen in an inherited cardiac disease unit. Of 180 families with genetically studied cardiomyopathies, 21 families (11.7%) were identified as having mutations in TNNT2: 10 families had Arg92Gln, 5 had Arg286His, 3 had Arg278Cys, 1 had Arg92Trp, 1 had Arg94His, and 1 had Ile221Thr. Thirty-three additional genetic carriers were identified through family assessment. The study included 54 genetic carriers: 56% were male, and the mean average age was 41 ± 17 years. There were 33 cases of hypertrophic cardiomyopathy, 9 of dilated cardiomyopathy, and 1 of noncompaction cardiomyopathy, and maximal myocardial thickness was 18.5 ± 6mm. Ventricular dysfunction was present in 30% of individuals and a history of sudden death in 62%. During follow-up, 4 patients died and 14 (33%) received a defibrillator (8 probands, 6 relatives). Mean survival was 54 years. Carriers of Arg92Gln had early disease development, high penetrance, a high risk of sudden death, a high rate of defibrillator implantation, and a high frequency of mixed phenotype. Mutations in the TNNT2 gene were more common in this series than in previous studies. The clinical and prognostic profiles depended on the mutation present. Carriers of the Arg92Gln mutation developed hypertrophic or dilated cardiomyopathy and had a significantly worse prognosis than those with other mutations in TNNT2 or other sarcomeric genes. Copyright © 2015 Sociedad Española de Cardiología. Published by Elsevier España, S.L.U. All rights reserved.

Transcriptional regulation of genes related to progesterone production.

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Mizutani, Tetsuya; Ishikane, Shin; Kawabe, Shinya; Umezawa, Akihiro; Miyamoto, Kaoru

2015-01-01

Steroid hormones are synthesized from cholesterol in various tissues, mainly in the adrenal glands and gonads. Because these lipid-soluble steroid hormones immediately diffuse through the cells in which they are produced, their secretion directly reflects the activity of the genes related to their production. Progesterone is important not only for luteinization and maintenance of pregnancy, but also as a substrate for most other steroids. Steroidogenic acute regulatory protein (STAR), cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc), and 3β-hydroxysteroid dehydrogenase/Δ(5)-Δ(4) isomerase (3β-HSD) are well-known proteins essential for progesterone production. In addition to them, glutathione S-transferase A1-1 and A3-3 are shown to exert Δ(5)-Δ(4) isomerization activity to produce progesterone in a cooperative fashion with 3β-HSD. 5-Aminolevulinic acid synthase 1, ferredoxin 1, and ferredoxin reductase also play a role in steroidogenesis as accessory factors. Members of the nuclear receptor 5A (NR5A) family (steroidogenic factor 1 and liver receptor homolog 1) play a crucial role in the transcriptional regulation of these genes. The NR5A family activates these genes by binding to NR5A responsive elements present within their promoter regions, as well as to the elements far from their promoters. In addition, various NR5A-interacting proteins including peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α), nuclear receptor subfamily 0, group B, member 1 (DAX-1), and CCAAT/enhancer-binding proteins (C/EBP) are involved in the transcription of NR5A target genes and regulate the transcription either positively or negatively under both basal and tropic hormone-stimulated conditions. In this review, we describe the transcriptional regulation of genes related to progesterone production.

Deletion of genes involved in glutamate metabolism to improve poly-gamma-glutamic acid production in B. amyloliquefaciens LL3.

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Zhang, Wei; He, Yulian; Gao, Weixia; Feng, Jun; Cao, Mingfeng; Yang, Chao; Song, Cunjiang; Wang, Shufang

2015-02-01

Here, we attempted to elevate poly-gamma-glutamic acid (γ-PGA) production by modifying genes involved in glutamate metabolism in Bacillus amyloliquefaciens LL3. Products of rocR, rocG and gudB facilitate the conversion from glutamate to 2-oxoglutarate in Bacillus subtillis. The gene odhA is responsible for the synthesis of a component of the 2-oxoglutarate dehydrogenase complex that catalyzes the oxidative decarboxylation of 2-oxoglutarate to succinyl coenzyme A. In-frame deletions of these four genes were performed. In shake flask experiments the gudB/rocG double mutant presented enhanced production of γ-PGA, a 38 % increase compared with wild type. When fermented in a 5-L fermenter with pH control, the γ-PGA yield of the rocR mutant was increased to 5.83 g/L from 4.55 g/L for shake flask experiments. The gudB/rocG double mutant produced 5.68 g/L γ-PGA compared with that of 4.03 g/L for the wild type, a 40 % increase. Those results indicated the possibility of improving γ-PGA production by modifying glutamate metabolism, and identified potential genetic targets to improve γ-PGA production.

Metabolic Engineering of the Regulators in Nitrogen Catabolite Repression To Reduce the Production of Ethyl Carbamate in a Model Rice Wine System

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Zhao, Xinrui; Zou, Huijun; Fu, Jianwei; Chen, Jian

2014-01-01

Rice wine has been one of the most popular traditional alcoholic drinks in China. However, the presence of potentially carcinogenic ethyl carbamate (EC) in rice wine has raised a series of food safety issues. During rice wine production, the key reason for EC formation is urea accumulation, which occurs because of nitrogen catabolite repression (NCR) in Saccharomyces cerevisiae. NCR represses urea utilization by retaining Gln3p in the cytoplasm when preferred nitrogen sources are present. In order to increase the nuclear localization of Gln3p, some possible phosphorylation sites on the nuclear localization signal were mutated and the nuclear localization regulation signal was truncated, and the disruption of URE2 provided an additional method of reducing urea accumulation. By combining these strategies, the genes involved in urea utilization (DUR1,2 and DUR3) could be significantly activated in the presence of glutamine. During shake flask fermentations of the genetically modified strains, very little urea accumulated in the medium. Furthermore, the concentrations of urea and EC were reduced by 63% and 72%, respectively, in a model rice wine system. Examination of the normal nutrients in rice wine indicated that there were few differences in fermentation characteristics between the wild-type strain and the genetically modified strain. These results show that metabolic engineering of the NCR regulators has great potential as a method for eliminating EC during rice wine production. PMID:24185848

The BRCA1 c. 5096G>A p.Arg1699Gln (R1699Q) intermediate risk variant

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Moghadasi, Setareh; Meeks, Huong D.; Vreeswijk, Maaike Pg

2018-01-01

studied families, to further define cancer risks and to propose adjusted clinical management of female BRCA1*R1699Q carriers. METHODS: Data were collected from 129 BRCA1*R1699Q families ascertained internationally by ENIGMA (Evidence-based Network for the Interpretation of Germline Mutant Alleles......BACKGROUND: We previously showed that the BRCA1 variant c.5096G>A p.Arg1699Gln (R1699Q) was associated with an intermediate risk of breast cancer (BC) and ovarian cancer (OC). This study aimed to assess these cancer risks for R1699Q carriers in a larger cohort, including follow-up of previously......) consortium members. A modified segregation analysis was used to calculate BC and OC risks. Relative risks were calculated under both monogenic model and major gene plus polygenic model assumptions. RESULTS: In this cohort the cumulative risk of BC and OC by age 70 years was 20% and 6%, respectively...

Uptake and metabolism of L-[3H]glutamate and L-[3H]glutamine in adult rat cerebellar slices

International Nuclear Information System (INIS)

de Barry, J.; Vincendon, G.; Gombos, G.

1983-01-01

Using very low concentrations (1 mumol range) of L-2-3-[ 3 H]glutamate, ( 3 H-Glu) or L-2-3-[ 3 H]glutamine ( 3 H-Gln), the authors have previously shown by autoradiography that these amino acids were preferentially taken up in the molecular layer of the cerebellar cortex. Furthermore, the accumulation of 3 H-Glu was essentially glial in these conditions. Uptake and metabolism of either ( 3 H-Glu) or ( 3 H-Gln) were studied in adult rat cerebellar slices. Both amino acids were rapidly converted into other metabolic compounds: after seven minutes of incubation in the presence of exogenous 3 H-Glu, 70% of the tissue accumulated radioactivity was found to be in compounds other than glutamate. The main metabolites were Gln (42%), alpha-ketoglutarate (25%) and GABA (1,4%). In the presence of exogenous 3 H-Gln the rate of metabolism was slightly slower (50% after seven minutes of incubation) and the metabolites were also Glu (29%), alpha-ketoglutarate (15%) and GABA (5%). Using depolarizing conditions (56 mM KCl) with either exogenous 3 H-Glu or 3 H-Gln, the radioactivity was preferentially accumulated in glutamate compared to control. From these results we conclude: i) there are two cellular compartments for the neurotransmission-glutamate-glutamine cycle; one is glial, the other neuronal; ii) these two cellular compartments contain both Gln and Glu; iii) transmitter glutamate is always in equilibrium with the so-called ''metabolic'' pool of glutamate; iv) the regulation of the glutamate-glutamine cycle occurs at least at two different levels: the uptake of glutamate and the enzymatic activity of the neuronal glutaminase

Allele and Genotype Distributions of DNA Repair Gene Polymorphisms in South Indian Healthy Population

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Katiboina Srinivasa Rao

2014-01-01

Full Text Available Various DNA repair pathways protect the structural and chemical integrity of the human genome from environmental and endogenous threats. Polymorphisms of genes encoding the proteins involved in DNA repair have been found to be associated with cancer risk and chemotherapeutic response. In this study, we aim to establish the normative frequencies of DNA repair genes in South Indian healthy population and compare with HapMap populations. Genotyping was done on 128 healthy volunteers from South India, and the allele and genotype distributions were established. The minor allele frequency of Xeroderma pigmentosum group A ( XPA G23A, Excision repair cross-complementing 2 ( ERCC2 /Xeroderma pigmentosum group D ( XPD Lys751Gln, Xeroderma pigmentosum group G ( XPG His46His, XPG Asp1104His, and X-ray repair cross-complementing group 1 ( XRCC1 Arg399Gln polymorphisms were 49.2%, 36.3%, 48.0%, 23.0%, and 34.0% respectively. Ethnic variations were observed in the frequency distribution of these polymorphisms between the South Indians and other HapMap populations. The present work forms the groundwork for cancer association studies and biomarker identification for treatment response and prognosis.

Lichenysin production is improved in codY null Bacillus licheniformis by addition of precursor amino acids.

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Zhu, Chengjun; Xiao, Fang; Qiu, Yimin; Wang, Qin; He, Zhili; Chen, Shouwen

2017-08-01

Lichenysin is categorized into the family of lipopeptide biosurfactants and has a variety of applications in the petroleum industry, bioremediation, pharmaceuticals, and the food industry. Currently, large-scale production is limited due to the low yield. This study found that lichenysin production was repressed by supplementation of extracellular amino acids. The global transcriptional factor CodY was hypothesized to prevent lichenysin biosynthesis under an amino acid-rich condition in Bacillus licheniformis. Thus, the codY null strain was constructed, and lichenysin production was increased by 31.0% to 2356 mg/L with the addition of precursor amino acids, and the lichenysin production efficiency was improved by 42.8% to 98.2 mg/L• h. Correspondingly, the transcription levels of the lichenysin synthetase gene lchAA, and its corresponding regulator genes comA, degQ, and degU, were upregulated. Also, the codY deletion enhanced biosynthesis of lichenysin precursor amino acids (Gln, Ile, Leu, and Val) and reduced the formation of byproducts, acetate, acetoin, and 2,3-butanediol. This study firstly reported that lichenysin biosynthesis was negatively regulated by CodY and lichenysin production could be further improved with the precursor amino acid amendment in the codY null strain.

A Study on Enhanced Expression of 3-Hydroxypropionic Acid Pathway Genes and Impact on Its Production in Lactobacillus reuteri

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Gopal Ramakrishnan Gopi

2015-01-01

Full Text Available 3-Hydroxypropionic acid (3-HP is a novel antimicrobial agent against foodborne pathogens like Salmonella and Staphylococcus species. Lactobacillus reuteri converts glycerol into 3-HP using a coenzyme A-dependent pathway, which is encoded by propanediol utilization operon (pdu subjected to catabolite repression. In a catabolite repression-deregulated L. reuteri RPRB3007, quantitative PCR revealed a 2.5-fold increase in the transcripts of the genes pduP, pduW and pduL during the mid-log phase of growth. The production of 3-HP was tested in resting cells in phosphate buff er and growing batch cultures in MRS broth of various glucose/glycerol ratios. Due to the upregulation of pathway genes, specific formation rate of 3-HP in the mutant strain was found to be enhanced from 0.167 to 0.257 g per g of cell dry mass per h. Furthermore, formation of 3-HP in resting cells was limited due to the substrate inhibition by reuterin at a concentration of (30±5 mM. In batch cultures, the formation of 3-HP was not observed during the logarithmic and stationary phases of growth of wild-type and mutant strains, which was confi rmed by NMR spectroscopy. However, the cells collected in these phases were found to produce 3-HP aft er washing and converting them to resting cells. Lactate and acetate, the primary end products of glucose catabolism, might be the inhibiting elements for 3-HP formation in batch cultures. This was confirmed when lactate (25±5 mM or acetate (20±5 mM were added to biotransformation medium, which prevented the 3-HP formation. Moreover, the removal of sodium acetate and glucose (carbon source for lactic acid production was found to restore 3-HP formation in the MRS broth in a similar manner to that of the phosphate buff er. Even though the genetic repression was circumvented by the up-regulation of pathway genes using a mutant strain, 3-HP formation was further limited by the substrate and catabolite inhibition.

Transcriptome of Proteus mirabilis in the Murine Urinary Tract: Virulence and Nitrogen Assimilation Gene Expression▿†

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Pearson, Melanie M.; Yep, Alejandra; Smith, Sara N.; Mobley, Harry L. T.

2011-01-01

The enteric bacterium Proteus mirabilis is a common cause of complicated urinary tract infections. In this study, microarrays were used to analyze P. mirabilis gene expression in vivo from experimentally infected mice. Urine was collected at 1, 3, and 7 days postinfection, and RNA was isolated from bacteria in the urine for transcriptional analysis. Across nine microarrays, 471 genes were upregulated and 82 were downregulated in vivo compared to in vitro broth culture. Genes upregulated in vivo encoded mannose-resistant Proteus-like (MR/P) fimbriae, urease, iron uptake systems, amino acid and peptide transporters, pyruvate metabolism enzymes, and a portion of the tricarboxylic acid (TCA) cycle enzymes. Flagella were downregulated. Ammonia assimilation gene glnA (glutamine synthetase) was repressed in vivo, while gdhA (glutamate dehydrogenase) was upregulated in vivo. Contrary to our expectations, ammonia availability due to urease activity in P. mirabilis did not drive this gene expression. A gdhA mutant was growth deficient in minimal medium with citrate as the sole carbon source, and loss of gdhA resulted in a significant fitness defect in the mouse model of urinary tract infection. Unlike Escherichia coli, which represses gdhA and upregulates glnA in vivo and cannot utilize citrate, the data suggest that P. mirabilis uses glutamate dehydrogenase to monitor carbon-nitrogen balance, and this ability contributes to the pathogenic potential of P. mirabilis in the urinary tract. PMID:21505083

Polymorphisms of the ghrelin/obestatin gene and ghrelin levels in Chinese children with short stature.

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Zou, Chao Chun; Huang, Ke; Liang, Li; Zhao, Zheng Yan

2008-07-01

To investigate the role of ghrelin and polymorphisms of ghrelin/obestatin gene in children with short stature. A total of 117 GH deficient (GHD) and 81 idiopathic short stature (ISS) children were studied. The controls consisted of 125 age and gender-matched healthy children. The Arg51Gln, Leu72Met and Gln90Leu polymorphisms were genotyped using MassArray and total plasma ghrelin was measured by radioimmunoassay. In this study, the frequency of the Arg51Gln polymorphism was very low (0% in controls and 1.0% in patients). The frequency of the Gln90Leu polymorphism was 1.6% in controls and 0.5% in patients, respectively. Higher frequencies of Leu72Met (34.4% in controls and 39.9% in patients) and Met72Met genotypes (4.0% in controls and 2.0% in patients) were found. The differences in the Arg51Gln, Leu72Met or Gln90Leu genotypes and allele frequencies between patients and controls were not significant. Also, there were no significant differences in the Leu72Met genotypes and allele frequencies between GHD and ISS subgroups. There were no significant differences in clinical characteristics and biochemistry markers (including ghrelin levels) among the different genotypes of Leu72Met. However, plasma ghrelin levels in the GHD group were significantly lower than those of controls (P = 0.001). These results suggest that ghrelin may have a role in GH secretion and controlling growth. Lower ghrelin levels, but not ghrelin/obestatin polymorphism, might contribute to GHD.

Study of oleanolic acid on the estrodiol production and the fat production of mouse preadipocyte 3T3-L1 in vitro.

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Wan, Qian; Lu, Hua; Liu, Xia; Yie, Shangmian; Xiang, Junbei; Yao, Zouying

2015-01-01

The women during the menopause period have an increased tendency for the obesity, which represents the more fat production than during the premenopausal period. Although this is not beneficial overall, it could provide a compensatory source for the estrogen production for the menopausal women. So it would be meaningful to find an agent that could inhibit the fat production while does not disturb the total estrogen production by fat tissues. In the present study, the effect of oleanolic acid (OA) on the fat production and the total estrogen production of the differentiating mouse preadipocyte 3T3-L1 as well as the mechanisms behind those effects were preliminarily investigated. The cell line 3T3-L1 was chosen as the model cell because it is usually used for the research about the obesity. During the induced differentiation of 3T3-L1 cells, cells were intervened continuously with OA. The fat production was determined with the oil red staining assay and the total estrogen production was measured with the ELISA assay. Finally, the expression patterns for important genes of the fat production and the estrogen production were studied, respectively with the real-time fluorescence quantitative PCR (qPCR). The results showed that for the differentiating 3T3-L1 cells, OA could significantly inhibit the fat production and did not disturb the total estrogen production significantly. In the mechanism studies, OA was found to significantly down-regulate ACC, the key gene for fat synthesis, which could explain the inhibitory effect of OA on the fat production; OA was also found to significantly up-regulate CYP11A1, CYP17, CYP19, the key genes for the estrogen synthesis and significantly down-regulate CYP1A1, the key gene for the estrogen decomposition, which preliminarily explained the lack of the effect of OA on the total estrogen production. In conclusion, OA was found able to inhibit the fat production while maintaining the total estrogen level and the mechanisms for the above

Correlating Information Contents of Gene Ontology Terms to Infer Semantic Similarity of Gene Products

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Mingxin Gan

2014-01-01

Full Text Available Successful applications of the gene ontology to the inference of functional relationships between gene products in recent years have raised the need for computational methods to automatically calculate semantic similarity between gene products based on semantic similarity of gene ontology terms. Nevertheless, existing methods, though having been widely used in a variety of applications, may significantly overestimate semantic similarity between genes that are actually not functionally related, thereby yielding misleading results in applications. To overcome this limitation, we propose to represent a gene product as a vector that is composed of information contents of gene ontology terms annotated for the gene product, and we suggest calculating similarity between two gene products as the relatedness of their corresponding vectors using three measures: Pearson’s correlation coefficient, cosine similarity, and the Jaccard index. We focus on the biological process domain of the gene ontology and annotations of yeast proteins to study the effectiveness of the proposed measures. Results show that semantic similarity scores calculated using the proposed measures are more consistent with known biological knowledge than those derived using a list of existing methods, suggesting the effectiveness of our method in characterizing functional relationships between gene products.

Leu72Met408 Polymorphism of the Ghrelin Gene Is Associated With Early Phase of Gastric Emptying in the Patients With Functional Dyspepsia in Japan.

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Yamawaki, Hiroshi; Futagami, Seiji; Shimpuku, Mayumi; Shindo, Tomotaka; Maruki, Yuuta; Nagoya, Hiroyuki; Kodaka, Yasuhiro; Sato, Hitomi; Gudis, Katya; Kawagoe, Tetsuro; Sakamoto, Choitsu

2015-01-01

There are no available data about the relationship between ghrelin gene genotypes and early phase of gastric emptying in functional dyspepsia (FD) as defined by Rome III classification. We enrolled 74 patients presenting with typical symptoms of FD and 64 healthy volunteers. Gastric motility was evaluated using the 13C-acetate breath test. We used Rome III criteria to evaluate upper abdominal symptoms and self-rating questionnaires for depression (SRQ-D) scores to determine status of depression. The Arg51Gln (346G->A), preproghrelin (3056T->C), Leu72Met (408C->A), Gln90Leu (3412T->A) and G-protein 3 (825C->T) polymorphisms were analyzed in the DNA from blood samples of enrolled subjects. Genotyping was performed by polymerase chain reaction. There was a significant relationship between the Gln90Leu3412 genotype and SRQ-D score in FD patients (P = 0.009). Area under the curve at 15 minutes (AUC15) value was significantly associated with the Leu72Met408 genotype (P = 0.015) but not with entire gastric emptying. The Leu72Met (408C->A) single nucleotide polymorphism was significantly associated with early phase of gastric emptying in FD patients. Further studies will be necessary to clarify the association between ghrelin gene single nucleotide polymorphisms and early phase of gastric emptying in FD patients.

Milder clinical and biochemical phenotypes associated with the c.482G>A (p.Arg161Gln) pathogenic variant in cobalamin C disease: Implications for management and screening.

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Almannai, Mohammed; Marom, Ronit; Divin, Kristian; Scaglia, Fernando; Sutton, V Reid; Craigen, William J; Lee, Brendan; Burrage, Lindsay C; Graham, Brett H

2017-09-01

Cobalamin C disease is a multisystemic disease with variable manifestations and age of onset. Genotype-phenotype correlations are well-recognized in this disorder. Here, we present a large cohort of individuals with cobalamin C disease, several of whom are heterozygous for the c.482G>A pathogenic variant (p.Arg161Gln). We compared clinical characteristics of individuals with this pathogenic variant to those who do not have this variant. To our knowledge, this study represents the largest single cohort of individuals with the c.482G>A (p.Arg161Gln) pathogenic variant. A retrospective chart review of 27 individuals from 21 families with cobalamin C disease who are followed at our facility was conducted. 13 individuals (48%) are compound heterozygous with the c.482G>A (p.Arg161Gln) on one allele and a second pathogenic variant on the other allele. Individuals with the c.482G>A (p.Arg161Gln) pathogenic variant had later onset of symptoms and easier metabolic control. Moreover, they had milder biochemical abnormalities at presentation which likely contributed to the observation that 4 individuals (31%) in this group were missed by newborn screening. The c.482G>A (p.Arg161Gln) pathogenic variant is associated with milder disease. These individuals may not receive a timely diagnosis as they may not be identified on newborn screening or because of unrecognized, late onset symptoms. Despite the milder presentation, significant complications can occur, especially if treatment is delayed. Copyright © 2017 Elsevier Inc. All rights reserved.

Role of transcription factor KLF11 and its diabetes-associated gene variants in pancreatic beta cell function

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Neve, Bernadette; Fernandez-Zapico, Martin E; Ashkenazi-Katalan, Vered

2005-01-01

in beta cells. Genetic analysis of the KLF11 gene revealed two rare variants (Ala347Ser and Thr220Met) that segregate with diabetes in families with early-onset type 2 diabetes, and significantly impair its transcriptional activity. In addition, analysis of 1,696 type 2 diabetes mellitus and 1......,776 normoglycemic subjects show a frequent polymorphic Gln62Arg variant that significantly associates with type 2 diabetes mellitus in North European populations (OR = 1.29, P = 0.00033). Moreover, this variant alters the corepressor mSin3A-binding activity of KLF11, impairs the activation of the insulin promoter...... and shows lower levels of insulin expression in pancreatic beta cells. In addition, subjects carrying the Gln62Arg allele show decreased plasma insulin after an oral glucose challenge. Interestingly, all three nonsynonymous KLF11 variants show increased repression of the catalase 1 promoter, suggesting...

Ammonia production from amino acid-based biomass-like sources by engineered Escherichia coli.

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Mikami, Yosuke; Yoneda, Hisanari; Tatsukami, Yohei; Aoki, Wataru; Ueda, Mitsuyoshi

2017-12-01

The demand for ammonia is expected to increase in the future because of its importance in agriculture, industry, and hydrogen transportation. Although the Haber-Bosch process is known as an effective way to produce ammonia, the process is energy-intensive. Thus, an environmentally friendly ammonia production process is desired. In this study, we aimed to produce ammonia from amino acids and amino acid-based biomass-like resources by modifying the metabolism of Escherichia coli. By engineering metabolic flux to promote ammonia production using the overexpression of the ketoisovalerate decarboxylase gene (kivd), derived from Lactococcus lactis, ammonia production from amino acids was 351 mg/L (36.6% yield). Furthermore, we deleted the glnA gene, responsible for ammonia assimilation. Using yeast extract as the sole source of carbon and nitrogen, the resultant strain produced 458 mg/L of ammonia (47.8% yield) from an amino acid-based biomass-like material. The ammonia production yields obtained are the highest reported to date. This study suggests that it will be possible to produce ammonia from waste biomass in an environmentally friendly process.

Aminoacyl-tRNA quality control is required for efficient activation of the TOR pathway regulator Gln3p.

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Mohler, Kyle; Mann, Rebecca; Kyle, Amanda; Reynolds, Noah; Ibba, Michael

2017-09-14

The aminoacylation status of the cellular tRNA pool regulates both general amino acid control (GAAC) and target of rapamycin (TOR) stress response pathways in yeast. Consequently, fidelity of translation at the level of aminoacyl-tRNA synthesis plays a central role in determining accuracy and sensitivity of stress responses. To investigate effects of translational quality control (QC) on cell physiology under stress conditions, phenotypic microarray analyses were used to identify changes in QC deficient cells. Nitrogen source growth assays showed QC deficient yeast grew differently compared to WT. The QC deficient strain was more tolerant to caffeine treatment than wild type through altered interactions with the TOR and GAAC pathways. Increased caffeine tolerance of the QC deficient strain was consistent with the observation that the activity of Gln3p, a transcription factor controlled by the TOR pathway, is decreased in the QC deficient strain compared to WT. GCN4 translation, which is typically repressed in the absence of nutritional stress, was enhanced in the QC deficient strain through TOR inhibition. QC did not impact cell cycle regulation; however, the chronological lifespan of QC deficient yeast strains decreased compared to wild type, likely due to translational errors and alteration of the TOR-associated regulon. These findings support the idea that changes in translational fidelity provide a mechanism of cellular adaptation by modulating TOR activity. This, in turn, supports a central role for aminoacyl-tRNA synthesis QC in the integrated stress response by maintaining the proper aa-tRNA pools necessary to coordinate the GAAC and TOR.

Radiation-induced damage to normal tissues after radiotherapy in patients treated for gynecologic tumors: Association with single nucleotide polymorphisms in XRCC1, XRCC3, and OGG1 genes and in vitro chromosomal radiosensitivity in lymphocytes

International Nuclear Information System (INIS)

Ruyck, Kim de; Eijkeren, Marc van; Claes, Kathleen; Morthier, Rudy; Paepe, Anne de; Vral, Anne; Ridder, Leo de; Thierens, Hubert

2005-01-01

Purpose: To examine the association of polymorphisms in XRCC1 (194Arg/Trp, 280Arg/His, 399Arg/Gln, 632Gln/Gln), XRCC3 (5' UTR 4.541A>G, IVS5-14 17.893A>G, 241Thr/Met), and OGG1 (326Ser/Cys) with the development of late radiotherapy (RT) reactions and to assess the correlation between in vitro chromosomal radiosensitivity and clinical radiosensitivity. Methods and Materials: Sixty-two women with cervical or endometrial cancer treated with RT were included in the study. According to the Common Terminology Criteria for Adverse Events, version 3.0, scale, 22 patients showed late adverse RT reactions. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assays were performed to examine polymorphic sites, the G2 assay was used to measure chromosomal radiosensitivity, and patient groups were compared using actuarial methods. Results: The XRCC3 IVS5-14 polymorphic allele was significantly associated with the risk of developing late RT reactions (odds ratio 3.98, p = 0.025), and the XRCC1 codon 194 variant showed a significant protective effect (p = 0.028). Patients with three or more risk alleles in XRCC1 and XRCC3 had a significantly increased risk of developing normal tissue reactions (odds ratio 10.10, p = 0.001). The mean number of chromatid breaks per cell was significantly greater in patients with normal tissue reactions than in patients with no reactions (1.16 and 1.34, respectively; p = 0.002). Patients with high chromosomal radiosensitivity showed a 9.2-fold greater annual risk of complications than patients with intermediate chromosomal radiosensitivity. Combining the G2 analysis with the risk allele model allowed us to identify 23% of the patients with late normal tissue reactions, without false-positive results. Conclusion: The results of the present study showed that clinical radiosensitivity is associated with an enhanced G2 chromosomal radiosensitivity and is significantly associated with a combination of different polymorphisms in

Electrostatic effects in the folding of the SH3 domain of the c-Src tyrosine kinase: pH-dependence in 3D-domain swapping and amyloid formation.

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Julio Bacarizo

Full Text Available The SH3 domain of the c-Src tyrosine kinase (c-Src-SH3 aggregates to form intertwined dimers and amyloid fibrils at mild acid pHs. In this work, we show that a single mutation of residue Gln128 of this SH3 domain has a significant effect on: (i its thermal stability; and (ii its propensity to form amyloid fibrils. The Gln128Glu mutant forms amyloid fibrils at neutral pH but not at mild acid pH, while Gln128Lys and Gln128Arg mutants do not form these aggregates under any of the conditions assayed. We have also solved the crystallographic structures of the wild-type (WT and Gln128Glu, Gln128Lys and Gln128Arg mutants from crystals obtained at different pHs. At pH 5.0, crystals belong to the hexagonal space group P6₅22 and the asymmetric unit is formed by one chain of the protomer of the c-Src-SH3 domain in an open conformation. At pH 7.0, crystals belong to the orthorhombic space group P2₁2₁2₁, with two molecules at the asymmetric unit showing the characteristic fold of the SH3 domain. Analysis of these crystallographic structures shows that the residue at position 128 is connected to Glu106 at the diverging β-turn through a cluster of water molecules. Changes in this hydrogen-bond network lead to the displacement of the c-Src-SH3 distal loop, resulting also in conformational changes of Leu100 that might be related to the binding of proline rich motifs. Our findings show that electrostatic interactions and solvation of residues close to the folding nucleation site of the c-Src-SH3 domain might play an important role during the folding reaction and the amyloid fibril formation.

Molecular evidence for the coordination of nitrogen and carbon metabolisms, revealed by a study on the transcriptional regulation of the agl3EFG operon that encodes a putative carbohydrate transporter in Streptomyces coelicolor.

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Cen, Xu-Feng; Wang, Jing-Zhi; Zhao, Guo-Ping; Wang, Ying; Wang, Jin

2016-03-18

In the agl3EFGXYZ operon (SCO7167-SCO7162, abbreviated as agl3 operon) of Streptomyces coelicolor M145, agl3EFG genes encode a putative ABC-type carbohydrate transporter. The transcription of this operon has been proved to be repressed by Agl3R (SCO7168), a neighboring GntR-family regulator, and this repression can be released by growth on poor carbon sources. Here in this study, we prove that the transcription of agl3 operon is also directly repressed by GlnR, a central regulator governing the nitrogen metabolism in S. coelicolor. The electrophoretic mobility shift assay (EMSA) employing the agl3 promoter and mixtures of purified recombinant GlnR and Agl3R indicates that GlnR and Agl3R bind to different DNA sequences within the promoter region of agl3 operon, which is further confirmed by the DNase I footprinting assay. As Agl3R and GlnR have been demonstrated to sense the extracellular carbon and nitrogen supplies, respectively, it is hypothesized that the transcription of agl3 operon is stringently governed by the availabilities of extracellular carbon and nitrogen sources. Consistent with the hypothesis, the agl3 operon is further found to be derepressed only under the condition of poor carbon and rich nitrogen supplies, when both regulators are inactivated. It is believed that activation of the expression of agl3 operon may facilitate the absorption of extracellular carbohydrates to balance the ratio of intracellular carbon to nitrogen. Copyright © 2016 Elsevier Inc. All rights reserved.

A new measure for functional similarity of gene products based on Gene Ontology

Directory of Open Access Journals (Sweden)

Lengauer Thomas

2006-06-01

Full Text Available Abstract Background Gene Ontology (GO is a standard vocabulary of functional terms and allows for coherent annotation of gene products. These annotations provide a basis for new methods that compare gene products regarding their molecular function and biological role. Results We present a new method for comparing sets of GO terms and for assessing the functional similarity of gene products. The method relies on two semantic similarity measures; simRel and funSim. One measure (simRel is applied in the comparison of the biological processes found in different groups of organisms. The other measure (funSim is used to find functionally related gene products within the same or between different genomes. Results indicate that the method, in addition to being in good agreement with established sequence similarity approaches, also provides a means for the identification of functionally related proteins independent of evolutionary relationships. The method is also applied to estimating functional similarity between all proteins in Saccharomyces cerevisiae and to visualizing the molecular function space of yeast in a map of the functional space. A similar approach is used to visualize the functional relationships between protein families. Conclusion The approach enables the comparison of the underlying molecular biology of different taxonomic groups and provides a new comparative genomics tool identifying functionally related gene products independent of homology. The proposed map of the functional space provides a new global view on the functional relationships between gene products or protein families.

RESEARCH ARTICLE A cross-sectional study of PRNP gene in two ...

Indian Academy of Sciences (India)

Navya

Prion protein gene (PRNP) analysis revealed that none of the scrapie cases carried the p.Gln222Lys ... genetic variation, with low presence of wild type goats and the lack of protective alleles. .... We analyzed 157 clinically healthy goats (94 males and 63 females) in more than 20 .... Results from a retrospective cohort study.

Yangian symmetry of long-range gl(N) integrable spin chains

International Nuclear Information System (INIS)

Beisert, Niklas; Erkal, Denis

2008-01-01

An interesting type of spin chain has appeared in the context of the planar AdS/CFT correspondence: it is based on an integrable nearest-neighbor spin chain, and it is perturbatively deformed by long-range interactions which apparently preserve the integrable structure. Similar models can be constructed by demanding the existence of merely one conserved local charge. Although the latter is not a sufficient integrability condition in general, the models often display convincing signs of full integrability. Here we consider a class of long-range spin chains with spins transforming in the fundamental representation of gl(N). For the most general such model with one conserved local charge we construct a conserved Yangian generator and show that it obeys the Serre relations. We thus provide a formal proof of integrability for this class of models

[Hot spot mutation screening of RYR1 gene in diagnosis of congenital myopathies].

Science.gov (United States)

Chang, Xing-zhi; Jin, Yi-wen; Wang, Jing-min; Yuan, Yun; Xiong, Hui; Wang, Shuang; Qin, Jiong

2014-10-18

To detect hot spot mutation of RYR1 gene in 15 cases of congenital myopathy with different subtypes, and to discuss the value of RYR1 gene hot spot mutation detection in the diagnosis of the disease. Clinical data were collected in all the patients, including clinical manifestations and signs, serum creatine kinase, electromyography. Fourteen of the patients accepted the muscle biopsy. Hot spot mutation in the C-terminal of RYR1 gene (extron 96-106) had been detected in all the 15 patients. All the patients presented with motor development delay, and they could walk at the age of 1 to 3.5 years,but were always easy to fall and could not run or jump. There were no progressive deteriorations. Physical examination showed different degrees of muscle weakness and hypotonia.High arched palates were noted in 3 patients. The serum levels of creatine kinase were mildly elevated in 3 cases, and normal in 12 cases. Electromyography showed "myogenic" features in 11 patients, being normal in the other 4 patients. Muscle biopsy pathologic diagnosis was the central core disease in 3 patients, the central nuclei in 2 patients, the congenital fiber type disproportion in 2 patients, the nameline myopathy in 3 patient, the multiminicore disease in 1 patient, and nonspecific minimal changes in the other 3 patients; one patient was diagnosed with central core disease according to positive family history and gene mutation. In the family case (Patient 2) of central core disease, the c.14678G>A (p.Arg4893Gln) mutation in 102 extron of RYR1 was identified in three members of the family, which had been reported to be a pathogenic mutation. The c.14596A>G(p.Lys4866Gln) mutation in 101 extron was found in one patient with central core disease(Patient 1), and the c.14719G>A(p.Gly4907Ser) mutation in 102 extron was found in another case of the central core disease(Patient 3).The same novel mutation was verified in one of the patients' (Patient 3) asymptomatic father. Congenital myopathies in

Regulation of dietary glutamine on the growth, intestinal function, immunity and antioxidant capacity of sea cucumber Apostichopus japonicus (Selenka).

Science.gov (United States)

Yu, Haibo; Gao, Qinfeng; Dong, Shuanglin; Lan, Ying; Ye, Zhi; Wen, Bin

2016-03-01

The present study examined the effects of dietary glutamine (Gln) on the growth, intestinal function, immunity and antioxidant capacity of sea cucumber Apostichopus japonicus (Selenka). The specific growth rate, intestinal morphology, activity of digestive enzymes, activity and gene expression of lysozyme and antioxidative enzymes of the sea cucumbers were determined after feeding 5 experimental diets with additions of increasing levels of Gln (at 0%, 0.4%, 0.8%,1.2% and 1.6%, respectively) for 60 days. We discovered that the specific growth rate of the sea cucumbers in 0.4%, 0.8% and 1.2% groups increased 35.3%, 27.3% and 24.1%, respectively, compared to the control (0%) group with significant differences. Dietary Gln can improve the intestinal function of the sea cucumbers by increasing the activities of trypsin and lipase in the intestine and the villus height and villus density of the intestine, eventhough significant differences were not observed in some groups. 0.4%-0.8% of dietary Gln can significantly increase the activity of lysozyme (LSZ) in the coelomic fluid of the sea cucumbers. Significant improvements were observed on the SOD activity in coelomic fluid of the sea cucumbers fed diets supplemented with 0.4%-1.6% of Gln compared to the control group. Similarly, the CAT activity in coelomic fluid of the sea cucumbers significantly increased in 0.8%, 1.2% and 1.6% groups compared to the control and 0.4% groups. Change pattern of the activity of CAT was consistent with the change pattern of the expression of CAT gene, indicating the dietary Gln can up-regulate the expression of CAT gene and consequently promote the secretion of CAT. However, the down-regulation of the expression of SOD gene by dietary Gln were observed in almost all of the treatment groups, which is in contrast with the change pattern of the activity of SOD, indicating the negative feedback regulation of the secretion of SOD on the expression of SOD gene. In summary, the suitable

Recurrent vomiting and ethylmalonic aciduria associated with rare mutations in the short-chain acyl-CoA dehydrogenase (SCAD) gene

DEFF Research Database (Denmark)

Seidel, J.; Streck, S.; Bellstedt, K.

2003-01-01

blood spots. Neither of the frequent SCAD gene variants 625G>A and 511C>T was present, but direct sequencing of the promoter and coding regions of the SCAD gene revealed that the patient had mutations on both alleles: 417G>C (Trpl15Cys) and 1095G>T (Gln341His). Neither mutation has been described before...

Ghrelin and obestatin plasma levels and ghrelin/obestatin prepropeptide gene polymorphisms in small for gestational age infants.

Science.gov (United States)

Zhang, Shulian; Zhai, Guanpeng; Zhang, Jinping; Zhou, Jianguo; Chen, Chao

2014-12-01

To investigate plasma ghrelin and obestatin levels, and ghrelin/obestatin prepropeptide gene polymorphisms, in sequentially enrolled small for gestational age (SGA) infants. Neonates were sequentially enrolled into this study and were then subdivided into different groups, according to different study aims and availability of study materials. Consequently, plasma ghrelin and obestatin levels were measured in term SGA, term appropriate for gestational age (AGA), term large for gestational age (LGA), preterm SGA and preterm AGA neonates. Levels of both peptides were also measured in AGA infants of different gestational ages, and in term AGA neonates at different days following birth. Three ghrelin/obestatin prepropeptide gene single nucleotide polymorphisms (SNPs), Arg51Gln, Leu72Met, and Gln90Leu, were measured in neonates. The study involved a total cohort of 581 neonates. Out of 150 neonates (30 term AGA, 30 term SGA, 30 term LGA, 30 preterm AGA, and 30 preterm SGA), plasma obestatin levels were significantly higher in term SGA versus term LGA neonates (0.21 ± 0.02 ng/ml versus 0.17 ± 0.01 ng/ml, respectively). Out of a wider cohort, there were no significant differences in genotypes and allele frequencies of Arg51Gln, Leu72Met, and Gln90Leu SNPs between term SGA and AGA neonates, or between preterm SGA and AGA neonates. Ghrelin/obestatin prepropeptide polymorphisms were not found to be associated with SGA status in neonates; however, ghrelin and obestatin levels may be involved in growth and development. Further studies are required to understand the relationship between ghrelin, obestatin and prenatal development. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

Polymorphisms of XRCC1 genes and risk of nasopharyngeal carcinoma in the Cantonese population

International Nuclear Information System (INIS)

Cao, Yun; Miao, Xiao-Ping; Huang, Ma-Yan; Deng, Ling; Hu, Li-Fu; Ernberg, Ingemar; Zeng, Yi-Xin; Lin, Dong-Xin; Shao, Jian-Yong

2006-01-01

Nasopharyngeal carcinoma (NPC) is one of the most common cancers in southern China. In addition to environmental factors such as Epstein-Barr virus infection and diet, genetic susceptibility has been reported to play a key role in the development of this disease. The x-ray repair cross-complementing group 1 (XRCC1) gene is important in DNA base excision repair. We hypothesized that two common single nucleotide polymorphisms of XRCC1 (codons 194 Arg→Trp and 399 Arg→Gln) are related to the risk of NPC and interact with tobacco smoking. We sought to determine whether these genetic variants of the XRCC1 gene were associated with the risk of NPC among the Cantonese population in a hospital-based case control study using polymerase chain reaction-restriction fragment length polymorphism analysis. We conducted this study in 462 NPC patients and 511 healthy controls. After adjustment for sex and age, we found a reduced risk of developing NPC in individuals with the Trp194Trp genotype (OR = 0.48; 95% CI, 0.27–0.86) and the Arg194Trp genotype (OR = 0.79; 95% CI, 0.60–1.05) compared with those with the Arg194Arg genotype. Compared with those with the Arg399Arg genotype, the risk for NPC was not significantly different in individuals with the Arg399Gln genotype (OR = 0.82; 95% CI, 0.62–1.08) and the Gln399Gln genotype (OR = 1.20; 95% CI, 0.69–2.06). Further analyses stratified by gender and smoking status revealed a significantly reduced risk of NPC among males (OR = 0.32; 95% CI, 0.14–0.70) and smokers (OR = 0.34; 95% CI, 0.14–0.82) carrying the XRCC1 194Trp/Trp genotype compared with those carrying the Arg/Arg genotype. No association was observed between Arg399Gln variant genotypes and the risk of NPC combined with smoking and gender. Our findings suggest that the XRCC1 Trp194Trp variant genotype is associated with a reduced risk of developing NPC in Cantonese population, particularly in males and smokers. Larger studies are needed to confirm our findings

A novel missense variant (Gln220Arg) of GNB4 encoding guanine nucleotide-binding protein, subunit beta-4 in a Japanese family with autosomal dominant motor and sensory neuropathy.

Science.gov (United States)

Miura, Shiroh; Morikawa, Takuya; Fujioka, Ryuta; Noda, Kazuhito; Kosaka, Kengo; Taniwaki, Takayuki; Shibata, Hiroki

2017-09-01

Dominant intermediate Charcot-Marie-Tooth disease F (CMTDIF) is an autosomal dominant hereditary form of Charcot-Marie-Tooth disease (CMT) caused by variations in the guanine nucleotide-binding protein, subunit beta-4 gene (GNB4). We examined two Japanese familial cases with CMT. Case 1 was a 49-year-old male whose chief complaint was slowly progressive gait disturbance and limb dysesthesia that appeared at the age of 47. On neurological examination, he showed hyporeflexia or areflexia, distal limb muscle weakness, and distal sensory impairment with lower dominancy. Nerve conduction studies demonstrated demyelinating sensorimotor neuropathy with reduced action potentials in the lower limbs. Case 2 was an 80-year-old man, Case 1's father, who reported difficulty in riding a bicycle at the age of 76. On neurological examination, he showed areflexia in the upper and lower limbs. Distal sensory impairment in the lower limbs was also observed. Nerve conduction studies revealed mainly axonal involvement. Exome sequencing identified a novel heterozygous nonsynonymous variant (NM_021629.3:c.659T > C [p.Gln220Arg]) in GNB4 exon 8, which is known to be responsible for CMT. Sanger sequencing confirmed that both patients are heterozygous for the variation, which causes an amino acid substitution, Gln220Arg, in the highly conserved region of the WD40 domain of GNB4. The frequency of this variant in the Exome Aggregation Consortium Database was 0.000008247, and we confirmed its absence in 502 Japanese control subjects. We conclude that this novel GNB4 variant is causative for CMTDIF in these patients, who represent the first record of the disease in the Japanese population. Copyright © 2017. Published by Elsevier Masson SAS.

Relative rates of amino acid import via the ABC transporter GlnPQ determine the growth performance of Lactococcus lactis

NARCIS (Netherlands)

Fulyani, Faizah; Schuurman-Wolters, Geesina; Slotboom, Dirk-Jan; Poolman, Bert

The GlnPQ transporter from Lactococcus lactis has the remarkable feature of having two substrate-binding domains (SBD) fused to the N-terminus of the transmembrane domain (TMD), and thus four SBDs are present in the homodimeric complex. Although X-ray structures and ligand binding data are available

Comprehensive Maturity Onset Diabetes of the Young (MODY) Gene Screening in Pregnant Women with Diabetes in India.

Science.gov (United States)

Doddabelavangala Mruthyunjaya, Mahesh; Chapla, Aaron; Hesarghatta Shyamasunder, Asha; Varghese, Deny; Varshney, Manika; Paul, Johan; Inbakumari, Mercy; Christina, Flory; Varghese, Ron Thomas; Kuruvilla, Kurien Anil; V Paul, Thomas; Jose, Ruby; Regi, Annie; Lionel, Jessie; Jeyaseelan, L; Mathew, Jiji; Thomas, Nihal

2017-01-01

Pregnant women with diabetes may have underlying beta cell dysfunction due to mutations/rare variants in genes associated with Maturity Onset Diabetes of the Young (MODY). MODY gene screening would reveal those women genetically predisposed and previously unrecognized with a monogenic form of diabetes for further clinical management, family screening and genetic counselling. However, there are minimal data available on MODY gene variants in pregnant women with diabetes from India. In this study, utilizing the Next generation sequencing (NGS) based protocol fifty subjects were screened for variants in a panel of thirteen MODY genes. Of these subjects 18% (9/50) were positive for definite or likely pathogenic or uncertain MODY variants. The majority of these variants was identified in subjects with autosomal dominant family history, of whom five were in women with pre-GDM and four with overt-GDM. The identified variants included one patient with HNF1A Ser3Cys, two PDX1 Glu224Lys, His94Gln, two NEUROD1 Glu59Gln, Phe318Ser, one INS Gly44Arg, one GCK, one ABCC8 Arg620Cys and one BLK Val418Met variants. In addition, three of the seven offspring screened were positive for the identified variant. These identified variants were further confirmed by Sanger sequencing. In conclusion, these findings in pregnant women with diabetes, imply that a proportion of GDM patients with autosomal dominant family history may have MODY. Further NGS based comprehensive studies with larger samples are required to confirm these finding.

Comprehensive Maturity Onset Diabetes of the Young (MODY Gene Screening in Pregnant Women with Diabetes in India.

Directory of Open Access Journals (Sweden)

Mahesh Doddabelavangala Mruthyunjaya

Full Text Available Pregnant women with diabetes may have underlying beta cell dysfunction due to mutations/rare variants in genes associated with Maturity Onset Diabetes of the Young (MODY. MODY gene screening would reveal those women genetically predisposed and previously unrecognized with a monogenic form of diabetes for further clinical management, family screening and genetic counselling. However, there are minimal data available on MODY gene variants in pregnant women with diabetes from India. In this study, utilizing the Next generation sequencing (NGS based protocol fifty subjects were screened for variants in a panel of thirteen MODY genes. Of these subjects 18% (9/50 were positive for definite or likely pathogenic or uncertain MODY variants. The majority of these variants was identified in subjects with autosomal dominant family history, of whom five were in women with pre-GDM and four with overt-GDM. The identified variants included one patient with HNF1A Ser3Cys, two PDX1 Glu224Lys, His94Gln, two NEUROD1 Glu59Gln, Phe318Ser, one INS Gly44Arg, one GCK, one ABCC8 Arg620Cys and one BLK Val418Met variants. In addition, three of the seven offspring screened were positive for the identified variant. These identified variants were further confirmed by Sanger sequencing. In conclusion, these findings in pregnant women with diabetes, imply that a proportion of GDM patients with autosomal dominant family history may have MODY. Further NGS based comprehensive studies with larger samples are required to confirm these finding.

Comprehensive Maturity Onset Diabetes of the Young (MODY) Gene Screening in Pregnant Women with Diabetes in India

Science.gov (United States)

Hesarghatta Shyamasunder, Asha; Varghese, Deny; Varshney, Manika; Paul, Johan; Inbakumari, Mercy; Christina, Flory; Varghese, Ron Thomas; Kuruvilla, Kurien Anil; V. Paul, Thomas; Jose, Ruby; Regi, Annie; Lionel, Jessie; Jeyaseelan, L.; Mathew, Jiji; Thomas, Nihal

2017-01-01

Pregnant women with diabetes may have underlying beta cell dysfunction due to mutations/rare variants in genes associated with Maturity Onset Diabetes of the Young (MODY). MODY gene screening would reveal those women genetically predisposed and previously unrecognized with a monogenic form of diabetes for further clinical management, family screening and genetic counselling. However, there are minimal data available on MODY gene variants in pregnant women with diabetes from India. In this study, utilizing the Next generation sequencing (NGS) based protocol fifty subjects were screened for variants in a panel of thirteen MODY genes. Of these subjects 18% (9/50) were positive for definite or likely pathogenic or uncertain MODY variants. The majority of these variants was identified in subjects with autosomal dominant family history, of whom five were in women with pre-GDM and four with overt-GDM. The identified variants included one patient with HNF1A Ser3Cys, two PDX1 Glu224Lys, His94Gln, two NEUROD1 Glu59Gln, Phe318Ser, one INS Gly44Arg, one GCK, one ABCC8 Arg620Cys and one BLK Val418Met variants. In addition, three of the seven offspring screened were positive for the identified variant. These identified variants were further confirmed by Sanger sequencing. In conclusion, these findings in pregnant women with diabetes, imply that a proportion of GDM patients with autosomal dominant family history may have MODY. Further NGS based comprehensive studies with larger samples are required to confirm these finding PMID:28095440

Detection of biosurfactants in Bacillus species: genes and products identification.

Science.gov (United States)

Płaza, G; Chojniak, J; Rudnicka, K; Paraszkiewicz, K; Bernat, P

2015-10-01

To screen environmental Bacillus strains for detection of genes encoding the enzymes involved in biosurfactant synthesis and to evaluate their products e.g. surfactin, iturin and fengycin. The taxonomic identification of isolated from the environment Bacillus strains was performed by Microgene ID Bacillus panel and GEN III Biolog system. The polymerase chain reaction (PCR) strategy for screening of genes in Bacillus strains was set up. Liquid chromatography-mass spectrometry (LC-MS/MS) method was used for the identification of lipopeptides (LPs). All studied strains exhibited the presence of srfAA gene and produced surfactin mostly as four homologues (C13 to C16). Moreover, in 2 strains (KP7, T'-1) simultaneous co-production of 3 biosurfactants: surfactin, iturin and fengycin was observed. Additionally, it was found out that isolate identified as Bacillus subtilis ssp. subtilis (KP7), beside LPs co-production, synthesizes surfactin with the efficiency much higher than other studied strains (40·2 mg l(-1) ) and with the yield ranging from 0·8 to 8·3 mg l(-1) . We showed that the combined methodology based on PCR and LC-MS/MS technique is an optimal tool for the detection of genes encoding enzymes involved in biosurfactant synthesis as well as their products, e.g. surfactin, iturin and fengycin. This approach improves the screening and the identification of environmental Bacillus co-producing biosurfactants-stimulating and facilitating the development of this area of science. The findings of this work will help to improve screening of biosurfactant producers. Discovery of novel biosurfactants and biosurfactants co-production ability has shed light on their new application fields and for the understanding of their interactions and properties. © 2015 The Society for Applied Microbiology.

Data of evolutionary structure change: 1BYUA-3RANC [Confc[Archive

Lifescience Database Archive (English)

Full Text Available 1BYUA-3RANC 1BYU 3RAN A C --EPQVQFKLVLVGDGGTGKTTFVKRHLTGEFEKKYVPT...142 CA TYR A 147 10.374 24.725 19.466 1.00 14.34 C e-map> TYR TYR ...in> 3RAN C 3RANC e-map> TYR TYR GLN LEU ASN LYS LYS ARG... GLN CA 6.93 3.86 TYR CA 3.88 TYR CA e-map> S

Impact of alg3 gene deletion on growth, development, pigment production, protein secretion, and functions of recombinant Trichoderma reesei cellobiohydrolases in Aspergillus niger

Energy Technology Data Exchange (ETDEWEB)

Dai, Ziyu; Aryal, Uma K.; Shukla, Anil; Qian, Wei-Jun; Smith, Richard D.; Magnuson, Jon K.; Adney, William S.; Beckham, Gregg T.; Brunecky, Roman; Himmel, Michael E.; Decker, Stephen R.; Ju, Xiaohui; Zhang, Xiao; Baker, Scott E.

2013-12-01

ALG3 is a Family 58 glycosyltransferase enzyme involved in early N-linked glycan synthesis. Here, we investigated the effect of the alg3 gene disruption on growth, development, metabolism, and protein secretion in Aspergillus niger. The alg3 gene deletion resulted in a significant reduction of growth on complete (CM) and potato dextrose agar (PDA) media and a substantial reduction of spore production on CM. It also delayed spore germination in the liquid cultures of both CM and PDA media, but led to a significant accumulation of red pigment on both CM and liquid modified minimal medium (MM) supplemented with yeast extract. The relative abundance of 55 proteins of the total 190 proteins identified in the secretome was significantly different as a result of alg3 gene deletion. Comparison of a Trichoderma reesei cellobiohydrolase (Cel7A) heterologously expressed in A. niger parental and Δalg3 strains showed that the recombinant Cel7A expressed in the mutant background was smaller in size than that from the parental strains. This study suggests that ALG3 is critical for growth and development, pigment production, and protein secretion in A. niger. Functional analysis of recombinant Cel7A with aberrant glycosylation demonstrates the feasibility of this alternative approach to evaluate the role of N-linked glycosylation in glycoprotein secretion and function.

Trans-activation function of a 3' truncated X gene-cell fusion product from integrated hepatitis B virus DNA in chronic hepatitis tissues

International Nuclear Information System (INIS)

Takada, Shinako; Koike, Katsuro

1990-01-01

To investigate the expression and transactivation function of the X gene in integrated hepatitis B virus (HBV) DNA from chronic hepatitis tissues, a series of transfectants containing cloned integrated HBV DNAs was made and analyzed for X mRNA expression and trans-activation activity by using a chloramphenicol acetyltransferase assay. Most of the integrated HBV DNAs expressed X mRNA and encoded a product with trans-activation activity in spite of the loss of the 3' end region of the X gene due to integration. From cDNA cloning and sequence analysis of X mRNA transcribed from native or integrated HBV DNA, the X protein was found to be translated from the X open reading frame without splicing. For integrated HBV DNA, transcription was extended to a cellular flanking DNA and an X gene-cell fusion transcript was terminated by using a cellular poly(A) signal. The amino acid sequence deduced from an X-cell fusion transcript indicated truncation of the carboxyl-terminal five amino acids, but the upstream region of seven amino acids conserved among hepadnaviruses was retained in the integrated HBV DNA, suggesting that this conserved region is essential for the transactivation function of the X protein. These findings support the following explanation for hepatocarcinogenesis by HBV DNA integration: the expression of a cellular oncogene(s) is transactivated at the time of chronic infection by the increasing amounts of the integrated HBV gene product(s), such as the X-cell fusion product

Mutation at codon 442 in the rpoB gene of Mycobacterium leprae does not confer resistance to rifampicin.

Science.gov (United States)

Lavania, Mallika; Hena, Abu; Reja, Hasanoor; Nigam, Astha; Biswas, Nibir Kumar; Singh, Itu; Turankar, Ravindra P; Gupta, Ud; Kumar, Senthil; Rewaria, Latika; Patra, Pradip K R; Sengupta, Utpal; Bhattacharya, Basudeb

2016-03-01

Rifampicin is the major drug in the treatment of leprosy. The rifampicin resistance of Mycobacterium leprae results from a mutation in the rpoB gene, encoding the β subunit of RNA polymerase. As M. leprae is a non-cultivable organism observation of its growth using mouse food-pad (MFP) is the only Gold Standard assay used for confirmation of "in-vivo" drug resistance. Any mutation at molecular level has to be verified by MFP assay for final confirmation of drug resistance in leprosy. In the present study, M. leprae strains showing a mutation only at codon 442 Gln-His and along with mutation either at codon 424 Val-Gly or at 438 Gln-Val within the Rifampicin Resistance Determining Region (RRDR) confirmed by DNA sequencing and by high resolution melting (HRM) analysis were subjected for its growth in MFP. The M. leprae strain having the new mutation at codon 442 Gln-His was found to be sensitive to all the three drugs and strains having additional mutations at 424 Val-Gly and 438 Gln-Val were conferring resistance with Multi drug therapy (MDT) in MFP. These results indicate that MFP is the gold standard method for confirming the mutations detected by molecular techniques.

Primary structure, gene organization and polypeptide expression of poliovirus RNA

Energy Technology Data Exchange (ETDEWEB)

Kitamura, N. (State Univ. of New York, Stony Brook); Semler, B.L.; Rothberg, P.G.

1981-06-18

The primary structure of the poliovirus genome has been determined. The RNA molecule is 7433 nucleotides long, polyadenylated at the 3' terminus, and covalently linked to a small protein (VPg) at the 5' terminus. An open reading frame of 2207 consecutive triplets spans over 89% of the nucleotide sequence and codes for the viral polyprotein NCVPOO. Twelve viral polypeptides have been mapped by amino acid sequence analysis and were found to be proteolytic cleavage products of the polyprotein, cleavages occurring predominantly at Gln-Gly pairs.

The chicken c-erbA alpha-product induces expression of thyroid hormone-responsive genes in 3,5,3'-triiodothyronine receptor-deficient rat hepatoma cells

DEFF Research Database (Denmark)

Muñoz, A; Höppner, W; Sap, J

1990-01-01

To determine the capacity of the chicken c-erbA (cTR-alpha) gene product in regulating expression of known thyroid hormone-responsive genes, both the cTR-alpha and the viral v-erbA genes were expressed in FAO cells, a rat hepatoma cell line defective for functional thyroid hormone receptors. Upon...

Identification of polymorphisms of XRCC1 gene in patients with cancer in a city of northern Brazil

Directory of Open Access Journals (Sweden)

Artemis Socorro N. Rodrigues

2015-06-01

Full Text Available ABSTRACT Introduction: Cancer is considered a genetic disease. For this reason, identification and characterization of the genes involved in its origin and progression are of fundamental importance in understanding its molecular basis. Objective: Our objective was to determine whether people from Macapá with a diagnosis of cancer have genetic polymorphisms related to the XRCC1 gene. Materials and methods: We analyzed 30 samples of deoxyribonucleic acid (DNA of cases with cancer and 30 control samples. All samples were amplified and analyzed by the polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP method, with the use of restriction enzyme MspI. Results: Regarding the 194T polymorphism, we found that all samples of the cases presented the polymorphic allele Trp (Arg/Trp. In control samples, 96.6% also identify the polymorphic allele Trp and, among these, one was homozygous for the same allele (Trp/Trp. Regarding the 399A polymorphism, 83.3% of the cases and 23.3% of the controls had the Arg/ Gln genotype, respectively. We found that 73.3% of controls and 16.6% of cases had the Arg/Arg genotype. Among the controls, we found only a sample that was homozygous for the polymorphic allele Trp/Trp. Conclusion: Our results demonstrated the allele frequency of 194Trp polymorphism in both sample groups analyzed. We also found a significant number of polymorphic allele 399A in people with cancer. Thus, we can highlight 399Gln polymorphism as a genetic marker of cancer risk in this population.

Novel homozygous nonsense mutations in the luteinizing hormone receptor (LHCGR) gene associated with 46,XY primary amenorrhea.

Science.gov (United States)

Ben Hadj Hmida, Imen; Mougou-Zerelli, Soumaya; Hadded, Anis; Dimassi, Sarra; Kammoun, Molka; Bignon-Topalovic, Joelle; Bibi, Mohamed; Saad, Ali; Bashamboo, Anu; McElreavey, Ken

2016-07-01

To determine the genetic cause of 46,XY primary amenorrhea in three 46,XY girls. Whole exome sequencing. University cytogenetics center. Three patients with unexplained 46,XY primary amenorrhea were included in the study. Potentially pathogenic variants were confirmed by Sanger sequencing, and familial segregation was determined where parents' DNA was available. Exome sequencing was performed in the three patients, and the data were analyzed for potentially pathogenic mutations. The functional consequences of mutations were predicted. Three novel homozygous nonsense mutations in the luteinizing hormone receptor (LHCGR) gene were identified:c.1573 C→T, p.Gln525Ter, c.1435 C→T p.Arg479Ter, and c.508 C→T, p.Gln170Ter. Inactivating mutations of the LHCGR gene may be a more common cause of 46,XY primary amenorrhea than previously considered. Copyright © 2016 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Pharmacogenetic Study in Rectal Cancer Patients Treated With Preoperative Chemoradiotherapy: Polymorphisms in Thymidylate Synthase, Epidermal Growth Factor Receptor, GSTP1, and DNA Repair Genes

International Nuclear Information System (INIS)

Páez, David; Salazar, Juliana; Paré, Laia; Pertriz, Lourdes; Targarona, Eduardo; Rio, Elisabeth del; Barnadas, Agusti; Marcuello, Eugenio; Baiget, Montserrat

2011-01-01

Purpose: Several studies have been performed to evaluate the usefulness of neoadjuvant treatment using oxaliplatin and fluoropyrimidines for locally advanced rectal cancer. However, preoperative biomarkers of outcome are lacking. We studied the polymorphisms in thymidylate synthase, epidermal growth factor receptor, glutathione S-transferase pi 1 (GSTP1), and several DNA repair genes to evaluate their usefulness as pharmacogenetic markers in a cohort of 128 rectal cancer patients treated with preoperative chemoradiotherapy. Methods and Materials: Blood samples were obtained from 128 patients with Stage II-III rectal cancer. DNA was extracted from the peripheral blood nucleated cells, and the genotypes were analyzed by polymerase chain reaction amplification and automated sequencing techniques or using a 48.48 dynamic array on the BioMark system. The germline polymorphisms studied were thymidylate synthase, (VNTR/5′UTR, 2R G>C single nucleotide polymorphism [SNP], 3R G>C SNP), epidermal growth factor receptor (Arg497Lys), GSTP1 (Ile105val), excision repair cross-complementing 1 (Asn118Asn, 8092C>A, 19716G>C), X-ray repair cross-complementing group 1 (XRCC1) (Arg194Trp, Arg280His, Arg399Gln), and xeroderma pigmentosum group D (Lys751Gln). The pathologic response, pathologic regression, progression-free survival, and overall survival were evaluated according to each genotype. Results: The ∗3/∗3 thymidylate synthase genotype was associated with a greater response rate (pathologic complete remission and microfoci residual tumor, 59% in ∗3/∗3 vs. 35% in ∗2/∗2 and ∗2/∗3; p = .013). For the thymidylate synthase genotype, the median progression-free survival was 103 months for the ∗3/∗3 patients and 84 months for the ∗2/∗2 and ∗2/∗3 patients (p = .039). For XRCC1 Arg399Gln SNP, the median progression-free survival was 101 months for the G/G, 78 months for the G/A, and 31 months for the A/A patients (p = .048). Conclusions: The thymidylate

Pharmacogenetic Study in Rectal Cancer Patients Treated With Preoperative Chemoradiotherapy: Polymorphisms in Thymidylate Synthase, Epidermal Growth Factor Receptor, GSTP1, and DNA Repair Genes

Energy Technology Data Exchange (ETDEWEB)

Paez, David, E-mail: dpaez@santpau.cat [Department of Medical Oncology, Hospital de la Santa Creu i Sant Pau, Universitat Autonoma de Barcelona, Barcelona (Spain); Salazar, Juliana; Pare, Laia [Centre for Biomedical Network Research on Rare Diseases, Barcelona (Spain); Department of Genetics, Hospital de la Santa Creu i Sant Pau, Universitat Autonoma de Barcelona, Barcelona (Spain); Pertriz, Lourdes [Department of Radiotherapy, Hospital de la Santa Creu i Sant Pau, Universitat Autonoma de Barcelona, Barcelona (Spain); Targarona, Eduardo [Department of Surgery, Hospital de la Santa Creu i Sant Pau, Universitat Autonoma de Barcelona, Barcelona (Spain); Rio, Elisabeth del [Centre for Biomedical Network Research on Rare Diseases, Barcelona (Spain); Department of Genetics, Hospital de la Santa Creu i Sant Pau, Universitat Autonoma de Barcelona, Barcelona (Spain); Barnadas, Agusti; Marcuello, Eugenio [Department of Medical Oncology, Hospital de la Santa Creu i Sant Pau, Universitat Autonoma de Barcelona, Barcelona (Spain); Baiget, Montserrat [Centre for Biomedical Network Research on Rare Diseases, Barcelona (Spain); Department of Genetics, Hospital de la Santa Creu i Sant Pau, Universitat Autonoma de Barcelona, Barcelona (Spain)

2011-12-01

Purpose: Several studies have been performed to evaluate the usefulness of neoadjuvant treatment using oxaliplatin and fluoropyrimidines for locally advanced rectal cancer. However, preoperative biomarkers of outcome are lacking. We studied the polymorphisms in thymidylate synthase, epidermal growth factor receptor, glutathione S-transferase pi 1 (GSTP1), and several DNA repair genes to evaluate their usefulness as pharmacogenetic markers in a cohort of 128 rectal cancer patients treated with preoperative chemoradiotherapy. Methods and Materials: Blood samples were obtained from 128 patients with Stage II-III rectal cancer. DNA was extracted from the peripheral blood nucleated cells, and the genotypes were analyzed by polymerase chain reaction amplification and automated sequencing techniques or using a 48.48 dynamic array on the BioMark system. The germline polymorphisms studied were thymidylate synthase, (VNTR/5 Prime UTR, 2R G>C single nucleotide polymorphism [SNP], 3R G>C SNP), epidermal growth factor receptor (Arg497Lys), GSTP1 (Ile105val), excision repair cross-complementing 1 (Asn118Asn, 8092C>A, 19716G>C), X-ray repair cross-complementing group 1 (XRCC1) (Arg194Trp, Arg280His, Arg399Gln), and xeroderma pigmentosum group D (Lys751Gln). The pathologic response, pathologic regression, progression-free survival, and overall survival were evaluated according to each genotype. Results: The Asterisk-Operator 3/ Asterisk-Operator 3 thymidylate synthase genotype was associated with a greater response rate (pathologic complete remission and microfoci residual tumor, 59% in Asterisk-Operator 3/ Asterisk-Operator 3 vs. 35% in Asterisk-Operator 2/ Asterisk-Operator 2 and Asterisk-Operator 2/ Asterisk-Operator 3; p = .013). For the thymidylate synthase genotype, the median progression-free survival was 103 months for the Asterisk-Operator 3/ Asterisk-Operator 3 patients and 84 months for the Asterisk-Operator 2/ Asterisk-Operator 2 and Asterisk-Operator 2/ Asterisk

DNA repair synthesis dependent on the uvrA,B gene products

International Nuclear Information System (INIS)

Moses, R.E.; Moody, E.E.M.

1975-01-01

Ultraviolet irradiation of toluene-treated Escherichia coli causes an inhibition of replicative DNA synthesis. This is followed by the appearance of nonconservative DNA repair synthesis which does not require either the polymerase or 5' → 3' exonucleolytic activities of DNA polymerase I. The repair synthesis may be catalyzed by DNA polymerase III activity but does not require a functional DNA polymerase II. The ultraviolet-induced synthesis requires ATP and is dependent on a functional uvrA and uvrB gene product. However, other uvr gene products are not required for the synthesis. The recB function is also not required

Revealing gene action for production characteristics by inbreeding ...

African Journals Online (AJOL)

Revealing gene action for production characteristics by inbreeding, based on a long-term selection ... The gene action involved in the expression of production characters was investigated, using the effect of the theoretical inbreeding ..... and predicted selection responses for growth, fat and lean traits in mice. J. Anim. Sci.

In vitro regulation of LH biosynthesis and release by GnRH and estradiol

International Nuclear Information System (INIS)

Ramey, J.W.

1986-01-01

Anterior pituitaries were taken from female rats at random stages of the estrous cycle, enzymatically dispersed, and cultured for 48h in steroid-free α-modified Eagles medium followed by 24h in fresh medium +/- estradiol (E 2 ). The pituitary cells were then incubated in fresh medium containing radiolabeled precursors +/- gonadotropin releasing hormone (GnRH). Radioactive precursor incorporation into LH was determined by immuno-precipitation. The dose-dependent effects of E 2 (10 -11 to 10 -8 M) on 3 H-glucosamine ( 3 H-Gln) and 35 S-methionine ( 35 S-Met) incorporation into LH +/- 1 nM GnRH (4h) were investigated. GnRH (10 -9 M) and E 2 (all doses) significantly increased total 3 H-Gln LH. Moreover, E 2 at 10 -9 M and 10 -8 M significantly enhanced GnRH stimulated LH glycosylation. In contrast, addition of GnRH and/or E 2 did not significantly increase 35 S-Met incorporation into LH over a 4h period. The effects of various GnRH concentrations (10 -11 to 10 -9 M; 8h) +/- E 2 (0.05 nM) on 3 H-Gln LH and 35 S-Met LH production were also investigated. In the absence of E 2 , only 10 -9 M GnRH was effective in increasing total 3 H-Gln LH and 35 S-Met LH synthesis. However, in the presence of E 2 , all concentrations of GnRH stimulated LH synthesis with 3 H-Gln LH production responding in a dose related manner whereas 35 S-Met LH production was maximally stimulated at all doses of GnRH. In the final series of experiments, pituitary cells previously exposed to estradiol were incubated for 4 h in normal calcium or low calcium medium containing 3 H-Gln or 35 S-Met +/- GnRH. Removal of extracellular calcium completely inhibited GnRH stimulated 3 H-Gln LH and 35 S-Met LH production

Integrating Ontological Knowledge and Textual Evidence in Estimating Gene and Gene Product Similarity

Energy Technology Data Exchange (ETDEWEB)

Sanfilippo, Antonio P.; Posse, Christian; Gopalan, Banu; Tratz, Stephen C.; Gregory, Michelle L.

2006-06-08

With the rising influence of the Gene On-tology, new approaches have emerged where the similarity between genes or gene products is obtained by comparing Gene Ontology code annotations associ-ated with them. So far, these approaches have solely relied on the knowledge en-coded in the Gene Ontology and the gene annotations associated with the Gene On-tology database. The goal of this paper is to demonstrate that improvements to these approaches can be obtained by integrating textual evidence extracted from relevant biomedical literature.

ROLE OF GENE POLYMORFISM OF PLASMINOGEN ACTIVATOR INGIBITOR TYPE I AS A RISK FACTOR FOR PREMATURE RUPTURE OF MEMBRANE AT TERM PREGNANCY

Directory of Open Access Journals (Sweden)

M. G. Nikolayeva

2013-01-01

Full Text Available The retrospective study was designed to identify association of premature rupture of the fetal membranes (PROM with carrying polymorphisms in genes encoding folate metabolism and hemostasis in 717 women. More than one hundred potential predictors were analyzed including carriage of thrombogenic genes polymorphisms and genes encoding folate metabolism: FV[Arg506Gln], F II [20210 G/A], MTHFR [Ala222Val], (PAI-I[-675 5G/4G]. Study revealed that plasminogen activator ingibitor-1 gene polymorphism increases significantly the risk of premature rupture of the fetal membranes in term pregnancy (PROM: heterozygous plasminogen activator ingibitor-1 gene polymorphism is associated with 3.6-fold (95% CI 2.4–5.4; p < 0.001, homozygous plasminogen activator ingibitor-1 gene polymorphism – with 1.7-fold (95% CI 1.1–2.6; p = 0.01 risk rise of PROM.

The Gln223Arg polymorphism of the leptin receptor in Pima Indians: influence on energy expenditure, physical activity and lipid metabolism

DEFF Research Database (Denmark)

Stefan, N; Vozarova, B; Del Parigi, A

2002-01-01

Leptin regulates body weight by its receptor-mediated anorectic, thermogenic and antisteatotic effects. Recently, lower leptin binding to the soluble form of the leptin receptor (LEPR) was shown in carriers of the Arg223-encoding allele of the Gln223Arg polymorphism of the LEPR. To investigate wh...

Detection of individual radiosensitivity by radiation–induced micronuclei in human peripheral blood lymphocytes and polymorphisms in DNA repair genes

Energy Technology Data Exchange (ETDEWEB)

Staynova, A.; Hadjidekova, V.; Popova, L.; Hristova, R. [Radiation Genetics Laboratory, National Centre of Radiobology and Radiation Protection, Sofia (Bulgaria); Savov, A. [National Genetic Laboratory, University Hospital of Obstetrics and Gynecology, Sofia (Bulgaria)

2013-07-01

Aim: To investigate the association of two polymorphisms – in XRCC1 gene (Arg399Gln) and in APE1 gene (Asp148Glu) and the radiation induced frequency of micronuclei in human peripheral blood lymphocytes. Material and methods: Genomic DNA from 34 cancer patients and 52 controls were genotyped using PCR–RFLP technique. Micronucleus test (MNT) was performed on 15 cancer patients and 15 controls, before and after in vitro irradiation with 2Gy gamma rays. Results: The data showed that cancer patients had a significantly higher spontaneous frequency of cells with micronuclei than controls (P=0.009). No statistical difference was registered when comparing the mean frequency of cells with micronuclei after in vitro irradiation between these groups. Four subjects were selected as radiosensitive after applying cut–off of the mean frequency of radiation induced micronuclei. Three of them are carriers of the XRCC1 399Gln allele and two of them are carriers of the APE1 148Glu allele. (author)

Guidelines for the naming of genes, gene products, and mutants in the opportunistic protists.

Science.gov (United States)

Limper, Andrew H; Weiss, Louis M

2011-01-01

The opportunistic protists encompass a wide diversity of organisms including Pneumocystis, Toxoplasma, cryptosporidia, microsporidia, and related genera. Recent advances in the molecular biology and cellular biochemistry of these organisms have led to the identification of an ever growing numbers of key genes and their cognate proteins. Until now, these molecules have not been designated using any consistent nomenclature system, leading to considerable confusion. The participants of the 11th International Workshop on Opportunistic Protists met on August 3, 2010 to reach consensus of a nomenclature system for genes, gene products, and mutants in the opportunistic protists. The following summary reports the consensus agreement to move toward a unified nomenclature system for these organisms. The system is adapted from that used for Saccharomyces cerevisiae. © 2011 The Author(s). Journal of Eukaryotic Microbiology © 2011 International Society of Protistologists.

Expression of cocoa genes in Saccharomyces cerevisiae improves cocoa butter production

DEFF Research Database (Denmark)

Wei, Yongjun; Bergenholm, David; Gossing, Michael

2018-01-01

Background: Cocoa butter (CB) extracted from cocoa beans (Theobroma cacao) is the main raw material for chocolate production, but CB supply is insufficient due to the increased chocolate demand and limited CB production. CB is mainly composed of three different kinds of triacylglycerols (TAGs), 1......), and it is essential to modulate the yeast TAG biosynthetic pathway for higher CBL production.Results: We cloned seven GPAT genes and three LPAT genes from cocoa cDNA, in order to screen for CBL biosynthetic gene candidates. By expressing these cloned cocoa genes and two synthesized cocoa DGAT genes in S. cerevisiae......, we successfully increased total fatty acid production, TAG production and CBL production in some of the strains. In the best producer, the potential CBL content was eightfold higher than the control strain, suggesting the cocoa genes expressed in this strain were functional and might be responsible...

Regulation of gene expression by tobacco product preparations in cultured human dermal fibroblasts

International Nuclear Information System (INIS)

Malpass, Gloria E.; Arimilli, Subhashini; Prasad, G.L.; Howlett, Allyn C.

2014-01-01

Skin fibroblasts comprise the first barrier of defense against wounds, and tobacco products directly contact the oral cavity. Cultured human dermal fibroblasts were exposed to smokeless tobacco extract (STE), total particulate matter (TPM) from tobacco smoke, or nicotine at concentrations comparable to those found in these extracts for 1 h or 5 h. Differences were identified in pathway-specific genes between treatments and vehicle using qRT-PCR. At 1 h, IL1α was suppressed significantly by TPM and less significantly by STE. Neither FOS nor JUN was suppressed at 1 h by tobacco products. IL8, TNFα, VCAM1, and NFκB1 were suppressed after 5 h with STE, whereas only TNFα and NFκB1 were suppressed by TPM. At 1 h with TPM, secreted levels of IL10 and TNFα were increased. Potentially confounding effects of nicotine were exemplified by genes such as ATF3 (5 h), which was increased by nicotine but suppressed by other components of STE. Within 2 h, TPM stimulated nitric oxide production, and both STE and TPM increased reactive oxygen species. The biological significance of these findings and utilization of the gene expression changes reported herein regarding effects of the tobacco product preparations on dermal fibroblasts will require additional research. - Highlights: • Tobacco product preparations (TPPs) alter gene expression in dermal fibroblasts. • Some immediate early genes critical to the inflammatory process are affected. • Different TPPs produce differential responses in certain pro-inflammatory genes

Regulation of gene expression by tobacco product preparations in cultured human dermal fibroblasts

Energy Technology Data Exchange (ETDEWEB)

Malpass, Gloria E., E-mail: gloria.malpass@gmail.com [Department of Physiology and Pharmacology, Wake Forest University Health Sciences, Winston-Salem, NC 27157 (United States); Arimilli, Subhashini, E-mail: sarimill@wakehealth.edu [Department of Microbiology and Immunology, Wake Forest University Health Sciences, Winston-Salem, NC 27157 (United States); Prasad, G.L., E-mail: prasadg@rjrt.com [R and D Department, R.J. Reynolds Tobacco Company, Winston-Salem, NC 27102 (United States); Howlett, Allyn C., E-mail: ahowlett@wakehealth.edu [Department of Physiology and Pharmacology, Wake Forest University Health Sciences, Winston-Salem, NC 27157 (United States)

2014-09-01

Skin fibroblasts comprise the first barrier of defense against wounds, and tobacco products directly contact the oral cavity. Cultured human dermal fibroblasts were exposed to smokeless tobacco extract (STE), total particulate matter (TPM) from tobacco smoke, or nicotine at concentrations comparable to those found in these extracts for 1 h or 5 h. Differences were identified in pathway-specific genes between treatments and vehicle using qRT-PCR. At 1 h, IL1α was suppressed significantly by TPM and less significantly by STE. Neither FOS nor JUN was suppressed at 1 h by tobacco products. IL8, TNFα, VCAM1, and NFκB1 were suppressed after 5 h with STE, whereas only TNFα and NFκB1 were suppressed by TPM. At 1 h with TPM, secreted levels of IL10 and TNFα were increased. Potentially confounding effects of nicotine were exemplified by genes such as ATF3 (5 h), which was increased by nicotine but suppressed by other components of STE. Within 2 h, TPM stimulated nitric oxide production, and both STE and TPM increased reactive oxygen species. The biological significance of these findings and utilization of the gene expression changes reported herein regarding effects of the tobacco product preparations on dermal fibroblasts will require additional research. - Highlights: • Tobacco product preparations (TPPs) alter gene expression in dermal fibroblasts. • Some immediate early genes critical to the inflammatory process are affected. • Different TPPs produce differential responses in certain pro-inflammatory genes.

Reduced Hsp70 and Glutamine in Pediatric Severe Malaria Anemia

DEFF Research Database (Denmark)

Kempaiah, Prakasha; Dokladny, Karol; Karim, Zachary

2016-01-01

by decreased HSPA1A, a heat shock protein (Hsp) 70 coding gene. Hsp70 is a ubiquitous chaperone that regulates Nuclear Factor-kappa B (NF-κB) signaling and production of pro-inflammatory cytokines known to be important in malaria pathogenesis (e.g., IL-1β, IL-6 and TNF-α). Since the role of host Hsp70...... in malaria pathogenesis is unexplored, we investigated Hsp70 and molecular pathways in children with SMA. Validation experiments revealed that leukocytic HSP70 transcripts were reduced in SMA relative to non-severe malaria, and that intraleukocytic hemozoin (PfHz) was associated with lower HSP70. HSP70...... was correlated with reticulocyte production and Hb. Since glutamine (Gln) up-regulates Hsp70, modulates NF-κB activation, and attenuates over-expression of pro-inflammatory cytokines, circulating Gln was measured in children with malaria. Reduced Gln was associated with increased risk of developing SMA...

Implications of XRCC1, XPD and APE1 gene polymorphism in North Indian population: a comparative approach in different ethnic groups worldwide.

Science.gov (United States)

Gangwar, Ruchika; Manchanda, Parmeet Kaur; Mittal, Rama Devi

2009-05-01

Identifying risk factors for human cancers should consider combinations of genetic variations and environmental exposures. Several polymorphisms in DNA repair genes have impact on repair and cancer susceptibility. We focused on X-ray repair cross-complementing group 1 (XRCC1), Xeroderma pigmentosum D (XPD) and apurinic/apyrimidinic endonuclease (APE1) as these are most extensively studied in cancer. Present study was conducted to determine distribution of XRCC1 C26304T, G27466A, G23591A, APE1 T2197G and XPD A35931C gene polymorphisms in North Indian population and compare with different populations globally. PCR-based analysis was conducted in 209 normal healthy individuals of similar ethnicity. Allelic frequencies in wild type of XRCC1 C26304T were 91.1% C(Arg); G27466A 62.9% G(Arg); G23591A 60.3% G(Arg); APE1 T2197G 75.1% T(Asp) and XPD A35931C 71.8% A(Lys). The variant allele frequency were 8.9% T(Trp) in XRCC1 C26304T; 37.1% A(His) in G27466A; 39.7% A(Gln) in G23591A; 24.9% G(Glu) in APE1 and 28.2% C(Gln) in XPD respectively. We further compared frequency distribution for these genes with various published studies in different ethnicity. Our results suggest that frequency in these DNA repair genes exhibit distinctive pattern in India that could be attributed to ethnicity variation. This could assist in high-risk screening of humans exposed to environmental carcinogens and cancer predisposition in different ethnic groups.

Genetic variation in the leptin receptor gene, leptin, and weight gain in young Dutch adults.

Science.gov (United States)

van Rossum, Caroline T M; Hoebee, Barbara; van Baak, Marleen A; Mars, Monica; Saris, Wim H M; Seidell, Jacob C

2003-03-01

To investigate the association between leptin levels, polymorphisms in the leptin receptor (LEPR) gene, and weight gain. From two large prospective cohorts in The Netherlands (n = 17,500), we compared the baseline leptin of 259 subjects who had gained an average of 12.6 kg (range 5.5 to 33 kg) with 277 subjects who kept stable weight (range -2.6 to 3.1 kg) after a mean follow-up of 6.8 years. Three polymorphisms in the LEPR gene (Lys109Arg, Gln223Arg, and Lys656Asn) were determined. Weight gainers had significantly higher baseline leptin levels than those who kept stable weight (odds ratio = 1.27, 95% confidence interval 1.1 to 1.5, per SD increase in log(e)-transformed leptin). Weight gainers with the Arg109 or the Arg223 alleles had higher leptin levels compared with the noncarriers of these alleles. Only among men, the association between leptin and weight gain tended to be stronger among those with an Arg223 allele compared with those without this mutation. Relatively high leptin levels predict weight gain, suggesting that leptin resistance plays a role in the development of obesity in the general population. Higher leptin levels for those with a Lys109Arg or Gln223Arg mutation (or a linked other marker) may imply that these subjects have a modified functional leptin receptor. However, the role of these mutations on weight gain is limited.

Analysing the dhaT gene in Colombian Clostridium sp. (Clostridia 1,3-propanediol-producing strains

Directory of Open Access Journals (Sweden)

Diana Milena Quilaguy-Ayure

2010-04-01

Full Text Available To analyze the dhaT gene, one of the genes responsible for the 1,3-propanediol (1,3-PD production, in two native Clostridiumstrains. Materials and methods: The dhaT gene was amplified by Polimerase Chain Reaction with specific primers designed fromClostridium butyricum VPI1718 operon. Bioinformatics tools like BLASTN, ORF finder, BLASTP and ClustalW were used to determinethe identity of the sequence and to assign a function. Results: DNA amplification products were obtained from Colombian Clostridium sp.native strains (IBUN 13A and IBUN 158B and the Clostridium butyricum DSM 2478 strain, which were sequenced. According to thebioinformatics analysis of the above sequences, a high degree of similarity was found with the dhaT gene of different bacterial species. Thehighest percentage of identity was obtained with the Clostridium butyricum VPI 1718 strain. Conclusion: knowledge of the physicalstructure of the 1,3-PD operon in native strains opens the way for developing genetic and metabolic engineering strategies for improvingprocesses productivity.

Impact of two common xeroderma pigmentosum group D (XPD gene polymorphisms on risk of prostate cancer.

Directory of Open Access Journals (Sweden)

Yuanyuan Mi

Full Text Available BACKGROUND: DNA repair genes (EG: xeroderma pigmentosum group D, XPD may affect the capacity of encoded DNA repair enzymes to effectively remove DNA adducts or lesions, which may result in enhanced cancer risk. The association between XPD gene polymorphisms and the susceptibility of prostate cancer (PCa was inconsistent in previous studies. METHODOLOGY/PRINCIPAL FINDINGS: A meta-analysis based on 9 independent case-control studies involving 3165 PCa patients and 3539 healthy controls for XPD Gln751Lys SNP (single nucleotide polymorphism and 2555 cases and 3182 controls for Asn312Asp SNP was performed to address this association. Meanwhile, odds ratio (OR and 95% confidence intervals (CIs were used to evaluate this relationship. Statistical analysis was performed with STATA10.0. No significant association was found between XPD Gln751Lys SNP and PCa risk. On the other hand, in subgroup analysis based on ethnicity, associations were observed in Asian (eg. Asn vs. Asp: OR = 1.34, 95%CI = 1.16-1.55; Asn/Asn+Asn/Asp vs. Asp/Asp: OR = 1.23, 95%CI = 1.07-1.42 and African (eg. Asn vs. Asp: OR = 1.31, 95%CI = 1.01-1.70; Asn/Asn vs. Asp/Asp: OR = 1.71, 95%CI = 1.03-7.10 populations for Asn312Asp SNP. Moreover, similar associations were detected in hospital-based controls studies; the frequency of Asn/Asn genotype in early stage of PCa men was poorly higher than those in advanced stage of PCa men (OR = 1.45, 95%CI = 1.00-2.11. CONCLUSION/SIGNIFICANCE: Our investigations demonstrate that XPD Asn312Asp SNP not the Gln751Lys SNP, might poorly increase PCa risk in Asians and Africans, moreover, this SNPs may associate with the tumor stage of PCa. Further studies based on larger sample size and gene-environment interactions should be conducted to determine the role of XPD gene polymorphisms in PCa risk.

Roles of PucR, G1nR, and TnrA in regulating expression of the Bacillus subtilis ure p3 promoter

DEFF Research Database (Denmark)

Brandenburg, J.L.; Wray, L.V.; Beier, Lars

2002-01-01

consensus GlnR/TnrA binding sites located in the ure P3 promoter region were shown to be required for negative regulation by GlnR. Mutational analysis indicates that a cooperative interaction occurs between GlnR dimers bound at these two sites. B. subtilis is the first example where urease expression...

Identification of potentially hazardous human gene products in GMO risk assessment.

Science.gov (United States)

Bergmans, Hans; Logie, Colin; Van Maanen, Kees; Hermsen, Harm; Meredyth, Michelle; Van Der Vlugt, Cécile

2008-01-01

Genetically modified organisms (GMOs), e.g. viral vectors, could threaten the environment if by their release they spread hazardous gene products. Even in contained use, to prevent adverse consequences, viral vectors carrying genes from mammals or humans should be especially scrutinized as to whether gene products that they synthesize could be hazardous in their new context. Examples of such potentially hazardous gene products (PHGPs) are: protein toxins, products of dominant alleles that have a role in hereditary diseases, gene products and sequences involved in genome rearrangements, gene products involved in immunomodulation or with an endocrine function, gene products involved in apoptosis, activated proto-oncogenes. For contained use of a GMO that carries a construct encoding a PHGP, the precautionary principle dictates that safety measures should be applied on a "worst case" basis, until the risks of the specific case have been assessed. The potential hazard of cloned genes can be estimated before empirical data on the actual GMO become available. Preliminary data may be used to focus hazard identification and risk assessment. Both predictive and empirical data may also help to identify what further information is needed to assess the risk of the GMO. A two-step approach, whereby a PHGP is evaluated for its conceptual dangers, then checked by data bank searches, is delineated here.

A novel 5-enolpyruvylshikimate-3-phosphate synthase from Rahnella aquatilis with significantly reduced glyphosate sensitivity.

Directory of Open Access Journals (Sweden)

Ri-He Peng

Full Text Available The 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS; EC 2.5.1.19 is a key enzyme in the shikimate pathway for the production of aromatic amino acids and chorismate-derived secondary metabolites in plants, fungi, and microorganisms. It is also the target of the broad-spectrum herbicide glyphosate. Natural glyphosate resistance is generally thought to occur within microorganisms in a strong selective pressure condition. Rahnella aquatilis strain GR20, an antagonist against pathogenic agrobacterial strains of grape crown gall, was isolated from the rhizosphere of grape in glyphosate-contaminated vineyards. A novel gene encoding EPSPS was identified from the isolated bacterium by complementation of an Escherichia coli auxotrophic aroA mutant. The EPSPS, named AroA(R. aquatilis, was expressed and purified from E. coli, and key kinetic values were determined. The full-length enzyme exhibited higher tolerance to glyphosate than the E. coli EPSPS (AroA(E. coli, while retaining high affinity for the substrate phosphoenolpyruvate. Transgenic plants of AroA(R. aquatilis were also observed to be more resistant to glyphosate at a concentration of 5 mM than that of AroA(E. coli. To probe the sites contributing to increased tolerance to glyphosate, mutant R. aquatilis EPSPS enzymes were produced with the c-strand of subdomain 3 and the f-strand of subdomain 5 (Thr38Lys, Arg40Val, Arg222Gln, Ser224Val, Ile225Val, and Gln226Lys substituted by the corresponding region of the E. coli EPSPS. The mutant enzyme exhibited greater sensitivity to glyphosate than the wild type R. aquatilis EPSPS with little change of affinity for its first substrate, shikimate-3-phosphate (S3P and phosphoenolpyruvate (PEP. The effect of the residues on subdomain 5 on glyphosate resistance was more obvious.

Role of the 2 adenine (g.11293_11294insAA) insertion polymorphism in the 3' untranslated region of the factor VII (FVII) gene: molecular characterization of a patient with severe FVII deficiency.

Science.gov (United States)

Peyvandi, F; Garagiola, I; Palla, R; Marziliano, N; Mannucci, P M

2005-11-01

Polymorphic variants in the gene encoding factor VII (F7) affect the plasma levels of this coagulation protein and modify the clinical phenotype of FVII deficiency in some patients. In this study we report the in vitro functional analysis of a novel polymorphic variant located in the 3' untranslated region of F7: g.11293_11294insAA. To determine whether this variant regulates FVII expression, we initially compared an expression vector containing FVII cDNA with g.11293_11294insAA with the FVII wild-type (WT) construct. The kinetics of mRNA production showed that the insertion decreases the steady-state FVII mRNA levels. To assess whether the insertion influences the phenotype of FVII-deficient patients, we evaluated its effect on the expression of FVII in a patient with severe FVII deficiency (undetectable FVII activity and antigen) carrying two additional homozygous missense variations (p.Arg277Cys and p.Arg353Gln). The two substitutions alone reduced the expression of FVII activity and antigen in vitro, but with the insertion polymorphism in our expression vector the patient's phenotype of undetectable plasma FVII was recapitulated. The insertion polymorphism in the 3' untranslated region of F7 is another modifier of FVII expression that might explain the poor genotype-phenotype correlation in some FVII-deficient patients. Copyright 2005 Wiley-Liss, Inc.

A two-cassette reporter system for assessing target gene translation and target gene product inclusion body formation

DEFF Research Database (Denmark)

2016-01-01

The present invention relates to a dual cassette reporter system capable of assessing target gene translation and target gene product folding. The present invention further relates to vectors and host cells comprising the dual cassette reporter system. In addition the invention relates to the use...... of the dual cassette reporter system for assessing target gene translation and target gene product folding....

Increasing cocoa butter-like lipid production of Saccharomyces cerevisiae by expression of selected cocoa genes

DEFF Research Database (Denmark)

Wei, Yongjun; Gossing, Michael; Bergenholm, David

2017-01-01

for CB biosynthesis from the cocoa genome using a phylogenetic analysis approach. By expressing the selected cocoa genes in S. cerevisiae, we successfully increased total fatty acid production, TAG production and CBL production in some S. cerevisiae strains. The relative CBL content in three yeast...... higher level of CBL compared with the control strain. In summary, CBL production by S. cerevisiae were increased through expressing selected cocoa genes potentially involved in CB biosynthesis.......Cocoa butter (CB) extracted from cocoa beans mainly consists of three different kinds of triacylglycerols (TAGs), 1,3-dipalmitoyl-2-oleoyl-glycerol (POP, C16:0-C18:1-C16:0), 1-palmitoyl-3-stearoyl-2-oleoyl-glycerol(POS,C16:0C18:1-C18:0) and 1,3-distearoyl-2-oleoyl-glycerol (SOS, C18:0-C18:1-C18...

Role of β/δ101Gln in Regulating the Effect of Temperature and Allosteric Effectors on Oxygen Affinity in Woolly Mammoth Hemoglobin†

Science.gov (United States)

Yuan, Yue; Byrd, Catherine; Shen, Tong-Jian; Simplaceanu, Virgil; Tam, Tsuey Chyi S.; Ho, Chien

2013-01-01

The oxygen affinity of woolly mammoth hemoglobin (rHb WM) is less affected by temperature change than that of Asian elephant hemoglobin (rHb AE) or human adult hemoglobin (Hb A). We report here a biochemical-biophysical study of Hb A, rHb AE, rHb WM and three rHb WM mutants with amino acid substitutions at β/δ101 (β/δ101Gln→Glu, Lys, or Asp) plus a double and a triple mutant, designed to clarify the role of the β/δ101 residue. The β/δ101Gln residue is important for responding to allosteric effectors, such as phosphate, inositol hexaphosphate (IHP), and chloride. The rHb WM mutants studied generally have higher affinity for oxygen under various conditions of pH, temperature, and salt concentration, and in the presence or absence of organic phosphate, than do rHb WM, rHb AE and Hb A. Titrations for the O2 affinity of these mutant rHbs as a function of chloride concentration indicate a lower heterotopic effect of this anion due to the replacement of β/δ101Gln in rHb WM. The alkaline Bohr effect of rHb WM and its mutants is reduced by 20–50% compared to that of Hb A and is independent of changes in temperature, in contrast to what has been observed in the hemoglobins of most mammalian species, including human. The results of our study on the temperature dependence of the O2 affinity of rHb WM and its mutant rHbs illustrate the important role of β/δ101Gln in regulating the functional properties of these hemoglobins. PMID:24228693

Relationship between polymorphisms of nucleotide excision repair genes and oral cancer risk in Taiwan: evidence for modification of smoking habit.

Science.gov (United States)

Bau, Da-Tian; Tsai, Ming-Hsui; Huang, Chih-Yang; Lee, Cheng-Chun; Tseng, Hsien-Chang; Lo, Yen-Li; Tsai, Yuhsin; Tsai, Fuu-Jen

2007-12-31

Inherited polymorphisms in DNA repair genes may be associated with differences in the repair capacity and contribute to individual's susceptibility to smoking-related cancers. Both XPA and XPD encode proteins that are part of the nucleotide excision repair (NER) pathway. In a hospital-based case-control study, we have investigated the influence of XPA A-23G and XPD Lys751Gln polymorphisms on oral cancer risk in a Taiwanese population. In total, 154 patients with oral cancer, and 105 age-matched controls recruited from the Chinese Medical Hospital in Central Taiwan were genotyped. No significant association was found between the heterozygous variant allele (AG), the homozygous variant allele (AA) at XPA A-23G, the heterozygous variant allele (AC), the homozygous variant allele (CC) at XPD Lys751Gln, and oral cancer risk. There was no significant joint effect of XPA A-23G and XPD Lys751Gln on oral cancer risk either. Since XPA and XPD are both NER genes, which are very important in removing tobacco-induced DNA adducts, further stratified analyses of both genotype and smoking habit were performed. We found a synergistic effect of variant genotypes of both XPA and XPD, and smoking status on oral cancer risk. Our results suggest that the genetic polymorphisms are modified by environmental carcinogen exposure status, and combined analyses of both genotype and personal habit record are a better access to know the development of oral cancer and useful for primary prevention and early intervention.

[Brd3 promotes IL-6 production via enhancing acetylase CBP recruitment and histone 3 acetylation within IL6 promoter].

Science.gov (United States)

Ren, Wenhui; Sun, Donghao; Wang, Chunmei; Li, Nan

2016-10-01

Objective To investigate the role of bromodomain containing 3 (Brd3) in LPS-triggered interleukin-6 (IL-6) production in macrophages and the underlying mechanism. Methods CRISPR-Cas9 technology was used to screen an RAW264.7 cell line with Brd3 knockout (Brd3 -/- ). The Brd3 -/- cells were used as an experimental group, and the parential cells expressing wide-type Brd3 as a control group. The IL-6 level in cell culture supernatant was detected by ELISA after 100 ng/mL LPS challenging. Effect of Brd3 knockout on the expression and activation of signal pathways involved in IL-6 expression, including the NF-κB and mitogen-activated protein kinase (MAPK) pathways were examined by Western blot analysis. Chromatin immunoprecipitation (ChIP) assay was used to evaluate the recruitment of acetylase CREB-binding protein (CBP) to IL6 gene promoter and the acetylation level of histone 3 within IL6 gene promoter. Results LPS treatment significantly downregulated Brd3 expression in mouse peritoneal macrophages. LPS-induced production of IL-6 was significantly inhibited in Brd3 -/- macrophages. The expressions and activation of signal molecules within NF-κB and MAPK pathways were barely affected. Brd3 knockout significantly decreased the recruitment of acetylase CBP to IL6 gene promoter, and the acetylation level of histone3 within IL6 gene promoter was also repressed. Conclusion Brd3 promotes LPS-triggered IL-6 production via promoting the recruitment of CBP to IL6 promoter and enhancing the acetylation level of histone 3 within IL6 promoter.

Isolation and characterization of PEP3, a gene required for vacuolar biogenesis in Saccharomyces cerevisiae.

OpenAIRE

Preston, R A; Manolson, M F; Becherer, K; Weidenhammer, E; Kirkpatrick, D; Wright, R; Jones, E W

1991-01-01

The Saccharomyces cerevisiae PEP3 gene was cloned from a wild-type genomic library by complementation of the carboxypeptidase Y deficiency in a pep3-12 strain. Subclone complementation results localized the PEP3 gene to a 3.8-kb DNA fragment. The DNA sequence of the fragment was determined; a 2,754-bp open reading frame predicts that the PEP3 gene product is a hydrophilic, 107-kDa protein that has no significant similarity to any known protein. The PEP3 predicted protein has a zinc finger (CX...

Eight novel F13A1 gene missense mutations in patients with mild FXIII deficiency: in silico analysis suggests changes in FXIII-A subunit structure/function.

Science.gov (United States)

Biswas, Arijit; Ivaskevicius, Vytautas; Thomas, Anne; Varvenne, Michael; Brand, Brigitte; Rott, Hannelore; Haussels, Iris; Ruehl, Heiko; Scholz, Ute; Klamroth, Robert; Oldenburg, Johannes

2014-10-01

Mild FXIII deficiency is an under-diagnosed disorder because the carriers of this deficiency are often asymptomatic and reveal a phenotype only under special circumstances like surgery or induced trauma. Mutational reports from this type of deficiency have been rare. In this study, we present the phenotypic and genotypic data of nine patients showing mild FXIII-A deficiency caused by eight novel heterozygous missense mutations (Pro166Leu, Arg171Gln, His342Tyr, Gln415Arg, Leu529Pro, Gln601Lys, Arg703Gln and Arg715Gly) in the F13A1 gene. None of these variants were seen in 200 healthy controls. In silico structural analysis of the local wild-type protein structures (activated and non-activated) from X-ray crystallographic models downloaded from the protein databank identified potential structural/functional effects for the identified mutations. The missense mutations in the core domain are suggested to be directly influencing the catalytic triad. Mutations on other domains might influence other critical factors such as activation peptide cleavage or the barrel domain integrity. In vitro expression and subsequent biochemical studies in the future will be able to confirm the pathophysiological mechanisms proposed for the mutations in this article.

A male with unilateral microphthalmia reveals a role for TMX3 in eye development

DEFF Research Database (Denmark)

Chao, Ryan; Nevin, Linda; Agarwal, Pooja

2010-01-01

Anophthalmia and microphthalmia are important birth defects, but their pathogenesis remains incompletely understood. We studied a patient with severe unilateral microphthalmia who had a 2.7 Mb deletion at chromosome 18q22.1 that was inherited from his mother. In-situ hybridization showed that one...... of the deleted genes, TMX3, was expressed in the retinal neuroepithelium and lens epithelium in the developing murine eye. We re-sequenced TMX3 in 162 patients with anophthalmia or microphthalmia, and found two missense substitutions in unrelated patients: c.116G>A, predicting p.Arg39Gln, in a male...

Epstein-Barr Virus BKRF4 Gene Product Is Required for Efficient Progeny Production.

Science.gov (United States)

Masud, H M Abdullah Al; Watanabe, Takahiro; Yoshida, Masahiro; Sato, Yoshitaka; Goshima, Fumi; Kimura, Hiroshi; Murata, Takayuki

2017-12-01

Epstein-Barr virus (EBV), a member of human gammaherpesvirus, infects mainly B cells. EBV has two alternative life cycles, latent and lytic, and is reactivated occasionally from the latent stage to the lytic cycle. To combat EBV-associated disorders, understanding the molecular mechanisms of the EBV lytic replication cycle is also important. Here, we focused on an EBV lytic gene, BKRF4. Using our anti-BKRF4 antibody, we revealed that the BKRF4 gene product is expressed during the lytic cycle with late kinetics. To characterize the role of BKRF4, we constructed BKRF4-knockout mutants using the bacterial artificial chromosome (BAC) and CRISPR/Cas9 systems. Although disruption of the BKRF4 gene had almost no effect on viral protein expression and DNA synthesis, it significantly decreased progeny virion levels in HEK293 and Akata cells. Furthermore, we show that BKRF4 is involved not only in production of progeny virions but also in increasing the infectivity of the virus particles. Immunoprecipitation assays revealed that BKRF4 interacted with a virion protein, BGLF2. We showed that the C-terminal region of BKRF4 was critical for this interaction and for efficient progeny production. Immunofluorescence analysis revealed that BKRF4 partially colocalized with BGLF2 in the nucleus and perinuclear region. Finally, we showed that BKRF4 is a phosphorylated, possible tegument protein and that the EBV protein kinase BGLF4 may be important for this phosphorylation. Taken together, our data suggest that BKRF4 is involved in the production of infectious virions. IMPORTANCE Although the latent genes of EBV have been studied extensively, the lytic genes are less well characterized. This study focused on one such lytic gene, BKRF4, which is conserved only among gammaherpesviruses (ORF45 of Kaposi's sarcoma-associated herpesvirus or murine herpesvirus 68). After preparing the BKRF4 knockout virus using B95-8 EBV-BAC, we demonstrated that the BKRF4 gene was involved in infectious

Ozone-induced gene expression occurs via ethylene-dependent and -independent signalling.

Science.gov (United States)

Grimmig, Bernhard; Gonzalez-Perez, Maria N; Leubner-Metzger, Gerhard; Vögeli-Lange, Regina; Meins, Fred; Hain, Rüdiger; Penuelas, Josep; Heidenreich, Bernd; Langebartels, Christian; Ernst, Dieter; Sandermann, Heinrich

2003-03-01

Recent studies suggest that ethylene is involved in signalling ozone-induced gene expression. We show here that application of ozone increased glucuronidase (GUS) expression of chimeric reporter genes regulated by the promoters of the tobacco class I beta-1,3-glucanases (GLB and Gln2) and the grapevine resveratrol synthase (Vst1) genes in transgenic tobacco leaves. 5'-deletion analysis of the class I beta-1,3-glucanase promoter revealed that ozone-induced gene regulation is mainly mediated by the distal enhancer region containing the positively acting ethylene-responsive element (ERE). In addition, application of 1-methylcyclopropene (1-MCP), an inhibitor of ethylene action, blocked ozone-induced class I beta-1,3-glucanase promoter activity. Enhancer activity and ethylene-responsiveness depended on the integrity of the GCC boxes, cis-acting elements present in the ERE of the class I beta-1,3-glucanase and the basic-type pathogenesis-related PR-1 protein (PRB-1b) gene promoters. The minimal PRB-1b promoter containing only the ERE with intact GCC boxes, was sufficient to confer 10-fold ozone inducibility to a GUS-reporter gene, while a substitution mutation in the GCC box abolished ozone responsiveness. The ERE region of the class I beta-1,3-glucanase promoter containing two intact GCC boxes confered strong ozone inducibility to a minimal cauliflower mosaic virus (CaMV) 35S RNA promoter, whereas two single-base substitution in the GCC boxes resulted in a complete loss of ozone inducibility. Taken together, these datastrongly suggest that ethylene is signalling ozone-induced expression of class I beta-l,3-glucanase and PRB-1b genes. Promoter analysis of the stilbene synthase Vst1 gene unravelled different regions for ozone and ethylene-responsiveness. Application of 1-MCP blocked ethylene-induced Vst1 induction, but ozone induction was not affected. This shows that ozone-induced gene expression occurs via at least two different signalling mechanisms and suggests an

A male with unilateral microphthalmia reveals a role for TMX3 in eye development.

Directory of Open Access Journals (Sweden)

Ryan Chao

Full Text Available Anophthalmia and microphthalmia are important birth defects, but their pathogenesis remains incompletely understood. We studied a patient with severe unilateral microphthalmia who had a 2.7 Mb deletion at chromosome 18q22.1 that was inherited from his mother. In-situ hybridization showed that one of the deleted genes, TMX3, was expressed in the retinal neuroepithelium and lens epithelium in the developing murine eye. We re-sequenced TMX3 in 162 patients with anophthalmia or microphthalmia, and found two missense substitutions in unrelated patients: c.116G>A, predicting p.Arg39Gln, in a male with unilateral microphthalmia and retinal coloboma, and c.322G>A, predicting p.Asp108Asn, in a female with unilateral microphthalmia and severe micrognathia. We used two antisense morpholinos targeted against the zebrafish TMX3 orthologue, zgc:110025, to examine the effects of reduced gene expression in eye development. We noted that the morphant larvae resulting from both morpholinos had significantly smaller eye sizes and reduced labeling with islet-1 antibody directed against retinal ganglion cells at 2 days post fertilization. Co-injection of human wild type TMX3 mRNA rescued the small eye phenotype obtained with both morpholinos, whereas co-injection of human TMX3(p.Arg39Gln mutant mRNA, analogous to the mutation in the patient with microphthalmia and coloboma, did not rescue the small eye phenotype. Our results show that haploinsufficiency for TMX3 results in a small eye phenotype and represents a novel genetic cause of microphthalmia and coloboma. Future experiments to determine if other thioredoxins are important in eye morphogenesis and to clarify the mechanism of function of TMX3 in eye development are warranted.

Partial rescue of in vivo insulin signalling in skeletal muscle by impaired insulin clearance in heterozygous carriers of a mutation in the insulin receptor gene

DEFF Research Database (Denmark)

Højlund, K.; Wojtaszewski, Jørgen; Birk, Jesper Bratz

2006-01-01

AIMS/HYPOTHESIS: Recently we reported the coexistence of postprandial hypoglycaemia and moderate insulin resistance in heterozygous carriers of the Arg1174Gln mutation in the insulin receptor gene (INSR). Controlled studies of in vivo insulin signalling in humans with mutant INSR are unavailable,...

The BAG3 gene variants in Polish patients with dilated cardiomyopathy: four novel mutations and a genotype-phenotype correlation.

Science.gov (United States)

Franaszczyk, Maria; Bilinska, Zofia T; Sobieszczańska-Małek, Małgorzata; Michalak, Ewa; Sleszycka, Justyna; Sioma, Agnieszka; Małek, Łukasz A; Kaczmarska, Dorota; Walczak, Ewa; Włodarski, Paweł; Hutnik, Łukasz; Milanowska, Blanka; Dzielinska, Zofia; Religa, Grzegorz; Grzybowski, Jacek; Zieliński, Tomasz; Ploski, Rafal

2014-07-09

BAG3 gene mutations have been recently implicated as a novel cause of dilated cardiomyopathy (DCM). Our aim was to evaluate the prevalence of BAG3 mutations in Polish patients with DCM and to search for genotype-phenotype correlations. We studied 90 unrelated probands by direct sequencing of BAG3 exons and splice sites. Large deletions/insertions were screened for by quantitative real time polymerase chain reaction (qPCR). We found 5 different mutations in 6 probands and a total of 21 mutations among their relatives: the known p.Glu455Lys mutation (2 families), 4 novel mutations: p.Gln353ArgfsX10 (c.1055delC), p.Gly379AlafsX45 (c.1135delG), p.Tyr451X (c.1353C>A) and a large deletion of 17,990 bp removing BAG3 exons 3-4. Analysis of mutation positive relatives of the probands from this study pooled with those previously reported showed higher DCM prevalence among those with missense vs. truncating mutations (OR = 8.33, P = 0.0058) as well as a difference in age at disease onset between the former and the latter in Kaplan-Meier survival analysis (P = 0.006). Clinical data from our study suggested that in BAG3 mutation carriers acute onset DCM with hemodynamic compromise may be triggered by infection. BAG3 point mutations and large deletions are relatively frequent cause of DCM. Delayed DCM onset associated with truncating vs. non-truncating mutations may be important for genetic counseling.

Id-1 gene and gene products as therapeutic targets for treatment of breast cancer and other types of carcinoma

Science.gov (United States)

Desprez, Pierre-Yves; Campisi, Judith

2014-08-19

A method for treatment of breast cancer and other types of cancer. The method comprises targeting and modulating Id-1 gene expression, if any, for the Id-1 gene, or gene products in breast or other epithelial cancers in a patient by delivering products that modulate Id-1 gene expression. When expressed, Id-1 gene is a prognostic indicator that cancer cells are invasive and metastatic.

Cadherin-related family member 3, a childhood asthma susceptibility gene product, mediates rhinovirus C binding and replication.

Science.gov (United States)

Bochkov, Yury A; Watters, Kelly; Ashraf, Shamaila; Griggs, Theodor F; Devries, Mark K; Jackson, Daniel J; Palmenberg, Ann C; Gern, James E

2015-04-28

Members of rhinovirus C (RV-C) species are more likely to cause wheezing illnesses and asthma exacerbations compared with other rhinoviruses. The cellular receptor for these viruses was heretofore unknown. We report here that expression of human cadherin-related family member 3 (CDHR3) enables the cells normally unsusceptible to RV-C infection to support both virus binding and replication. A coding single nucleotide polymorphism (rs6967330, C529Y) was previously linked to greater cell-surface expression of CDHR3 protein, and an increased risk of wheezing illnesses and hospitalizations for childhood asthma. Compared with wild-type CDHR3, cells transfected with the CDHR3-Y529 variant had about 10-fold increases in RV-C binding and progeny yields. We developed a transduced HeLa cell line (HeLa-E8) stably expressing CDHR3-Y529 that supports RV-C propagation in vitro. Modeling of CDHR3 structure identified potential binding sites that could impact the virus surface in regions that are highly conserved among all RV-C types. Our findings identify that the asthma susceptibility gene product CDHR3 mediates RV-C entry into host cells, and suggest that rs6967330 mutation could be a risk factor for RV-C wheezing illnesses.

Increased enzyme production under liquid culture conditions in the industrial fungus Aspergillus oryzae by disruption of the genes encoding cell wall α-1,3-glucan synthase.

Science.gov (United States)

Miyazawa, Ken; Yoshimi, Akira; Zhang, Silai; Sano, Motoaki; Nakayama, Mayumi; Gomi, Katsuya; Abe, Keietsu

2016-09-01

Under liquid culture conditions, the hyphae of filamentous fungi aggregate to form pellets, which reduces cell density and fermentation productivity. Previously, we found that loss of α-1,3-glucan in the cell wall of the fungus Aspergillus nidulans increased hyphal dispersion. Therefore, here we constructed a mutant of the industrial fungus A. oryzae in which the three genes encoding α-1,3-glucan synthase were disrupted (tripleΔ). Although the hyphae of the tripleΔ mutant were not fully dispersed, the mutant strain did form smaller pellets than the wild-type strain. We next examined enzyme productivity under liquid culture conditions by transforming the cutinase-encoding gene cutL1 into A. oryzae wild-type and the tripleΔ mutant (i.e. wild-type-cutL1, tripleΔ-cutL1). A. oryzae tripleΔ-cutL1 formed smaller hyphal pellets and showed both greater biomass and increased CutL1 productivity compared with wild-type-cutL1, which might be attributable to a decrease in the number of tripleΔ-cutL1 cells under anaerobic conditions.

Interaction of GlnK with the GAF domain of Herbaspirillum seropedicae NifA mediates NH₄⁺-regulation.

Science.gov (United States)

Oliveira, Marco A S; Aquino, Bruno; Bonatto, Ana Claudia; Huergo, Luciano F; Chubatsu, Leda S; Pedrosa, Fábio O; Souza, Emanuel M; Dixon, Ray; Monteiro, Rose A

2012-04-01

Nitrogen fixation in Herbaspirillum seropedicae is transcriptionally regulated by NifA, a σ(54) transcriptional activator with three structural domains: an N-terminal GAF domain, a catalytic AAA+ domain and a C-terminal DNA-binding domain. NifA is only active in H. seropedicae when cultures are grown in the absence of fixed nitrogen and at low oxygen tensions. There is evidence that the inactivation of NifA in response to fixed nitrogen is mediated by the regulatory GAF domain. However, the mechanism of NifA repression by the GAF domain, as well as the transduction of nitrogen status to NifA, is not understood. In order to study the regulation of NifA activity by fixed nitrogen independently of oxygen regulation, we constructed a chimeric protein containing the GAF domain of H. seropedicae NifA fused to the AAA+ and C-terminal domains of Azotobacter vinelandii NifA. This chimeric protein (NifAQ1) lacks the cysteine motif found in oxygen sensitive NifA proteins and is not oxygen responsive in vivo. Our results demonstrate that NifAQ1 responds to fixed nitrogen and requires GlnK protein for activity, a behavior similar to H. seropedicae NifA. In addition, protein footprinting analysis indicates that this response probably involves a protein-protein contact between the GAF domain and the GlnK protein. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

Systematic metabolic engineering of Methylomicrobium alcaliphilum 20Z for 2,3-butanediol production from methane.

Science.gov (United States)

Nguyen, Anh Duc; Hwang, In Yeub; Lee, Ok Kyung; Kim, Donghyuk; Kalyuzhnaya, Marina G; Mariyana, Rina; Hadiyati, Susila; Kim, Min Sik; Lee, Eun Yeol

2018-04-16

Methane is considered a next-generation feedstock, and methanotrophic cell-based biorefinery is attractive for production of a variety of high-value compounds from methane. In this work, we have metabolically engineered Methylomicrobium alcaliphilum 20Z for 2,3-butanediol (2,3-BDO) production from methane. The engineered strain 20Z/pBudK.p, harboring the 2,3-BDO synthesis gene cluster (budABC) from Klebsiella pneumoniae, accumulated 2,3-BDO in methane-fed shake flask cultures with a titer of 35.66 mg/L. Expression of the most efficient gene cluster was optimized using selection of promoters, translation initiation rates (TIR), and the combination of 2,3-BDO synthesis genes from different sources. A higher 2,3-BDO titer of 57.7 mg/L was measured in the 20Z/pNBM-Re strain with budA of K. pneumoniae and budB of Bacillus subtilis under the control of the Tac promoter. The genome-scale metabolic network reconstruction of M. alcaliphilum 20Z enabled in silico gene knockout predictions using an evolutionary programming method to couple growth and 2,3-BDO production. The ldh, ack, and mdh genes in M. alcaliphilum 20Z were identified as potential knockout targets. Pursuing these targets, a triple-mutant strain ∆ldh ∆ack ∆mdh was constructed, resulting in a further increase of the 2,3-BDO titer to 68.8 mg/L. The productivity of this optimized strain was then tested in a fed-batch stirred tank bioreactor, where final product concentrations of up to 86.2 mg/L with a yield of 0.0318 g-(2,3-BDO) /g-CH 4 were obtained under O 2 -limited conditions. This study first demonstrates the strategy of in silico simulation-guided metabolic engineering and represents a proof-of-concept for the production of value-added compounds using systematic approaches from engineered methanotrophs. Copyright © 2018 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

Risk of colorectal cancer for people with a mutation in both a MUTYH and a DNA mismatch repair gene

Science.gov (United States)

Win, Aung Ko; Reece, Jeanette C.; Buchanan, Daniel D.; Clendenning, Mark; Young, Joanne P.; Cleary, Sean P.; Kim, Hyeja; Cotterchio, Michelle; Dowty, James G.; MacInnis, Robert J.; Tucker, Katherine M.; Winship, Ingrid M.; Macrae, Finlay A.; Burnett, Terrilea; Le Marchand, Loïc; Casey, Graham; Haile, Robert W.; Newcomb, Polly A.; Thibodeau, Stephen N.; Lindor, Noralane M.; Hopper, John L.; Gallinger, Steven; Jenkins, Mark A.

2015-01-01

The base excision repair protein, MUTYH, functionally interacts with the DNA mismatch repair (MMR) system. As genetic testing moves from testing one gene at a time, to gene panel and whole exome next generation sequencing approaches, understanding the risk associated with co-existence of germline mutations in these genes will be important for clinical interpretation and management. From the Colon Cancer Family Registry, we identified 10 carriers who had both a MUTYH mutation (6 with c.1187G>A p.(Gly396Asp), 3 with c.821G>A p.(Arg274Gln), and 1 with c.536A>G p.(Tyr179Cys)) and a MMR gene mutation (3 in MLH1, 6 in MSH2, and 1 in PMS2), 375 carriers of a single (monoallelic) MUTYH mutation alone, and 469 carriers of a MMR gene mutation alone. Of the 10 carriers of both gene mutations, 8 were diagnosed with colorectal cancer. Using a weighted cohort analysis, we estimated that risk of colorectal cancer for carriers of both a MUTYH and a MMR gene mutation was substantially higher than that for carriers of a MUTYH mutation alone [hazard ratio (HR) 21.5, 95 % confidence interval (CI) 9.19–50.1; p colorectal cancer for carriers of a MMR gene mutation alone. Our finding suggests MUTYH mutation testing in MMR gene mutation carriers is not clinically informative. PMID:26202870

[Analysis of the NDP gene in a Chinese family with X-linked recessive Norrie disease].

Science.gov (United States)

Mei, Libin; Huang, Yanru; Pan, Qian; Liang, Desheng; Wu, Lingqian

2015-05-01

The purpose of the current research was to investigate the NDP (Norrie disease protein) gene in one Chinese family with Norrie disease (ND) and to characterize the related clinical features. Clinical data of the proband and his family members were collected. Complete ophthalmic examinations were carried out on the proband. Genomic DNA was extracted from peripheral blood leukocytes of 35 family members. Molecular analysis of the NDP gene was performed by polymerase chain reaction and direct sequencing of all exons and flanking regions. A hemizygous NDP missense mutation c.362G > A (p.Arg121Gln) in exon 3 was identified in the affected members, but not in any of the unaffected family individuals. The missense mutation c.362G > A in NDP is responsible for the Norrie disease in this family. This discovery will help provide the family members with accurate and reliable genetic counseling and prenatal diagnosis.

Roles of Beta2- and Beta3-Adrenoceptor Polymorphisms in Hypertension and Metabolic Syndrome

Directory of Open Access Journals (Sweden)

Kazuko Masuo

2010-01-01

gain, and the sympathetic nervous system participates in regulating energy balance through thermogenesis. The thermogenic effects of catecholamines in obesity have been mainly mediated via the 2- and 3-adrenergic receptors in humans. Further, 2-adrenoceptors importantly influence vascular reactivity and may regulate blood pressure. Genetic polymorphistns of the -adrenoceptor gene have been shown to alter the function of several adrenoceptor subtypes and thus to modify the response to catecholamine. 2-adrenoceptor polymorphisms (Arg16Gly, Gln27Glu, and Thr164Ile have been studied in relation to hypertension. Genetic variations in the 3-adrenoceptor (i.e. Try64Arg variant are also associated with both obesity and hypertension. However, the precise relationships of the polymorphisms of 2- and 3-adrenoceptor genes with sympathetic nervous system activity, hypertension, and metabolic syndrome have not been fully clarified. This paper will discuss the current topics involving the influence of the sympathetic nervous system and 2- and 3- adrenoceptor polymorphisms in hypertension and metabolic syndrome.

A novel autolysis system controlled by magnesium and its application to poly (3-hydroxypropionate) production in engineered Escherichia coli.

Science.gov (United States)

Tamekou Lacmata, Stephen; Yao, Lan; Xian, Mo; Liu, Hui; Kuiate, Jules-Roger; Liu, Huizhou; Feng, Xinjun; Zhao, Guang

2017-09-03

The release of intracellular products, especially polyhydroxyalkanoates, is still a great challenge in industry. To solve this bottleneck, a novel autolysis system strictly controlled with magnesium was constructed and applied to poly(3-hydroxypropionate) production in engineered Escherichia coli. The autolysis system was constructed by inserting the 5'untranslated region (5'UTR) behind promoter PmgtA with lysis genes (S, R, and Rz, from E. coli) overexpressed. The autolysis system functioned well (lysis efficiency of more than 90%) in the P3HP producer with double plasmids containing lysis genes and P3HP biosynthesis genes, whereas the P3HP production was reduced due to plasmid losses. After the autolysis genes and P3HP biosynthesis genes were integrated into one plasmid, the P3HP content of 72.7% (2.4 times of the control) and the plasmid stability of 79.8 ± 3.1% were achieved in strain Q2646 with promoter PmgtA-UTR. However, the strain Q2647 with promoter PmgtA could not accumulate P3HP because of rapid cell lysis. The novel autolysis system activated in Mg 2+ -depleted conditions proves to be feasible for polyhydroxyalkanoates production, which may have great application potential for other intracellular products.

[Identification of new genes that affect [PSI^(+)] prion toxicity in Saccharomyces cerevisiae yeast].

Science.gov (United States)

Matveenko, A G; Belousov, M V; Bondarev, S A; Moskalenko, S E; Zhouravleva, G A

2016-01-01

Translation termination is an important step in gene expression. Its correct processing is governed by eRF1 (Sup45) and eRF3 (Sup35) proteins. In Saccharomyces cerevisiae, mutations in the corresponding genes, as well as Sup35 aggregation in [PSI^(+)] cells that propagate the prion form of Sup35 lead to inaccurate stop codon recognition and, consequently, nonsense suppression. The presence of stronger prion variants results in the more efficient suppression of nonsense mutations. Previously, we proposed a synthetic lethality test that enables the identification of genes that may influence either translation termination factors or [PSI^(+)] manifestation. This is based on the fact that the combination of sup45 mutations with the strong [PSI^(+)] prion variant in diploids is lethal. In this work, a set of genes that were previously shown to enhance nonsense suppression was analyzed. It was found that ABF1, FKH2, and REB1 overexpression decreased the growth of strains in a prion-dependent manner and, thus, might influence [PSI^(+)] prion toxicity. It was also shown that the synthetic lethality of [PSI^(+)] and sup45 mutations increased with the overexpression of GLN3 and MOT3 that encode Q/N-rich transcription factors. An analysis of the effects of their expression on the transcription of the release factors genes revealed an increase in SUP35 transcription in both cases. Since SUP35 overexpression is known to be toxic in [PSI^(+)] strains, these genes apparently enhance [PSI^(+)] toxicity via the regulation of SUP35 transcription.

Efficient production of antibody Fab fragment by transient gene expression in insect cells.

Science.gov (United States)

Mori, Keita; Hamada, Hirotsugu; Ogawa, Takafumi; Ohmuro-Matsuyama, Yuki; Katsuda, Tomohisa; Yamaji, Hideki

2017-08-01

Transient gene expression allows a rapid production of diverse recombinant proteins in early-stage preclinical and clinical developments of biologics. Insect cells have proven to be an excellent platform for the production of functional recombinant proteins. In the present study, the production of an antibody Fab fragment by transient gene expression in lepidopteran insect cells was investigated. The DNA fragments encoding heavy-chain (Hc; Fd fragment) and light-chain (Lc) genes of an Fab fragment were individually cloned into the plasmid vector pIHAneo, which contained the Bombyx mori actin promoter downstream of the B. mori nucleopolyhedrovirus (BmNPV) IE-1 transactivator and the BmNPV HR3 enhancer for high-level expression. Trichoplusia ni BTI-TN-5B1-4 (High Five) cells were co-transfected with the resultant plasmid vectors using linear polyethyleneimine. When the transfection efficiency was evaluated, a plasmid vector encoding an enhanced green fluorescent protein (EGFP) gene was also co-transfected. Transfection and culture conditions were optimized based on both the flow cytometry of the EGFP expression in transfected cells and the yield of the secreted Fab fragments determined by enzyme-linked immunosorbent assay (ELISA). Under optimal conditions, a yield of approximately 120 mg/L of Fab fragments was achieved in 5 days in a shake-flask culture. Transient gene expression in insect cells may offer a promising approach to the high-throughput production of recombinant proteins. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Ghrelin gene: identification of missense variants and a frameshift mutation in extremely obese children and adolescents and healthy normal weight students.

Science.gov (United States)

Hinney, Anke; Hoch, Anne; Geller, Frank; Schäfer, Helmut; Siegfried, Wolfgang; Goldschmidt, Hanspeter; Remschmidt, Helmut; Hebebrand, Johannes

2002-06-01

Ghrelin induces obesity via central and peripheral mechanisms. Administration of ghrelin leads to increased food intake and decreased fat utilisation in rodents. Ghrelin levels are decreased in obese individuals. Recently, a polymorphism (Arg-51-Gln) within the ghrelin gene (GHRL) was described to be associated with obesity. We screened the GHRL coding region in 215 extremely obese German Children and adolescents (study group 1) and 93 normal weight students (study group 2) by single strand conformation polymorphism analysis (SSCP). We found the two previously described single nucleotide polymorphisms (SNP: Arg-51-Gln and Leu-72-Met) in similar frequencies in study groups 1 and 2 (allele frequencies were: 0.019 and 0.016 for the 51-Gln allele and 0.091 and 0.086 for the 72-Met allele, respectively). Hence, we could not confirm the previous finding. Additionally, two novel variants were identified within the coding region: (1) We detected one healthy normal weight individual with a frameshift mutation (2bp deletion at codon 34). This frameshift mutation affects the coding region of the mature ghrelin. Hence, it is highly likely that the normal weight student is haplo-insufficient for ghrelin. (2) An A to T transversion leads to an amino acid exchange from Gln to Leu at amino acid position 90. The frequency of the 90-Leu allele was significantly higher in the extremely obese children and adolescents (0.063) than in the normal weight students (0.016; nominal p = 0.011). Additionally, we genotyped 134 underweight students and 44 normal weight adults for this SNP. Genotype frequencies were similar in extremely obese children and adolescents, underweight students and normal weight adults (p > 0.8). In conclusion, we identified four sequence variants in the coding region of the ghrelin gene in individuals belonging to different weight extremes. A frameshift mutation was detected in a normal weight individual. None of the variants seem to influence weight regulation.

Cloning and characterization of a gene (UVR3) required for photorepair of 6-4 photoproducts in Arabidopsis thaliana

International Nuclear Information System (INIS)

Nakajima, S.; Sugiyama, M.; Iwai, S.; Hitomi, K.; Otoshi, E.; Kim SangTae; Jiang CaiZhong; Todo, T.; Britt, A.B.; Yamamoto, K.

1998-01-01

UV radiation induces two major classes of pyrimidine dimers: the pyrimidine [6-4] pyrimidone photoproduct (6-4 product) and the cyclobutane pyrimidine dimer (CPD). Many organisms produce enzymes, termed photolyases, that specifically bind to these damage products and split them via a UV-A/blue light-dependent mechanism, thereby reversing the damage. These photolyases are specific for either CPDs or 6-4 products. A gene that expresses a protein with 6-4 photolyase activity in vitro was recently cloned from Drosophila melanogaster and Xenopus laevis. We report here the isolation of a homolog of this gene, cloned on the basis of sequence similarity, from the higher plant Arabidopsis thaliana. This cloned gene produces a protein with 6-4 photolyase activity when expressed in Escherichia coli. We also find that a previously described mutant of Arabidopsis (uvr3) that is defective in photoreactivation of 6-4 products carries a nonsense mutation in this 6-4 photolyase homolog. We have therefore termed this gene UVR3. Although homologs of this gene have previously been shown to produce a functional 6-4 photolyase when expressed in heterologous systems, this is the first demonstration of a requirement for this gene for photoreactivation of 6-4 products in vivo

Abundance and distribution of Macrolide-Lincosamide-Streptogramin resistance genes in an anaerobic-aerobic system treating spiramycin production wastewater.

Science.gov (United States)

Liu, Miaomiao; Ding, Ran; Zhang, Yu; Gao, Yingxin; Tian, Zhe; Zhang, Tong; Yang, Min

2014-10-15

The behaviors of the Macrolide-Lincosamide-Streptogramin (MLS) resistance genes were investigated in an anaerobic-aerobic pilot-scale system treating spiramycin (SPM) production wastewater. After screening fifteen typical MLS resistance genes with different mechanisms using conventional PCR, eight detected genes were determined by quantitative PCR, together with three mobile elements. Aerobic sludge in the pilot system exhibited a total relative abundance of MLS resistance genes (per 16S rRNA gene) 2.5 logs higher than those in control samples collected from sewage and inosine wastewater treatment systems (P resistance genes. However, the total relative gene abundance in anaerobic sludge (4.3 × 10(-1)) was lower than that in aerobic sludge (3.7 × 10(0)) despite of the higher SPM level in anaerobic reactor, showing the advantage of anaerobic treatment in reducing the production of MLS resistance genes. The rRNA methylase genes (erm(B), erm(F), erm(X)) were the most abundant in the aerobic sludge (5.3 × 10(-1)-1.7 × 10(0)), followed by esterase gene ere(A) (1.3 × 10(-1)) and phosphorylase gene mph(B) (5.7 × 10(-2)). In anaerobic sludge, erm(B), erm(F), ere(A), and msr(D) were the major ones (1.2 × 10(-2)-3.2 × 10(-1)). These MLS resistance genes (except for msr(D)) were positively correlated with Class 1 integron (r(2) = 0.74-0.93, P < 0.05), implying the significance of horizontal transfer in their proliferation. Copyright © 2014 Elsevier Ltd. All rights reserved.

Regulation of Cell and Gene Therapy Medicinal Products in Taiwan.

Science.gov (United States)

Lin, Yi-Chu; Wang, Po-Yu; Tsai, Shih-Chih; Lin, Chien-Liang; Tai, Hsuen-Yung; Lo, Chi-Fang; Wu, Shiow-Ing; Chiang, Yu-Mei; Liu, Li-Ling

2015-01-01

Owing to the rapid and mature development of emerging biotechnology in the fields of cell culture, cell preservation, and recombinant DNA technology, more and more cell or gene medicinal therapy products have been approved for marketing, to treat serious diseases which have been challenging to treat with current medical practice or medicine. This chapter will briefly introduce the Taiwan Food and Drug Administration (TFDA) and elaborate regulation of cell and gene therapy medicinal products in Taiwan, including regulatory history evolution, current regulatory framework, application and review procedures, and relevant jurisdictional issues. Under the promise of quality, safety, and efficacy of medicinal products, it is expected the regulation and environment will be more flexible, streamlining the process of the marketing approval of new emerging cell or gene therapy medicinal products and providing diverse treatment options for physicians and patients.

164Ile allele in the beta2-Adrenergic receptor gene is associated with risk of elevated blood pressure in women. The Copenhagen City Heart Study

DEFF Research Database (Denmark)

Sethi, AA; Tybjærg-Hansen, A; Jensen, Gorm Boje

2005-01-01

Since beta2-adrenergic receptors are important regulators of blood pressure, genetic variation in this receptor could explain risk of elevated blood pressure in selected individuals. We tested the hypothesis that Gly16Arg, Gln27Glu, and Thr164Ile in the beta2-adrenergic receptor gene associated w...

Association of TGF-β1 and XPD polymorphisms with severe acute radiation-induced esophageal toxicity in locally advanced lung cancer patients treated with radiotherapy

International Nuclear Information System (INIS)

Zhang Li; Yang Ming; Bi Nan; Ji Wei; Wu Chen; Tan Wen; Zhao Lujun; Yu Dianke; Lin Dongxin; Wang Luhua

2010-01-01

Purpose: Radiation-induced esophageal toxicity (RIET) is a dose-limiting toxicity in lung cancer patients receiving radiotherapy. Accumulating evidence indicates that DNA repair and the cytokine pathways play essential roles in radiation-induced diseases. Genetic polymorphisms of genes in these pathways may affect gene function and/or gene expression and lead to different treatment-related esophageal toxicity. Materials and methods: This study investigated the association of 21 polymorphisms in 14 genes, with the occurrence of ≥grade 2 acute RIET. Genotypes were analyzed among 213 stage III lung cancer patients receiving radiotherapy. Results: We used Cox proportional hazard model to examine the effects of genotypes on ≥grade 2 acute RIET risk and Kaplan-Meier estimator to compare effects of different genotypes on such risk. Multivariate analysis showed that CT or TT genotype of TGF-β1-509C/T polymorphism was associated with a significantly higher RIET risk (adjusted hazard ratio [HR] = 2.47; 95% confidence interval (CI) = 1.17-5.24; P = 0.018, or HR = 3.86; 95% CI = 1.50-9.92; P = 0.005), respectively, compared with the CC genotype. Moreover, Lys/Gln+Gln/Gln genotypes of XPD Lys751Gln polymorphism were also associated with a significantly decreased RIET risk (adjusted HR = 0.55; 95% CI = 0.32-0.96; P = 0.030). Conclusions: This report, for the first time, examined the influence of inherited variation in the DNA repair and the cytokine pathways on RIET.

The Androgen Receptor Gene Mutations Database.

Science.gov (United States)

Gottlieb, B; Lehvaslaiho, H; Beitel, L K; Lumbroso, R; Pinsky, L; Trifiro, M

1998-01-01

The current version of the androgen receptor (AR) gene mutations database is described. The total number of reported mutations has risen from 272 to 309 in the past year. We have expanded the database: (i) by giving each entry an accession number; (ii) by adding information on the length of polymorphic polyglutamine (polyGln) and polyglycine (polyGly) tracts in exon 1; (iii) by adding information on large gene deletions; (iv) by providing a direct link with a completely searchable database (courtesy EMBL-European Bioinformatics Institute). The addition of the exon 1 polymorphisms is discussed in light of their possible relevance as markers for predisposition to prostate or breast cancer. The database is also available on the internet (http://www.mcgill. ca/androgendb/ ), from EMBL-European Bioinformatics Institute (ftp. ebi.ac.uk/pub/databases/androgen ), or as a Macintosh FilemakerPro or Word file (MC33@musica.mcgill.ca).

Mitochondrial DNA Variants Mediate Energy Production and Expression Levels for CFH, C3 and EFEMP1 Genes: Implications for Age-Related Macular Degeneration

Science.gov (United States)

Kenney, M. Cristina; Chwa, Marilyn; Atilano, Shari R.; Pavlis, Janelle M.; Falatoonzadeh, Payam; Ramirez, Claudio; Malik, Deepika; Hsu, Tiffany; Woo, Grace; Soe, Kyaw; Nesburn, Anthony B.; Boyer, David S.; Kuppermann, Baruch D.; Jazwinski, S. Michal; Miceli, Michael V.; Wallace, Douglas C.; Udar, Nitin

2013-01-01

Background Mitochondrial dysfunction is associated with the development and progression of age-related macular degeneration (AMD). Recent studies using populations from the United States and Australia have demonstrated that AMD is associated with mitochondrial (mt) DNA haplogroups (as defined by combinations of mtDNA polymorphisms) that represent Northern European Caucasians. The aim of this study was to use the cytoplasmic hybrid (cybrid) model to investigate the molecular and biological functional consequences that occur when comparing the mtDNA H haplogroup (protective for AMD) versus J haplogroup (high risk for AMD). Methodology/Principal Findings Cybrids were created by introducing mitochondria from individuals with either H or J haplogroups into a human retinal epithelial cell line (ARPE-19) that was devoid of mitochondrial DNA (Rho0). In cybrid lines, all of the cells carry the same nuclear genes but vary in mtDNA content. The J cybrids had significantly lower levels of ATP and reactive oxygen/nitrogen species production, but increased lactate levels and rates of growth. Q-PCR analyses showed J cybrids had decreased expressions for CFH, C3, and EFEMP1 genes, high risk genes for AMD, and higher expression for MYO7A, a gene associated with retinal degeneration in Usher type IB syndrome. The H and J cybrids also have comparatively altered expression of nuclear genes involved in pathways for cell signaling, inflammation, and metabolism. Conclusion/Significance Our findings demonstrate that mtDNA haplogroup variants mediate not only energy production and cell growth, but also cell signaling for major molecular pathways. These data support the hypothesis that mtDNA variants play important roles in numerous cellular functions and disease processes, including AMD. PMID:23365660

Mitochondrial DNA variants mediate energy production and expression levels for CFH, C3 and EFEMP1 genes: implications for age-related macular degeneration.

Directory of Open Access Journals (Sweden)

M Cristina Kenney

Full Text Available Mitochondrial dysfunction is associated with the development and progression of age-related macular degeneration (AMD. Recent studies using populations from the United States and Australia have demonstrated that AMD is associated with mitochondrial (mt DNA haplogroups (as defined by combinations of mtDNA polymorphisms that represent Northern European Caucasians. The aim of this study was to use the cytoplasmic hybrid (cybrid model to investigate the molecular and biological functional consequences that occur when comparing the mtDNA H haplogroup (protective for AMD versus J haplogroup (high risk for AMD.Cybrids were created by introducing mitochondria from individuals with either H or J haplogroups into a human retinal epithelial cell line (ARPE-19 that was devoid of mitochondrial DNA (Rho0. In cybrid lines, all of the cells carry the same nuclear genes but vary in mtDNA content. The J cybrids had significantly lower levels of ATP and reactive oxygen/nitrogen species production, but increased lactate levels and rates of growth. Q-PCR analyses showed J cybrids had decreased expressions for CFH, C3, and EFEMP1 genes, high risk genes for AMD, and higher expression for MYO7A, a gene associated with retinal degeneration in Usher type IB syndrome. The H and J cybrids also have comparatively altered expression of nuclear genes involved in pathways for cell signaling, inflammation, and metabolism.Our findings demonstrate that mtDNA haplogroup variants mediate not only energy production and cell growth, but also cell signaling for major molecular pathways. These data support the hypothesis that mtDNA variants play important roles in numerous cellular functions and disease processes, including AMD.

Genome Wide Association Analysis Reveals New Production Trait Genes in a Male Duroc Population.

Directory of Open Access Journals (Sweden)

Kejun Wang

Full Text Available In this study, 796 male Duroc pigs were used to identify genomic regions controlling growth traits. Three production traits were studied: food conversion ratio, days to 100 KG, and average daily gain, using a panel of 39,436 single nucleotide polymorphisms. In total, we detected 11 genome-wide and 162 chromosome-wide single nucleotide polymorphism trait associations. The Gene ontology analysis identified 14 candidate genes close to significant single nucleotide polymorphisms, with growth-related functions: six for days to 100 KG (WT1, FBXO3, DOCK7, PPP3CA, AGPAT9, and NKX6-1, seven for food conversion ratio (MAP2, TBX15, IVL, ARL15, CPS1, VWC2L, and VAV3, and one for average daily gain (COL27A1. Gene ontology analysis indicated that most of the candidate genes are involved in muscle, fat, bone or nervous system development, nutrient absorption, and metabolism, which are all either directly or indirectly related to growth traits in pigs. Additionally, we found four haplotype blocks composed of suggestive single nucleotide polymorphisms located in the growth trait-related quantitative trait loci and further narrowed down the ranges, the largest of which decreased by ~60 Mb. Hence, our results could be used to improve pig production traits by increasing the frequency of favorable alleles via artificial selection.

Genetic Resources for Advanced Biofuel Production Described with the Gene Ontology

Directory of Open Access Journals (Sweden)

Trudy eTorto-Alalibo

2014-10-01

Full Text Available Dramatic increases in research in the area of microbial biofuel production coupled with high-throughput data generation on bioenergy-related microbes has led to a deluge of information in the scientific literature and in databases. Consolidating this information and making it easily accessible requires a unified vocabulary. The Gene Ontology (GO fulfills that requirement, as it is a well-developed structured vocabulary that describes the activities and locations of gene products in a consistent manner across all kingdoms of life. The Microbial Energy Gene Ontology (MENGO: http://www.mengo.biochem.vt.edu project is extending the GO to include new terms to describe microbial processes of interest to bioenergy production. Our effort has added over 600 bioenergy related terms to the Gene Ontology. These terms will aid in the comprehensive annotation of gene products from diverse energy-related microbial genomes. An area of microbial energy research that has received a lot of attention is microbial production of advanced biofuels. These include alcohols such as butanol, isopropanol, isobutanol, and fuels derived from fatty acids, isoprenoids, and polyhydroxyalkanoates. These fuels are superior to first generation biofuels (ethanol and biodiesel esterified from vegetable oil or animal fat, can be generated from non-food feedstock sources, can be used as supplements or substitutes for gasoline, diesel and jet fuels, and can be stored and distributed using existing infrastructure. Here we review the roles of genes associated with synthesis of advanced biofuels, and at the same time introduce the use of the GO to describe the functions of these genes in a standardized way.

Next-generation sequencing identifies transportin 3 as the causative gene for LGMD1F.

Directory of Open Access Journals (Sweden)

Annalaura Torella

Full Text Available Limb-girdle muscular dystrophies (LGMD are genetically and clinically heterogeneous conditions. We investigated a large family with autosomal dominant transmission pattern, previously classified as LGMD1F and mapped to chromosome 7q32. Affected members are characterized by muscle weakness affecting earlier the pelvic girdle and the ileopsoas muscles. We sequenced the whole exome of four family members and identified a shared heterozygous frame-shift variant in the Transportin 3 (TNPO3 gene, encoding a member of the importin-β super-family. The TNPO3 gene is mapped within the LGMD1F critical interval and its 923-amino acid human gene product is also expressed in skeletal muscle. In addition, we identified an isolated case of LGMD with a new missense mutation in the same gene. We localized the mutant TNPO3 around the nucleus, but not inside. The involvement of gene related to the nuclear transport suggests a novel disease mechanism leading to muscular dystrophy.

Enhanced citrate production through gene insertion in Aspergillus niger

DEFF Research Database (Denmark)

Jongh, Wian de; Nielsen, Jens

2007-01-01

The effect of inserting genes involved in the reductive branch of the tricarboxylic acid (TCA) cycle on citrate production by Aspergillus niger was evaluated. Several different genes were inserted individually and in combination, i.e. malate dehydrogenase (mdh2) from Saccharomyces cerevisiae, two...

Epstein-Barr virus immediate-early gene product trans-activates gene expression from the human immunodeficiency virus long terminal repeat

International Nuclear Information System (INIS)

Kenney, S.; Kamine, J.; Markovitz, D.; Fenrick, R.; Pagano, J.

1988-01-01

Acquired immunodeficiency syndrome patients are frequently coinfected with Epstein-Barr virus (EBV). In this report, the authors demonstrate that an EBV immediate-early gene product, BamHI MLF1, stimulates expression of the bacterial chloramphenicol acetyltransferase (CAT) gene linked to the human immunodeficiency virus (HIV) promoter. The HIV promoter sequences necessary for trans-activation by EBV do not include the tat-responsive sequences. In addition, in contrast to the other herpesvirus trans-activators previously studied, the EBV BamHI MLF1 gene product appears to function in part by a posttranscriptional mechanism, since it increases pHIV-CAT protein activity more than it increases HIV-CAT mRNA. This ability of an EBV gene product to activate HIV gene expression may have biologic consequences in persons coinfected with both viruses

Nano-nutrition of chicken embryos

DEFF Research Database (Denmark)

Grodzik, Marta; Sawosz, Filip; Sawosz, Ewa

2013-01-01

factors of chicken embryo pectoral muscles. ND, Gln, and Gln/ND solutions (50 mg/L) were injected into fertilized broiler chicken eggs at the beginning of embryogenesis. Muscle tissue was dissected at day 20 of incubation and analysed for gene expression of FGF2, VEGF-A, and MyoD1. ND and especially Gln...

Glycogen Synthase Kinase-3 regulates IGFBP-1 gene transcription through the Thymine-rich Insulin Response Element

Directory of Open Access Journals (Sweden)

Marquez Rodolfo

2004-09-01

Full Text Available Abstract Background Hepatic expression of several gene products involved in glucose metabolism, including phosphoenolpyruvate carboxykinase (PEPCK, glucose-6-phosphatase (G6Pase and insulin-like growth factor binding protein-1 (IGFBP-1, is rapidly and completely inhibited by insulin. This inhibition is mediated through the regulation of a DNA element present in each of these gene promoters, that we call the Thymine-rich Insulin Response Element (TIRE. The insulin signalling pathway that results in the inhibition of these gene promoters requires the activation of phosphatidylinositol 3-kinase (PI 3-kinase. However, the molecules that connect PI 3-kinase to these gene promoters are not yet fully defined. Glycogen Synthase Kinase 3 (GSK-3 is inhibited following activation of PI 3-kinase. We have shown previously that inhibitors of GSK-3 reduce the activity of two TIRE-containing gene promoters (PEPCK and G6Pase, whose products are required for gluconeogenesis. Results In this report we demonstrate that in H4IIE-C3 cells, four distinct classes of GSK-3 inhibitor mimic the effect of insulin on a third TIRE-containing gene, IGFBP-1. We identify the TIRE as the minimum requirement for inhibition by these agents, and demonstrate that the target of GSK-3 is unlikely to be the postulated TIRE-binding protein FOXO-1. Importantly, overexpression of GSK-3 in cells reduces the insulin regulation of TIRE activity as well as endogenous IGFBP-1 expression. Conclusions These results implicate GSK-3 as an intermediate in the pathway from the insulin receptor to the TIRE. Indeed, this is the first demonstration of an absolute requirement for GSK-3 inhibition in insulin regulation of gene transcription. These data support the potential use of GSK-3 inhibitors in the treatment of insulin resistant states such as Type 2 diabetes mellitus, but suggest that it will be important to identify all TIRE-containing genes to assess potential side effects of these agents.

Variations in CCL3L gene cluster sequence and non-specific gene copy numbers

Directory of Open Access Journals (Sweden)

Edberg Jeffrey C

2010-03-01

Full Text Available Abstract Background Copy number variations (CNVs of the gene CC chemokine ligand 3-like1 (CCL3L1 have been implicated in HIV-1 susceptibility, but the association has been inconsistent. CCL3L1 shares homology with a cluster of genes localized to chromosome 17q12, namely CCL3, CCL3L2, and, CCL3L3. These genes are involved in host defense and inflammatory processes. Several CNV assays have been developed for the CCL3L1 gene. Findings Through pairwise and multiple alignments of these genes, we have shown that the homology between these genes ranges from 50% to 99% in complete gene sequences and from 70-100% in the exonic regions, with CCL3L1 and CCL3L3 being identical. By use of MEGA 4 and BioEdit, we aligned sense primers, anti-sense primers, and probes used in several previously described assays against pre-multiple alignments of all four chemokine genes. Each set of probes and primers aligned and matched with overlapping sequences in at least two of the four genes, indicating that previously utilized RT-PCR based CNV assays are not specific for only CCL3L1. The four available assays measured median copies of 2 and 3-4 in European and African American, respectively. The concordance between the assays ranged from 0.44-0.83 suggesting individual discordant calls and inconsistencies with the assays from the expected gene coverage from the known sequence. Conclusions This indicates that some of the inconsistencies in the association studies could be due to assays that provide heterogenous results. Sequence information to determine CNV of the three genes separately would allow to test whether their association with the pathogenesis of a human disease or phenotype is affected by an individual gene or by a combination of these genes.

Weaver gene 3'UTR novel mutations: Associations with production ...

African Journals Online (AJOL)

use

2011-11-21

Nov 21, 2011 ... College of Animal Science and Technology, Northwest A&F University, Shaanxi ... To further understand the effects of weaver gene variant on fat, protein, .... one kid, but does that lambed twins were also part of the dataset.

The immunomodulatory gene products of myxoma virus

Indian Academy of Sciences (India)

Unknown

273. Keywords. Gene products; myxoma virus; Oryctolagus cuniculus; poxvirus; skin lesions ...... these data is that these viral proteins do not promote class .... Cudmore S, Reckmann I and Way M 1997 Viral manipulations of the actin ...

How Posttranslational Modification of Nitrogenase Is Circumvented in Rhodopseudomonas palustris Strains That Produce Hydrogen Gas Constitutively

Science.gov (United States)

Heiniger, Erin K.; Oda, Yasuhiro; Samanta, Sudip K.

2012-01-01

Nitrogenase catalyzes the conversion of dinitrogen gas (N2) and protons to ammonia and hydrogen gas (H2). This is a catalytically difficult reaction that requires large amounts of ATP and reducing power. Thus, nitrogenase is not normally expressed or active in bacteria grown with a readily utilized nitrogen source like ammonium. nifA* mutants of the purple nonsulfur phototrophic bacterium Rhodopseudomonas palustris have been described that express nitrogenase genes constitutively and produce H2 when grown with ammonium as a nitrogen source. This raised the regulatory paradox of why these mutants are apparently resistant to a known posttranslational modification system that should switch off the activity of nitrogenase. Microarray, mutation analysis, and gene expression studies showed that posttranslational regulation of nitrogenase activity in R. palustris depends on two proteins: DraT2, an ADP-ribosyltransferase, and GlnK2, an NtrC-regulated PII protein. GlnK2 was not well expressed in ammonium-grown NifA* cells and thus not available to activate the DraT2 nitrogenase modification enzyme. In addition, the NifA* strain had elevated nitrogenase activity due to overexpression of the nif genes, and this increased amount of expression overwhelmed a basal level of activity of DraT2 in ammonium-grown cells. Thus, insufficient levels of both GlnK2 and DraT2 allow H2 production by an nifA* mutant grown with ammonium. Inactivation of the nitrogenase posttranslational modification system by mutation of draT2 resulted in increased H2 production by ammonium-grown NifA* cells. PMID:22179236

Vasopressin Gene-Related Products in the Management of Breast Cancer

National Research Council Canada - National Science Library

North, William

1998-01-01

.... The VP gene is expressed by seemingly all breast cancers and by all DCIS, and this information coupled with an absence of VP gene-related products from fibrocystic disease potentially provides us...

[Mutation analysis of FGFR3 gene in a family featuring hereditary dwarfism].

Science.gov (United States)

Zhang, Qiong; Jiang, Hai-ou; Quan, Qing-li; Li, Jun; He, Ting; Huang, Xue-shuang

2011-12-01

To investigate the clinical symptoms and potential mutation in FGFR3 gene for a family featuring hereditary dwarfism in order to attain diagnosis and provide prenatal diagnosis. Five patients and two unaffected relatives from the family, in addition with 100 healthy controls, were recruited. Genome DNA was extracted. Exons 10 and 13 of the FGFR3 gene were amplified using polymerase chain reaction (PCR). PCR products were sequenced in both directions. All patients had similar features including short stature, short limbs, lumbar hyperlordosis but normal craniofacial features. A heterozygous mutation G1620T (N540K) was identified in the cDNA from all patients but not in the unaffected relatives and 100 control subjects. A heterozygous G380R mutation was excluded. The hereditary dwarfism featured by this family has been caused by hypochondroplasia (HCH) due to a N540K mutation in the FGFR3 gene.

Associations between variants of the HAL gene and milk production traits in Chinese Holstein cows.

Science.gov (United States)

Wang, Haifei; Jiang, Li; Wang, Wenwen; Zhang, Shengli; Yin, Zongjun; Zhang, Qin; Liu, Jian-Feng

2014-11-25

The histidine ammonia-lyse gene (HAL) encodes the histidine ammonia-lyase, which catalyzes the first reaction of histidine catabolism. In our previous genome-wide association study in Chinese Holstein cows to identify genetic variants affecting milk production traits, a SNP (rs41647754) located 357 bp upstream of HAL, was found to be significantly associated with milk yield and milk protein yield. In addition, the HAL gene resides within the reported QTLs for milk production traits. The aims of this study were to identify genetic variants in HAL and to test the association between these variants and milk production traits. Fifteen SNPs were identified within the regions under study of the HAL gene, including three coding mutations, seven intronic mutations, one promoter region mutation, and four 3'UTR mutations. Nine of these identified SNPs were chosen for subsequent genotyping and association analyses. Our results showed that five SNP markers (ss974768522, ss974768525, ss974768531, ss974768533 and ss974768534) were significantly associated with one or more milk production traits. Haplotype analysis showed that two haplotype blocks were significantly associated with milk yield and milk protein yield, providing additional support for the association between HAL variants and milk production traits in dairy cows (P HAL gene and milk production traits in Chinese Holstein cows, indicating the potential role of HAL variants in these traits. These identified SNPs may serve as genetic markers used in genomic selection schemes to accelerate the genetic gains of milk production traits in dairy cattle.

NDP gene mutations in 14 French families with Norrie disease.

Science.gov (United States)

Royer, Ghislaine; Hanein, Sylvain; Raclin, Valérie; Gigarel, Nadine; Rozet, Jean-Michel; Munnich, Arnold; Steffann, Julie; Dufier, Jean-Louis; Kaplan, Josseline; Bonnefont, Jean-Paul

2003-12-01

Norrie disease is a rare X-inked recessive condition characterized by congenital blindness and occasionally deafness and mental retardation in males. This disease has been ascribed to mutations in the NDP gene on chromosome Xp11.1. Previous investigations of the NDP gene have identified largely sixty disease-causing sequence variants. Here, we report on ten different NDP gene allelic variants in fourteen of a series of 21 families fulfilling inclusion criteria. Two alterations were intragenic deletions and eight were nucleotide substitutions or splicing variants, six of them being hitherto unreported, namely c.112C>T (p.Arg38Cys), c.129C>G (p.His43Gln), c.133G>A (p.Val45Met), c.268C>T (p.Arg90Cys), c.382T>C (p.Cys128Arg), c.23479-1G>C (unknown). No NDP gene sequence variant was found in seven of the 21 families. This observation raises the issue of misdiagnosis, phenocopies, or existence of other X-linked or autosomal genes, the mutations of which would mimic the Norrie disease phenotype. Copyright 2003 Wiley-Liss, Inc.

Structural organization of the transfer RNA operon I of Vibrio cholerae

Indian Academy of Sciences (India)

Nine major transfer RNA (tRNA) gene clusters were analysed in various Vibrio cholerae strains. Of these, only the tRNA operon I was found to differ significantly in V. cholerae classical (sixth pandemic) and El Tor (seventh pandemic) strains. Amongst the sixteen tRNA genes contained in this operon, genes for tRNA Gln3 ...

Glutamic acid promotes monacolin K production and monacolin K biosynthetic gene cluster expression in Monascus.

Science.gov (United States)

Zhang, Chan; Liang, Jian; Yang, Le; Chai, Shiyuan; Zhang, Chenxi; Sun, Baoguo; Wang, Chengtao

2017-12-01

This study investigated the effects of glutamic acid on production of monacolin K and expression of the monacolin K biosynthetic gene cluster. When Monascus M1 was grown in glutamic medium instead of in the original medium, monacolin K production increased from 48.4 to 215.4 mg l -1 , monacolin K production increased by 3.5 times. Glutamic acid enhanced monacolin K production by upregulating the expression of mokB-mokI; on day 8, the expression level of mokA tended to decrease by Reverse Transcription-polymerase Chain Reaction. Our findings demonstrated that mokA was not a key gene responsible for the quantity of monacolin K production in the presence of glutamic acid. Observation of Monascus mycelium morphology using Scanning Electron Microscope showed glutamic acid significantly increased the content of Monascus mycelium, altered the permeability of Monascus mycelium, enhanced secretion of monacolin K from the cell, and reduced the monacolin K content in Monascus mycelium, thereby enhancing monacolin K production.

XPC gene mutations in families with xeroderma pigmentosum from Pakistan; prevalent founder effect.

Science.gov (United States)

Ijaz, Ambreen; Basit, Sulman; Gul, Ajab; Batool, Lilas; Hussain, Abrar; Afzal, Sibtain; Ramzan, Khushnooda; Ahmad, Jamil; Wali, Abdul

2018-03-23

Xeroderma pigmentosum (XP) is a rare autosomal recessive skin disorder characterized by hyperpigmentation, premature skin aging, ocular and cutaneous photosensitivity, and increased risk of skin carcinoma. We investigated seven consanguineous XP families with nine patients from Pakistan. All the Patients exhibited typical clinical symptoms of XP since first year of life. Whole genome SNP genotyping identified a 14 Mb autozygous region segregating with the disease phenotype on chromosome 3p25.1. DNA sequencing of XPC gene revealed a founder homozygous splice site mutation (c.2251-1G>C) in patients from six families (A-F) and a homozygous nonsense mutation (c.1399C>T; p.Gln467*) in patients of family G. This is the first report of XPC mutations, underlying XP phenotype, in Pakistani population. © 2018 Japanese Teratology Society.

Form gene clustering method about pan-ethnic-group products based on emotional semantic

Science.gov (United States)

Chen, Dengkai; Ding, Jingjing; Gao, Minzhuo; Ma, Danping; Liu, Donghui

2016-09-01

The use of pan-ethnic-group products form knowledge primarily depends on a designer's subjective experience without user participation. The majority of studies primarily focus on the detection of the perceptual demands of consumers from the target product category. A pan-ethnic-group products form gene clustering method based on emotional semantic is constructed. Consumers' perceptual images of the pan-ethnic-group products are obtained by means of product form gene extraction and coding and computer aided product form clustering technology. A case of form gene clustering about the typical pan-ethnic-group products is investigated which indicates that the method is feasible. This paper opens up a new direction for the future development of product form design which improves the agility of product design process in the era of Industry 4.0.

Clinical features and prognostic implications of familial hypertrophic cardiomyopathy related to the cardiac myosin-binding protein C gene.

Science.gov (United States)

Charron, P; Dubourg, O; Desnos, M; Bennaceur, M; Carrier, L; Camproux, A C; Isnard, R; Hagege, A; Langlard, J M; Bonne, G; Richard, P; Hainque, B; Bouhour, J B; Schwartz, K; Komajda, M

1998-06-09

Little information is available on phenotype-genotype correlations in familial hypertrophic cardiomyopathy that are related to the cardiac myosin binding protein C (MYBPC3) gene. The aim of this study was to perform this type of analysis. We studied 76 genetically affected subjects from nine families with seven recently identified mutations (SASint20, SDSint7, SDSint23, branch point int23, Glu542Gln, a deletion in exon 25, and a duplication/deletion in exon 33) in the MYBPC3 gene. Detailed clinical, ECG, and echocardiographic parameters were analyzed. An intergene analysis was performed by comparing the MYBPC3 group to seven mutations in the beta-myosin heavy-chain gene (beta-MHC) group (n=52). There was no significant phenotypic difference among the different mutations in the MYBPC3 gene. However, in the MYBPC3 group compared with the beta-MHC group, (1) prognosis was significantly better (P<0.0001), and no deaths occurred before the age of 40 years; (2) the age at onset of symptoms was delayed (41+/-19 versus 35+/-17 years, P<0.002); and (3) before 30 years of age, the phenotype was particularly mild because penetrance was low (41% versus 62%), maximal wall thicknesses lower (12+/-4 versus 16+/-7 mm, P<0.03), and abnormal T waves less frequent (9% versus 45%, P<0.02). These results are consistent with specific clinical features related to the MYBPC3 gene: onset of the disease appears delayed and the prognosis is better than that associated with the beta-MHC gene. These findings could be particularly important for the purpose of clinical management and genetic counseling in familial hypertrophic cardiomyopathy.

Increased isobutanol production in Saccharomyces cerevisiae by overexpression of genes in valine metabolism

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Karhumaa Kaisa

2011-07-01

Full Text Available Abstract Background Isobutanol can be a better biofuel than ethanol due to its higher energy density and lower hygroscopicity. Furthermore, the branched-chain structure of isobutanol gives a higher octane number than the isomeric n-butanol. Saccharomyces cerevisiae was chosen as the production host because of its relative tolerance to alcohols, robustness in industrial fermentations, and the possibility for future combination of isobutanol production with fermentation of lignocellulosic materials. Results The yield of isobutanol was improved from 0.16 to 0.97 mg per g glucose by simultaneous overexpression of biosynthetic genes ILV2, ILV3, and ILV5 in valine metabolism in anaerobic fermentation of glucose in mineral medium in S. cerevisiae. Isobutanol yield was further improved by twofold by the additional overexpression of BAT2, encoding the cytoplasmic branched-chain amino-acid aminotransferase. Overexpression of ILV6, encoding the regulatory subunit of Ilv2, in the ILV2 ILV3 ILV5 overexpression strain decreased isobutanol production yield by threefold. In aerobic cultivations in shake flasks in mineral medium, the isobutanol yield of the ILV2 ILV3 ILV5 overexpression strain and the reference strain were 3.86 and 0.28 mg per g glucose, respectively. They increased to 4.12 and 2.4 mg per g glucose in yeast extract/peptone/dextrose (YPD complex medium under aerobic conditions, respectively. Conclusions Overexpression of genes ILV2, ILV3, ILV5, and BAT2 in valine metabolism led to an increase in isobutanol production in S. cerevisiae. Additional overexpression of ILV6 in the ILV2 ILV3 ILV5 overexpression strain had a negative effect, presumably by increasing the sensitivity of Ilv2 to valine inhibition, thus weakening the positive impact of overexpression of ILV2, ILV3, and ILV5 on isobutanol production. Aerobic cultivations of the ILV2 ILV3 ILV5 overexpression strain and the reference strain showed that supplying amino acids in cultivation media

Interleukin 1-beta upregulates brain-derived neurotrophic factor, neurotrophin 3 and neuropilin 2 gene expression and NGF production in annulus cells.

Science.gov (United States)

Gruber, H E; Hoelscher, G L; Bethea, S; Hanley, E N

2012-11-01

The relationship between disc cells, nerves and pain production in the intervertebral disc is poorly understood. Neurotrophins, signaling molecules involved in the survival, differentiation and migration of neurons, and neurite outgrowth, are expressed in non-neuronal tissues including the disc. We hypothesized that three-dimensional exposure of human disc cells to the proinflammatory cytokine IL-1ß in vitro would elevate neurotrophin gene expression levels and production of nerve growth factor (NGF). Cells isolated from Thompson grade III and IV discs were cultured for 14 days under control conditions or with addition of 10(2) pM IL-1ß; mRNA was isolated and conditioned media assayed for NGF content. IL-1ß exposure in three-dimensional culture significantly increased expression of neurotrophin 3, brain-derived neurotrophic factor, and neuropilin 2 compared to controls. IL-1ß-exposed cells showed significantly increased NGF production compared to controls. Findings support our hypothesis, expand previous data concerning expression of neurotrophins, and provide the first documented expression of neurotrophin 3 and neuropilin 2. Our results have direct translational relevance, because they address the primary clinical issue of low back pain and open the possibility of novel analgesic therapies using specific small-molecular antagonists to neurotrophins.

rpoB gene mutations among Mycobacterium tuberculosis isolates from extrapulmonary sites.

Science.gov (United States)

Khosravi, Azar Dokht; Meghdadi, Hossein; Ghadiri, Ata A; Alami, Ameneh; Sina, Amir Hossein; Mirsaeidi, Mehdi

2018-03-01

The aim of this study was to analyze mutations occurring in the rpoB gene of Mycobacterium tuberculosis (MTB) isolates from clinical samples of extrapulmonary tuberculosis (EPTB). Seventy formalin-fixed, paraffin-embedded samples and fresh tissue samples from confirmed EPTB cases were analyzed. Nested PCR based on the rpoB gene was performed on the extracted DNAs, combined with cloning and subsequent sequencing. Sixty-seven (95.7%) samples were positive for nester PCR. Sequence analysis of the 81 bp region of the rpoB gene demonstrated mutations in 41 (61.2%) of 67 sequenced samples. Several point mutations including deletion mutations at codons 510, 512, 513 and 515, with 45% and 51% of the mutations in codons 512 and 513 respectively were seen, along with 26% replacement mutations at codons 509, 513, 514, 518, 520, 524 and 531. The most common alteration was Gln → His, at codon 513, presented in 30 (75.6%) isolates. This study demonstrated sequence alterations in codon 513 of the 81 bp region of the rpoB gene as the most common mutation occurred in 75.6% of molecularly confirmed rifampin-resistant strains. In addition, simultaneous mutation at codons 512 and 513 was demonstrated in 34.3% of the isolates. © 2018 APMIS. Published by John Wiley & Sons Ltd.

Opposing roles of STAT4 and Dnmt3a in Th1 gene regulation

Science.gov (United States)

Pham, Duy; Yu, Qing; Walline, Crystal C.; Muthukrishnan, Rajarajeswari; Blum, Janice S.; Kaplan, Mark H.

2013-01-01

The Signal Transducer and Activator of Transcription factor STAT4 is a critical regulator of Th1 differentiation and inflammatory disease. Yet, how STAT4 regulates gene expression is still unclear. In this report, we define a STAT4-dependent sequence of events including H3K4 methylation, Jmjd3 association with STAT4 target loci, and a Jmjd3-dependent decrease in H3K27 trimethylation and DNA methyltransferase (Dnmt) 3a association with STAT4 target loci. Dnmt3a has an obligate role in repressing Th1 gene expression, and in Th1 cultures deficient in both STAT4 and Dnmt3a, there is recovery in the expression of a subset of Th1 genes that is sufficient to increase IFNγ production. Moreover, although STAT4-deficient mice are protected from the development of EAE, mice deficient in STAT4 and conditionally-deficient in Dnmt3a in T cells develop paralysis. Th1 genes that are de-repressed in the absence of Dnmt3a have greater induction following the ectopic expression of the Th1-associated transcription factors T-bet and Hlx1. Together, these data demonstrate that STAT4 and Dnmt3a play opposing roles in regulating Th1 gene expression, and that one mechanism for STAT4-dependent gene programming is in establishing a de-repressed genetic state susceptible to transactivation by additional fate-determining transcription factors. PMID:23772023

A process optimization for bio-catalytic production of substituted catechols (3-nitrocatechol and 3-methylcatechol

Directory of Open Access Journals (Sweden)

Tiwary Bhupendra N

2010-06-01

Full Text Available Abstract Background Substituted catechols are important precursors for large-scale synthesis of pharmaceuticals and other industrial products. Most of the reported chemical synthesis methods are expensive and insufficient at industrial level. However, biological processes for production of substituted catechols could be highly selective and suitable for industrial purposes. Results We have optimized a process for bio-catalytic production of 3-substituted catechols viz. 3-nitrocatechol (3-NC and 3-methylcatechol (3-MC at pilot scale. Amongst the screened strains, two strains viz. Pseudomonas putida strain (F1 and recombinant Escherichia coli expression clone (pDTG602 harboring first two genes of toluene degradation pathway were found to accumulate 3-NC and 3-MC respectively. Various parameters such as amount of nutrients, pH, temperature, substrate concentration, aeration, inoculums size, culture volume, toxicity of substrate and product, down stream extraction, single step and two-step biotransformation were optimized at laboratory scale to obtain high yields of 3-substituted catechols. Subsequently, pilot scale studies were performed in 2.5 liter bioreactor. The rate of product accumulation at pilot scale significantly increased up to ~90-95% with time and high yields of 3-NC (10 mM and 3-MC (12 mM were obtained. Conclusion The biocatalytic production of 3-substituted catechols viz. 3-NC and 3-MC depend on some crucial parameters to obtain maximum yields of the product at pilot scale. The process optimized for production of 3-substituted catechols by using the organisms P. putida (F1 and recombinant E. coli expression clone (pDTG602 may be useful for industrial application.

In silico aided metabolic engineering of Klebsiella oxytoca and fermentation optimization for enhanced 2,3-butanediol production.

Science.gov (United States)

Park, Jong Myoung; Song, Hyohak; Lee, Hee Jong; Seung, Doyoung

2013-09-01

Klebsiella oxytoca naturally produces a large amount of 2,3-butanediol (2,3-BD), a promising bulk chemical with wide industrial applications, along with various byproducts. In this study, the in silico gene knockout simulation of K. oxytoca was carried out for 2,3-BD overproduction by inhibiting the formation of byproducts. The knockouts of ldhA and pflB genes were targeted with the criteria of maximization of 2,3-BD production and minimization of byproducts formation. The constructed K. oxytoca ΔldhA ΔpflB strain showed higher 2,3-BD yields and higher final concentrations than those obtained from the wild-type and ΔldhA strains. However, the simultaneous deletion of both genes caused about a 50 % reduction in 2,3-BD productivity compared with K. oxytoca ΔldhA strain. Based on previous studies and in silico investigation that the agitation speed during 2,3-BD fermentation strongly affected cell growth and 2,3-BD synthesis, the effect of agitation speed on 2,3-BD production was investigated from 150 to 450 rpm in 5-L bioreactors containing 3-L culture media. The highest 2,3-BD productivity (2.7 g/L/h) was obtained at 450 rpm in batch fermentation. Considering the inhibition of acetoin for 2,3-BD production, fed-batch fermentations were performed using K. oxytoca ΔldhA ΔpflB strain to enhance 2,3-BD production. Altering the agitation speed from 450 to 350 rpm at nearly 10 g/L of acetoin during the fed-batch fermentation allowed for the production of 113 g/L 2,3-BD, with a yield of 0.45 g/g, and for the production of 2.1 g/L/h of 2,3-BD.

Association study between XRCC1 gene polymorphisms and sporadic amyotrophic lateral sclerosis.

Science.gov (United States)

Coppedè, Fabio; Migheli, Francesca; Lo Gerfo, Annalisa; Fabbrizi, Maria Rita; Carlesi, Cecilia; Mancuso, Michelangelo; Corti, Stefania; Mezzina, Nicoletta; del Bo, Roberto; Comi, Giacomo P; Siciliano, Gabriele; Migliore, Lucia

2010-01-01

The aim of the present study was to investigate the possible contribution of three common functional polymorphisms in the DNA repair protein X-ray repair cross-complementing group 1 (XRCC1), namely Arg194Trp (rs1799782), Arg280His (rs25489) and Arg399Gln (rs25487), to sporadic amyotrophic lateral sclerosis (SALS). We genotyped 206 Italian SALS patients and 203 matched controls for XRCC1 Arg194Trp, Arg280His and Arg399Gln polymorphisms by means of PCR/RFLP technique, searching for association between any of the studied polymorphisms and disease risk, age and site of onset. We observed a statistically significant difference in XRCC1 Gln399 allele frequencies between SALS cases and controls (0.39/0.28; p=0.001). The present study suggests that the XRCC1 Arg399Gln polymorphism might contribute to SALS risk.

Interaction between lifestyle factors and the XRCC1, XPD, and XRCC3 genetic variations modulates the risk for sporadic colorectal cancer

Directory of Open Access Journals (Sweden)

Procopciuc Lucia Maria

2014-03-01

Full Text Available Introducere: Variațiile genetice, cum ar fi cele care influențează sistemele de reparare a defectelor de replicare a ADN, pot reprezenta factori de susceptibilitate în cancerul colorectal sporadic (CCR ca urmare a interacțiunii cu factori de mediu. Material și metodă: 80 de femei și 70 de bărbați, pacienți diagnosticați cu CCR sporadic în Clinica Chirurgie III Cluj au fost genotipați pentru Arg399Gln-XRCC1, Lys751Gln-XPD și Met241Thr-XRCC3 utilizând metodele PCR-RFLP. Am determinat de asemenea, genotipurile pentru 100 femei și 62 bărbați , care au format grupul de control. Rezultatele au fost analizate din punct de vedere al relației cu factorii de risc de mediu, fumatul și dieta. Rezultate: Bărbații fumători purtători ai variațiilor genetice Arg399Gln, Lys751Gln, Met241Thr au avut un risc semnificativ crescut de 4.09 (95%IC[0.96-19.98],p=0.05, 5.95(95%IC[1.08-43.22],p=0.03 și respectiv 3.73(95%IC[0.86-18.53],p=0.05 de a dezvolta cancer colorectal sporadic. Un risc semnificativ crescut de a dezvolta cancer colorectal sporadic a fost observat în cazul femeilor și bărbaților cu o dietă bogată în carne roșie prăjită purtători ai variațiilor genetice Arg399Gln (OR 2.77 95%IC [1.34-6.82],p=0.015 și OR 8.64 95%IC[2.67-29.14],p<0.001, Lys751Gln (OR 4.12 95%IC[1.37-12.74],p=0.007 și OR 5.06 95%IC[1.4- 19.02],p=0.006, Met241Thr (OR5.92 95%IC[2.21-16.23],p<0.001 și OR 5.64 95%IC[1.52-21.7],p=0.022. Femeile a căror dietă a inclus cantități mari de carne roșie prăjită au avut un risc semnificativ crescut de a dezvolta timpuriu cancer colorectal sporadic dacă au fost purtătoare a variațiilor genetice Arg399Gln-XRCC1 (OR 5.14 95%IC[0.99-28.3],p=0.047, Thr241Met-XRCC3 (OR 6.67 95%IC[1.05-46.67],p=0.025 și Lys751Gln-XPD (OR 4.7 95%IC[0.99-23.32],p=0.034. Concluzii: În cazul populației de origine română, asocierea genotipurilor mutante cu factori de mediu modulează riscul pentru CCR sporadic. La femei

Characterization and cloning of laccase gene from Hericium coralloides NBRC 7716 suitable for production of epitheaflagallin 3-O-gallate.

Science.gov (United States)

Itoh, Nobuya; Takagi, Shinya; Miki, Asami; Kurokawa, Junji

2016-01-01

Epitheaflagallin 3-O-gallate (ETFGg) is a minor polyphenol found in black tea extract, which has good physiological functions. It is synthesized from epigallocatechin gallate (EGCg) with gallic acid via laccase oxidation. Various basidiomycetes and fungi were screened to find a suitable laccase for the production of ETFGg. A basidiomycete, Hericium coralloides NBRC 7716, produced an appropriate extracellular laccase. The purified laccase produced twice the level of ETFGg compared with commercially available laccase from Trametes sp. The enzyme, termed Lcc2, is a monomeric protein with an apparent molecular mass of 67.2 kDa. The N-terminal amino acid sequence of Lcc2 is quite different from laccase isolated from the fruiting bodies of Hericium. Lcc2 showed similar substrate specificity to known laccases and could oxidize various phenolic substrates, including pyrogallol, gallic acid, and 2,6-dimethoxyphenol. The full-length lcc2 gene was obtained by PCR using degenerate primers, which were designed based on the N-terminal amino acid sequence of Lcc2 and conserved copper-binding sites of laccases, and 5'-, and 3'-RACE PCR with mRNA. The Lcc2 gene showed homology with Lentinula edodes laccase (sharing 77% amino acid identity with Lcc6). We successfully produced extracellular Lcc2 using a heterologous expression system with Saccharomyces cerevisiae. Moreover, it was confirmed that the recombinant laccase generates similar levels of ETFGg as the native enzyme. Copyright © 2015 Elsevier Inc. All rights reserved.

76 FR 9028 - Guidance for Industry: Potency Tests for Cellular and Gene Therapy Products; Availability

Science.gov (United States)

2011-02-16

...] Guidance for Industry: Potency Tests for Cellular and Gene Therapy Products; Availability AGENCY: Food and... Therapy Products'' dated January 2011. The guidance document provides manufacturers of cellular and gene... for Industry: Potency Tests for Cellular and Gene Therapy Products'' dated January 2011. The guidance...

Employing a Recombinant Strain of Advenella mimigardefordensis for Biotechnical Production of Homopolythioesters from 3,3′-Dithiodipropionic Acid

Science.gov (United States)

Xia, Yongzhen; Wübbeler, Jan Hendrik; Qi, Qingsheng

2012-01-01

Advenella mimigardefordensis strain DPN7T was genetically modified to produce poly(3-mercaptopropionic acid) (PMP) homopolymer by exploiting the recently unraveled process of 3,3′-dithiodipropionic acid (DTDP) catabolism. Production was achieved by systematically engineering the metabolism of this strain as follows: (i) deletion of its inherent 3MP dioxygenase-encoding gene (mdo), (ii) introduction of the buk-ptb operon (genes encoding the butyrate kinase, Buk, and the phosphotransbutyrylase, Ptb, from Clostridium acetobutylicum), and (iii) overexpression of its own polyhydroxyalkanoate synthase (phaCAm). These measures yielded the potent PMP production strain A. mimigardefordensis strain SHX22. The deletion of mdo was required for adequate synthesis of PMP due to the resulting accumulation of 3MP during utilization of DTDP. Overexpression of the plasmid-borne buk-ptb operon caused a severe growth repression. This effect was overcome by inserting this operon into the genome. Polyhydroxyalkanoate (PHA) synthases from different origins were compared. The native PHA synthase of A. mimigardefordensis (phaCAm) was obviously the best choice to establish homopolythioester production in this strain. In addition, the cultivation conditions, including an appropriate provision of the carbon source, were further optimized to enhance PMP production. The engineered strain accumulated PMP up to approximately 25% (wt/wt) of the cell dry weight when cultivated in mineral salts medium containing glycerol as the carbon source in addition to DTDP as the sulfur-providing precursor. According to our knowledge, this is the first report of PMP homopolymer production by a metabolically engineered bacterium using DTDP, which is nontoxic, as the precursor substrate. PMID:22344658

Regulatory structures for gene therapy medicinal products in the European Union.

Science.gov (United States)

Klug, Bettina; Celis, Patrick; Carr, Melanie; Reinhardt, Jens

2012-01-01

Taking into account the complexity and technical specificity of advanced therapy medicinal products: (gene and cell therapy medicinal products and tissue engineered products), a dedicated European regulatory framework was needed. Regulation (EC) No. 1394/2007, the "ATMP Regulation" provides tailored regulatory principles for the evaluation and authorization of these innovative medicines. The majority of gene or cell therapy product development is carried out by academia, hospitals, and small- and medium-sized enterprises (SMEs). Thus, acknowledging the particular needs of these types of sponsors, the legislation also provides incentives for product development tailored to them. The European Medicines Agency (EMA) and, in particular, its Committee for Advanced Therapies (CAT) provide a variety of opportunities for early interaction with developers of ATMPs to enable them to have early regulatory and scientific input. An important tool to promote innovation and the development of new medicinal products by micro-, small-, and medium-sized enterprises is the EMA's SME initiative launched in December 2005 to offer financial and administrative assistance to smaller companies. The European legislation also foresees the involvement of stakeholders, such as patient organizations, in the development of new medicines. Considering that gene therapy medicinal products are developed in many cases for treatment of rare diseases often of monogenic origin, the involvement of patient organizations, which focus on rare diseases and genetic and congenital disorders, is fruitful. Two such organizations are represented in the CAT. Research networks play another important role in the development of gene therapy medicinal products. The European Commission is funding such networks through the EU Sixth Framework Program. Copyright © 2012 Elsevier Inc. All rights reserved.

Candidate genes for drought tolerance and improved productivity in ...

Indian Academy of Sciences (India)

Madhu

tropics. Improving drought tolerance and productivity is one of the most difficult tasks for cereal breeders. The diffi- culty arises from the diverse strategies adopted by plants themselves to combat drought stress depending on the timing,. Candidate genes for drought tolerance and improved productivity in rice (Oryza sativa L.).

Functional Analysis of Genes Involved in the Biosynthesis of Enterocin NKR-5-3B, a Novel Circular Bacteriocin.

Science.gov (United States)

Perez, Rodney H; Ishibashi, Naoki; Inoue, Tomoko; Himeno, Kohei; Masuda, Yoshimitsu; Sawa, Narukiko; Zendo, Takeshi; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2016-01-15

A putative biosynthetic gene cluster of the enterocin NKR-5-3B (Ent53B), a novel circular bacteriocin, was analyzed by sequencing the flanking regions around enkB, the Ent53B structural gene, using a fosmid library. A region approximately 9 kb in length was obtained, and the enkB1, enkB2, enkB3, and enkB4 genes, encoding putative biosynthetic proteins involved in the production, maturation, and secretion of Ent53B, were identified. We also determined the identity of proteins mediating self-immunity against the effects of Ent53B. Heterologous expression systems in various heterologous hosts, such as Enterococcus faecalis and Lactococcus lactis strains, were successfully established. The production and secretion of the mature Ent53B required the cooperative functions of five genes. Ent53B was produced only by those heterologous hosts that expressed protein products of the enkB, enkB1, enkB2, enkB3, and enkB4 genes. Moreover, self-immunity against the antimicrobial action of Ent53B was conferred by at least two independent mechanisms. Heterologous hosts harboring the intact enkB4 gene and/or a combination of intact enkB1 and enkB3 genes were immune to the inhibitory action of Ent53B. In addition to their potential application as food preservatives, circular bacteriocins are now considered possible alternatives to therapeutic antibiotics due to the exceptional stability conferred by their circular structure. The successful practical application of circular bacteriocins will become possible only if the molecular details of their biosynthesis are fully understood. The results of the present study offer a new perspective on the possible mechanism of circular bacteriocin biosynthesis. In addition, since some enterococcal strains are associated with pathogenicity, virulence, and drug resistance, the establishment of the first multigenus host heterologous production of Ent53B has very high practical significance, as it widens the scope of possible Ent53B applications

Effects of Copper on Hemocyte Apoptosis, ROS Production, and Gene Expression in White Shrimp Litopenaeus vannamei.

Science.gov (United States)

Guo, Hui; Li, Kexu; Wang, Wei; Wang, Chenggui; Shen, Yuchun

2017-10-01

Copper, a common chemical contaminant in aquatic environment, is known to be toxic to aquatic life at high concentrations. In the present study, we evaluated the apoptotic cell ratio and ROS production in hemocytes of the white shrimp Litopenaeus vannamei exposed to 1 or 5 mg L -1 Cu for 0, 3, 6, 12, 24, and 48 h. The expression changes of antioxidant biomarker genes, i.e., copper-zinc superoxide dismutase (Cu-Zn SOD) and catalase (CAT), apoptosis-related genes, i.e., caspase-3 and inhibitor of apoptosis protein (IAP), and a specific biomarker gene of heavy metal pollution, i.e., metallothionein (MT), were also determined in hemocytes. Significant increases in ROS production were observed in both treatment groups at each time points. The apoptotic cell ratios were significantly increased at 6-48 h among shrimp exposed to 1 mg L -1 Cu and at each time points in 5 mg L -1 Cu group. These results indicated that Cu would induce oxidative stress and apoptosis in the hemocyte of L. vannamei. Quantitative real-time PCR analysis revealed that the relative expression levels of Cu-Zn SOD, CAT, caspase-3, IAP, and MT were upregulated in a dose-dependent and time-dependent manner, suggesting the involvement of these genes in stress response against Cu exposure.

Escherichia coli yjjPB genes encode a succinate transporter important for succinate production.

Science.gov (United States)

Fukui, Keita; Nanatani, Kei; Hara, Yoshihiko; Yamakami, Suguru; Yahagi, Daiki; Chinen, Akito; Tokura, Mitsunori; Abe, Keietsu

2017-09-01

Under anaerobic conditions, Escherichia coli produces succinate from glucose via the reductive tricarboxylic acid cycle. To date, however, no genes encoding succinate exporters have been established in E. coli. Therefore, we attempted to identify genes encoding succinate exporters by screening an E. coli MG1655 genome library. We identified the yjjPB genes as candidates encoding a succinate transporter, which enhanced succinate production in Pantoea ananatis under aerobic conditions. A complementation assay conducted in Corynebacterium glutamicum strain AJ110655ΔsucE1 demonstrated that both YjjP and YjjB are required for the restoration of succinate production. Furthermore, deletion of yjjPB decreased succinate production in E. coli by 70% under anaerobic conditions. Taken together, these results suggest that YjjPB constitutes a succinate transporter in E. coli and that the products of both genes are required for succinate export.

Data of evolutionary structure change: 1JMEB-3EKFA [Confc[Archive

Lifescience Database Archive (English)

Full Text Available 1JMEB-3EKFA 1JME 3EKF B A --KEMPQPKTFGELKNLPLLNTDKPVQALMKIADELGEI...e>PHE CA 283 GLN CA 245 3EKF ...A 3EKFA MNKLQRANQFQ HHGGG ...ax>11.26553726196289 1 3EKF... A 3EKFA NQFQE--KVMND

Enhanced pest resistance and increased phenolic production in maize callus transgenically expressing a maize chalcone isomerase -3 like gene

Science.gov (United States)

Significant losses in maize production are due to damage by insects and ear rot fungi. A gene designated as chalcone-isomerase-like, located in a quantitative trait locus for resistance to Fusarium ear rot fungi, was cloned from a Fusarium ear rot resistant inbred and transgenically expressed in mai...

The ipdC, hisC1 and hisC2 genes involved in indole-3-acetic production used as alternative phylogenetic markers in Azospirillum brasilense.

Science.gov (United States)

Jijón-Moreno, Saúl; Marcos-Jiménez, Cynthia; Pedraza, Raúl O; Ramírez-Mata, Alberto; de Salamone, I García; Fernández-Scavino, Ana; Vásquez-Hernández, Claudia A; Soto-Urzúa, Lucia; Baca, Beatriz E

2015-06-01

Plant growth-promoting bacteria of the genus Azospirillum are present in the rhizosphere and as endophytes of many crops. In this research we studied 40 Azospirillum strains isolated from different plants and geographic regions. They were first characterized by 16S rDNA restriction analysis, and their phylogenetic position was established by sequencing the genes 16S rDNA, ipdC, hisC1, and hisC2. The latter three genes are involved in the indole-3-pyruvic acid (IPyA) biosynthesis pathway of indole-3-acetic acid (IAA). Furthermore, the suitability of the 16S-23S rDNA intergenic spacer sequence (IGS) for the differentiation of closely related Azospirillum taxa and development of PCR protocols allows for specific detection of strains. The IGS-RFLP analysis enabled intraspecies differentiation, particularly of Azospirillum brasilense and Azospirillum lipoferum strains. Results demonstrated that the ipdC, hisC1, and hisC2 genes are highly conserved in all the assessed A. brasilense isolates, suggesting that these genes can be used as an alternative phylogenetic marker. In addition, IAA production determined by HPLC ranged from 0.17 to 98.2 μg mg(-1) protein. Southern hybridization with the A. brasilense ipdC gene probe did not show, a hybridization signal with A. lipoferum, Azospirillum amazonense, Azospirillum halopreferans and Azospirillum irakense genomic DNA. This suggests that these species produce IAA by other pathways. Because IAA is mainly synthesized via the IPyA pathway in A. brasilense strains, a species that is used worldwide in agriculture, the identification of ipdC, hisC1, and hisC2 genes by PCR may be suitable for selecting exploitable strains.

The role of base excision repair in the development of primary open angle glaucoma in the Polish population

Energy Technology Data Exchange (ETDEWEB)

Cuchra, Magda; Markiewicz, Lukasz; Mucha, Bartosz [Department of Clinical Chemistry and Biochemistry, Medical University of Lodz (Poland); Pytel, Dariusz [The Abramson Family Cancer Research Institute, Department of Cancer Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104 (United States); Department of Biochemistry and Molecular Biology, Hollings Cancer Center, Medical University of South Carolina, Charleston, SC 29425 (United States); Szymanek, Katarzyna [Department of Ophthalmology, Medical University of Warsaw, SPKSO Hospital, Warsaw (Poland); Szemraj, Janusz [Department of Medical Biochemistry, Medical University of Lodz, Lodz (Poland); Szaflik, Jerzy; Szaflik, Jacek P. [Department of Ophthalmology, Medical University of Warsaw, SPKSO Hospital, Warsaw (Poland); Majsterek, Ireneusz, E-mail: ireneusz.majsterek@umed.lodz.pl [Department of Clinical Chemistry and Biochemistry, Medical University of Lodz (Poland)

2015-08-15

Highlights: • We suggested the association of XRCC1 gene with the increase risk of POAG development. • We indicated the association of clinical factor and XRCC1, MUTYH, ADPRT and APE1 genes with POAG progression. • We postulated the increase level of oxidative DNA damage in group of patients with POAG in relation to healthy controls. • We suggested the slightly decrease ability to repair of oxidative DNA damage. • This is the first data that showed the role of BER mechanism in POAG pathogenesis. - Abstract: Glaucoma is a leading cause of irreversible blindness in developing countries. Previous data have shown that progressive loss of human TM cells may be connected with chronic exposure to oxidative stress. This hypothesis may suggest a role of the base excision repair (BER) pathway of oxidative DNA damage in primary open angle glaucoma (POAG) patients. The aim of our study was to evaluate an association of BER gene polymorphism with a risk of POAG. Moreover, an association of clinical parameters was examined including cup disk ratio (c/d), rim area (RA) and retinal nerve fiber layer (RNFL) with glaucoma progression according to BER gene polymorphisms. Our research included 412 patients with POAG and 454 healthy controls. Gene polymorphisms were analyzed by PCR-RFLP. Heidelberg Retinal Tomography (HRT) clinical parameters were also analyzed. The 399Arg/Gln genotype of the XRCC1 gene (OR 1.38; 95% CI 1.02–1.89 p = 0.03) was associated with an increased risk of POAG occurrence. It was indicated that the 399Gln/Gln XRCC1 genotype might increase the risk of POAG progression according to the c/d ratio (OR 1.67; 95% CI 1.07–2.61 P = 0.02) clinical parameter. Moreover, the association of VF factor with 148Asp/Glu of APE1 genotype distribution and POAG progression (OR 2.25; 95% CI 1.30–3.89) was also found. Additionally, the analysis of the 324Gln/His MUTYH polymorphism gene distribution in the patient group according to RNFL factor showed that it might

Association of XPC Gene Polymorphisms with Colorectal Cancer Risk in a Southern Chinese Population: A Case-Control Study and Meta-Analysis

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Rui-Xi Hua

2016-09-01

Full Text Available Xeroderma pigmentosum group C (XPC is a key component of the nucleotide excision repair (NER pathway. Dysfunctional XPC protein may impair NER-mediated DNA repair capacity and further lead to genomic instability and carcinogenesis. Two common nonsynonymous polymorphisms in the XPC gene, Lys939Gln (rs2228001 A > C and Ala499Val (rs2228000 C > T, have been investigated in various types of cancer. We genotyped these two polymorphisms in 1141 cases with histologically confirmed colorectal cancer (CRC and 1173 healthy controls to explore their causative association with CRC susceptibility. Overall, no association was observed between these two variants and the risk of CRC. Our meta-analysis also confirmed a lack of overall association. Stratified analyses were performed by age, gender, smoking status, pack-year, drinking status, tumor sites, and Duke’s stages. We found that XPC Lys939Gln polymorphism was significantly associated with an increased CRC risk in subjects at 57 years of age or younger (adjusted odds ratio (OR = 1.37, 95% confidence interval (CI = 1.004–1.86, p = 0.047 and non-drinkers (adjusted OR = 1.53, 95% CI = 1.10–2.12, p = 0.011. Our results indicated that XPC Lys939Gln may be a low-penetrance CRC susceptibility polymorphism. Our findings warrant further validation.

Whole-exome sequencing identifies novel candidate predisposition genes for familial polycythemia vera.

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Hirvonen, Elina A M; Pitkänen, Esa; Hemminki, Kari; Aaltonen, Lauri A; Kilpivaara, Outi

2017-04-20

Polycythemia vera (PV), characterized by massive production of erythrocytes, is one of the myeloproliferative neoplasms. Most patients carry a somatic gain-of-function mutation in JAK2, c.1849G > T (p.Val617Phe), leading to constitutive activation of JAK-STAT signaling pathway. Familial clustering is also observed occasionally, but high-penetrance predisposition genes to PV have remained unidentified. We studied the predisposition to PV by exome sequencing (three cases) in a Finnish PV family with four patients. The 12 shared variants (maximum allowed minor allele frequency  G (p.Phe418Leu) in ZXDC, c.1931C > G (p.Pro644Arg) in ATN1, and c.701G > A (p.Arg234Gln) in LRRC3. We also observed a rare, predicted benign germline variant c.2912C > G (p.Ala971Gly) in BCORL1 in all four patients. Somatic mutations in BCORL1 have been reported in myeloid malignancies. We further screened the variants in eight PV patients in six other Finnish families, but no other carriers were found. Exome sequencing provides a powerful tool for the identification of novel variants, and understanding the familial predisposition of diseases. This is the first report on Finnish familial PV cases, and we identified three novel candidate variants that may predispose to the disease.

Glutamine-enriched enteral diet increases renal arginine production

NARCIS (Netherlands)

Houdijk, A. P.; van Leeuwen, P. A.; Teerlink, T.; FLINKERBUSCH, E. L.; Boermeester, M. A.; Sauerwein, H. P.; Wesdorp, R. I.

1994-01-01

Arginine (Arg) is generated in the kidney by the conversion of circulating citrulline. The most important source for circulating citrulline is the metabolism of glutamine (Gln) by the gut. In this study, we investigated the influence of an enteral diet enriched with Gln on renal Arg synthesis in the

Case report of novel CACNA1A gene mutation causing episodic ataxia type 2

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David Alan Isaacs

2017-05-01

Full Text Available Background: Episodic ataxia type 2 (OMIM 108500 is an autosomal dominant channelopathy characterized by paroxysms of ataxia, vertigo, nausea, and other neurologic symptoms. More than 50 mutations of the CACNA1A gene have been discovered in families with episodic ataxia type 2, although 30%–50% of all patients with typical episodic ataxia type 2 phenotype have no detectable mutation of the CACNA1A gene. Case: A 46-year-old Caucasian man, with a long history of bouts of imbalance, vertigo, and nausea, presented to our hospital with 2 weeks of ataxia and headache. Subsequent evaluation revealed a novel mutation in the CACNA1A gene: c.1364 G > A Arg455Gln. Acetazolamide was initiated with symptomatic improvement. Conclusion: This case report expands the list of known CACNA1A mutations associated with episodic ataxia type 2.

Effects of short-term dietary restriction and glutamine supplementation in vitro on the modulation of inflammatory properties.

Science.gov (United States)

C de Oliveira, Dalila; Santos, Ed Wilson; Nogueira-Pedro, Amanda; Xavier, José Guilherme; Borelli, Primavera; Fock, Ricardo Ambrósio

2018-04-01

Dietary restriction (DR) is a nutritional intervention that exerts profound effects on biochemical and immunologic parameters, modulating some inflammatory properties. Glutamine (GLN) is a conditionally essential amino acid that can modulate inflammatory properties. However, there is a lack of data evaluating the effects of DR and GLN supplementation, especially in relation to inflammatory cytokine production and the expression of transcription factors such as nuclear factor (NF)-κB. We subjected 3-mo-old male Balb/c mice to DR by reducing their food intake by 30%. DR animals lost weight and showed reduced levels of serum triacylglycerols, glucose, cholesterol, and calcium as well as a reduction in bone density. Additionally, blood, peritoneal, and spleen cellularity were reduced, lowering the number of peritoneal F4/80- and CD86-positive cells and the total number of splenic CD4- and CD8-positive cells. The production of interleukin (IL)-10 and the expression of NF-κB in splenic cells were not affected by DR or by GLN supplementation. However, peritoneal macrophages from DR animals showed reduced IL-12 and tumor necrosis factor-α production and increased IL-10 production with reduced phosphorylation of NF-κB expression. Additionally, GLN was able to modulate cytokine production by peritoneal cells from the control group, although no effects were observed in cells from the DR group. DR induces biochemical and immunologic changes, in particular by reducing IL-12 and tumor necrosis factor-α production by macrophages and clearly upregulating IL-10 production, whereas GLN supplementation did not modify these parameters in cells from DR animals. Copyright © 2017 Elsevier Inc. All rights reserved.

De Novo Mutations in CHD4, an ATP-Dependent Chromatin Remodeler Gene, Cause an Intellectual Disability Syndrome with Distinctive Dysmorphisms.

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Weiss, Karin; Terhal, Paulien A; Cohen, Lior; Bruccoleri, Michael; Irving, Melita; Martinez, Ariel F; Rosenfeld, Jill A; Machol, Keren; Yang, Yaping; Liu, Pengfei; Walkiewicz, Magdalena; Beuten, Joke; Gomez-Ospina, Natalia; Haude, Katrina; Fong, Chin-To; Enns, Gregory M; Bernstein, Jonathan A; Fan, Judith; Gotway, Garrett; Ghorbani, Mohammad; van Gassen, Koen; Monroe, Glen R; van Haaften, Gijs; Basel-Vanagaite, Lina; Yang, Xiang-Jiao; Campeau, Philippe M; Muenke, Maximilian

2016-10-06

Chromodomain helicase DNA-binding protein 4 (CHD4) is an ATP-dependent chromatin remodeler involved in epigenetic regulation of gene transcription, DNA repair, and cell cycle progression. Also known as Mi2β, CHD4 is an integral subunit of a well-characterized histone deacetylase complex. Here we report five individuals with de novo missense substitutions in CHD4 identified through whole-exome sequencing and web-based gene matching. These individuals have overlapping phenotypes including developmental delay, intellectual disability, hearing loss, macrocephaly, distinct facial dysmorphisms, palatal abnormalities, ventriculomegaly, and hypogonadism as well as additional findings such as bone fusions. The variants, c.3380G>A (p.Arg1127Gln), c.3443G>T (p.Trp1148Leu), c.3518G>T (p.Arg1173Leu), and c.3008G>A, (p.Gly1003Asp) (GenBank: NM_001273.3), affect evolutionarily highly conserved residues and are predicted to be deleterious. Previous studies in yeast showed the equivalent Arg1127 and Trp1148 residues to be crucial for SNF2 function. Furthermore, mutations in the same positions were reported in malignant tumors, and a de novo missense substitution in an equivalent arginine residue in the C-terminal helicase domain of SMARCA4 is associated with Coffin Siris syndrome. Cell-based studies of the p.Arg1127Gln and p.Arg1173Leu mutants demonstrate normal localization to the nucleus and HDAC1 interaction. Based on these findings, the mutations potentially alter the complex activity but not its formation. This report provides evidence for the role of CHD4 in human development and expands an increasingly recognized group of Mendelian disorders involving chromatin remodeling and modification. Published by Elsevier Inc.

Replication of alfalfa mosaic virus RNA 3 with movement and coat protein genes replaced by corresponding genes of Prunus necrotic ringspot ilarvirus.

Science.gov (United States)

Sánchez-Navarro, J A; Reusken, C B; Bol, J F; Pallás, V

1997-12-01

Alfalfa mosaic virus (AMV) and Prunus necrotic ringspot virus (PNRSV) are tripartite positive-strand RNA plant viruses that encode functionally similar translation products. Although the two viruses are phylogenetically closely related, they infect a very different range of natural hosts. The coat protein (CP) gene, the movement protein (MP) gene or both genes in AMV RNA 3 were replaced by the corresponding genes of PNRSV. The chimeric viruses were tested for heterologous encapsidation, replication in protoplasts from plants transformed with AMV replicase genes P1 and P2 (P12 plants) and for cell-to-cell transport in P12 plants. The chimeric viruses exhibited basic competence for encapsidation and replication in P12 protoplasts and for a low level of cell-to-cell movement in P12 plants. The potential involvement of the MP gene in determining host specificity in ilarviruses is discussed.

Role of transcription factor KLF11 and its diabetes-associated gene variants in pancreatic beta cell function

Science.gov (United States)

Neve, Bernadette; Fernandez-Zapico, Martin E.; Ashkenazi-Katalan, Vered; Dina, Christian; Hamid, Yasmin H.; Joly, Erik; Vaillant, Emmanuel; Benmezroua, Yamina; Durand, Emmanuelle; Bakaher, Nicolas; Delannoy, Valerie; Vaxillaire, Martine; Cook, Tiffany; Dallinga-Thie, Geesje M.; Jansen, Hans; Charles, Marie-Aline; Clément, Karine; Galan, Pilar; Hercberg, Serge; Helbecque, Nicole; Charpentier, Guillaume; Prentki, Marc; Hansen, Torben; Pedersen, Oluf; Urrutia, Raul; Melloul, Danielle; Froguel, Philippe

2005-01-01

KLF11 (TIEG2) is a pancreas-enriched transcription factor that has elicited significant attention because of its role as negative regulator of exocrine cell growth in vitro and in vivo. However, its functional role in the endocrine pancreas remains to be established. Here, we report, for the first time, to our knowledge, the characterization of KLF11 as a glucose-inducible regulator of the insulin gene. A combination of random oligonucleotide binding, EMSA, luciferase reporter, and chromatin immunoprecipitation assays shows that KLF11 binds to the insulin promoter and regulates its activity in beta cells. Genetic analysis of the KLF11 gene revealed two rare variants (Ala347Ser and Thr220Met) that segregate with diabetes in families with early-onset type 2 diabetes, and significantly impair its transcriptional activity. In addition, analysis of 1,696 type 2 diabetes mellitus and 1,776 normoglycemic subjects show a frequent polymorphic Gln62Arg variant that significantly associates with type 2 diabetes mellitus in North European populations (OR = 1.29, P = 0.00033). Moreover, this variant alters the corepressor mSin3A-binding activity of KLF11, impairs the activation of the insulin promoter and shows lower levels of insulin expression in pancreatic beta cells. In addition, subjects carrying the Gln62Arg allele show decreased plasma insulin after an oral glucose challenge. Interestingly, all three nonsynonymous KLF11 variants show increased repression of the catalase 1 promoter, suggesting a role in free radical clearance that may render beta cells more sensitive to oxidative stress. Thus, both functional and genetic analyses reveal that KLF11 plays a role in the regulation of pancreatic beta cell physiology, and its variants may contribute to the development of diabetes. PMID:15774581

Radiochemical identification of the kil gene product of bacteriophage lambda

International Nuclear Information System (INIS)

Greer, H.; Ausubel, F.M.

1979-01-01

The coliphage lambda kil gene product has been identified using a differential labeling technique . The kil gene polypeptide has a molecular weight of about 16,000, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Gel filtration of the kil protein indicates that it may exist as a tetramer in native form

Data of evolutionary structure change: 1DFFA-3CPMA [Confc[Archive

Lifescience Database Archive (English)

Full Text Available 1DFFA-3CPMA 1DFF 3CPM A A -----SVLQVLHIPDERLRKVAKPVEEVNAEIQRIVDDM...line> 3CPM A 3CPMA THR CA 218 GLY CA 249 ILE CA 245 3CPM... A 3CPMA IKKYSDKLVPFILE CA 305 ARG CA 316 GLN CA 241 3CPM

Co-expression of the Thermotoga neapolitana aglB gene with an upstream 3'-coding fragment of the malG gene improves enzymatic characteristics of recombinant AglB cyclomaltodextrinase.

Science.gov (United States)

Lunina, Natalia A; Agafonova, Elena V; Chekanovskaya, Lyudmila A; Dvortsov, Igor A; Berezina, Oksana V; Shedova, Ekaterina N; Kostrov, Sergey V; Velikodvorskaya, Galina A

2007-07-01

A cluster of Thermotoga neapolitana genes participating in starch degradation includes the malG gene of sugar transport protein and the aglB gene of cyclomaltodextrinase. The start and stop codons of these genes share a common overlapping sequence, aTGAtg. Here, we compared properties of expression products of three different constructs with aglB from T. neapolitana. The first expression vector contained the aglB gene linked to an upstream 90-bp 3'-terminal region of the malG gene with the stop codon overlapping with the start codon of aglB. The second construct included the isolated coding sequence of aglB with two tandem potential start codons. The expression product of this construct in Escherichia coli had two tandem Met residues at its N terminus and was characterized by low thermostability and high tendency to aggregate. In contrast, co-expression of aglB and the 3'-terminal region of malG (the first construct) resulted in AglB with only one N-terminal Met residue and a much higher specific activity of cyclomaltodextrinase. Moreover, the enzyme expressed by such a construct was more thermostable and less prone to aggregation. The third construct was the same as the second one except that it contained only one ATG start codon. The product of its expression had kinetic and other properties similar to those of the enzyme with only one N-terminal Met residue.

Synthetic operon for (R,R)-2,3-butanediol production in Bacillus subtilis and Escherichia coli.

Science.gov (United States)

de Oliveira, Rafael R; Nicholson, Wayne L

2016-01-01

To reduce dependence on petroleum, an alternative route to production of the chemical feedstock 2,3-butanediol (2,3-BD) from renewable lignocellulosic sources is desirable. In this communication, the genes encoding the pathway from pyruvate to 2,3-BD (alsS, alsD, and bdhA encoding acetolactate synthase, acetolactate decarboxylase, and butanediol dehydrogenase, respectively) from Bacillus subtilis were engineered into a single tricistronic operon under control of the isopropyl β-D-1-thiogalactopyranoside (IPTG)-inducible Pspac promoter in a shuttle plasmid capable of replication and expression in either B. subtilis or Escherichia coli. We describe the construction and performance of a shuttle plasmid carrying the IPTG-inducible synthetic operon alsSDbdhA coding for 2,3-BD pathway capable of (i) expression in two important representative model microorganisms, the gram-positive B. subtilis and the gram-negative E. coli; (ii) increasing 2,3-BD production in B. subtilis; and (iii) successfully introducing the B. subtilis 2,3-BD pathway into E. coli. The synthetic alsSDbdhA operon constructed using B. subtilis native genes not only increased the 2,3-BD production in its native host but also efficiently expressed the pathway in the heterologous organism E. coli. Construction of an efficient shuttle plasmid will allow investigation of 2,3-BD production performance in related organisms with industrial potential for production of bio-based chemicals.

Homozygous sequence variants in the WNT10B gene underlie split hand/foot malformation

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Asmat Ullah

2018-01-01

Full Text Available Abstract Split-hand/split-foot malformation (SHFM, also known as ectrodactyly is a rare genetic disorder. It is a clinically and genetically heterogeneous group of limb malformations characterized by absence/hypoplasia and/or median cleft of hands and/or feet. To date, seven genes underlying SHFM have been identified. This study described four consanguineous families (A-D segregating SHFM in an autosomal recessive manner. Linkage in the families was established to chromosome 12p11.1–q13.13 harboring WNT10B gene. Sequence analysis identified a novel homozygous nonsense variant (p.Gln154* in exon 4 of the WNT10B gene in two families (A and B. In the other two families (C and D, a previously reported variant (c.300_306dupAGGGCGG; p.Leu103Argfs*53 was detected. This study further expands the spectrum of the sequence variants reported in the WNT10B gene, which result in the split hand/foot malformation.

Genome-engineered Sinorhizobium meliloti for the production of poly(lactic-co-3-hydroxybutyric) acid copolymer.

Science.gov (United States)

Tran, Tam T; Charles, Trevor C

2016-02-01

Economically competitive commercial production of biodegradable bioplastics with desirable properties is an important goal. In this study, we demonstrate the use of chromosome engineering of an alternative bacterial host, Sinorhizobium meliloti, for production of the copolymer, poly(lactate-co-3-hydroxybutyrate). Codon-optimized genes for 2 previously engineered enzymes, Clostridium propionicum propionate CoA transferase (Pct532Cp) and Pseudomonas sp. strain MBEL 6-19 polyhydroxyalkanoate (PHA) synthase 1 (PhaC1400Ps6-19), were introduced into S. meliloti Rm1021 by chromosome integration, replacing the native phbC gene. On the basis of phenotypic analysis and detection of polymer product by gas chromatography analysis, synthesis and accumulation of the copolymer was confirmed. The chromosome integrant strain, with the introduced genes under the control of the native phbC promoter, is able to produce over 15% cell dry mass of poly(lactate-co-3-hydroxybutyrate), containing 30 mol% lactate, from growth on mannitol. We were also able to purify the polymer from the culture and confirm the structure by NMR and GC-MS. To our knowledge, this is the first demonstration of production of this copolymer in the Alphaproteobacteria. Further optimization of this system may eventually yield strains that are able to produce economically viable commercial product.

The R2R3-MYB-like regulatory factor EOBI, acting downstream of EOBII, regulates scent production by activating ODO1 and structural scent-related genes in petunia.

Science.gov (United States)

Spitzer-Rimon, Ben; Farhi, Moran; Albo, Boaz; Cna'ani, Alon; Ben Zvi, Michal Moyal; Masci, Tania; Edelbaum, Orit; Yu, Yixun; Shklarman, Elena; Ovadis, Marianna; Vainstein, Alexander

2012-12-01

Flower scent is a highly dynamic trait, under developmental, spatial, and diurnal regulation. The mechanism governing scent production is only beginning to be unraveled. In petunia (Petunia hybrida), EMISSION OF BENZENOIDS II (EOBII) controls transcription of both the shikimate pathway-regulating MYB factor ODORANT1 (ODO1) and phenylpropanoid scent-related structural genes. A promoter-activation screen identified an R2R3-MYB-like regulatory factor of phenylpropanoid volatile biosynthesis acting downstream of EOBII, designated EOBI. EOBI silencing led to downregulation of ODO1 and numerous structural scent-related genes from both the shikimate and phenylpropanoid pathways. The ability of EOBI to directly activate ODO1, as revealed by electrophoretic mobility shift assay and yeast one-hybrid analysis, place EOBI upstream of ODO1 in regulating substrate availability for volatile biosynthesis. Interestingly, ODO1-silenced transgenic petunia flowers accumulated higher EOBI transcript levels than controls, suggesting a complex feedback loop between these regulatory factors. The accumulation pattern of EOBI transcript relative to EOBII and ODO1, and the effect of up/downregulation of EOBII on transcript levels of EOBI and ODO1, further support these factors' hierarchical relationships. The dependence of scent production on EOBI expression and its direct interaction with both regulatory and structural genes provide evidence for EOBI's wide-ranging involvement in the production of floral volatiles.

Identification of immediate early gene products of bovine herpes virus 1 (BHV-1) as dominant antigens recognized by CD8 T cells in immune cattle

DEFF Research Database (Denmark)

Hart, Jane; MacHugh, Niall D.; Sheldrake, Tara

2017-01-01

candidate viral gene products with CD8 T-cell lines from 3 BHV-1-immune cattle of defined MHC genotypes identified 4 antigens, including 3 immediate early (IE) gene products (ICP4, ICP22 and Circ) and a tegument protein (UL49). Identification of the MHC restriction specificities revealed that the antigens...... cases refined, the identity of the epitopes. Analyses of the epitope specificity of the CD8 T-cell lines showed that a large component of the response is directed against these IE epitopes. The results indicate that these IE gene products are dominant targets of the CD8 T-cell response in BHV...

Comparative genomic analysis of the multispecies probiotic-marketed product VSL#3.

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François P Douillard

Full Text Available Several probiotic-marketed formulations available for the consumers contain live lactic acid bacteria and/or bifidobacteria. The multispecies product commercialized as VSL#3 has been used for treating various gastro-intestinal disorders. However, like many other products, the bacterial strains present in VSL#3 have only been characterized to a limited extent and their efficacy as well as their predicted mode of action remain unclear, preventing further applications or comparative studies. In this work, the genomes of all eight bacterial strains present in VSL#3 were sequenced and characterized, to advance insights into the possible mode of action of this product and also to serve as a basis for future work and trials. Phylogenetic and genomic data analysis allowed us to identify the 7 species present in the VSL#3 product as specified by the manufacturer. The 8 strains present belong to the species Streptococcus thermophilus, Lactobacillus acidophilus, Lactobacillus paracasei, Lactobacillus plantarum, Lactobacillus helveticus, Bifidobacterium breve and B. animalis subsp. lactis (two distinct strains. Comparative genomics revealed that the draft genomes of the S. thermophilus and L. helveticus strains were predicted to encode most of the defence systems such as restriction modification and CRISPR-Cas systems. Genes associated with a variety of potential probiotic functions were also identified. Thus, in the three Bifidobacterium spp., gene clusters were predicted to encode tight adherence pili, known to promote bacteria-host interaction and intestinal barrier integrity, and to impact host cell development. Various repertoires of putative signalling proteins were predicted to be encoded by the genomes of the Lactobacillus spp., i.e. surface layer proteins, LPXTG-containing proteins, or sortase-dependent pili that may interact with the intestinal mucosa and dendritic cells. Taken altogether, the individual genomic characterization of the strains

Glutamine reduces myocardial cell apoptosis in a rat model of sepsis by promoting expression of heat shock protein 90.

Science.gov (United States)

Li, Wanxia; Tao, Shaoyu; Wu, Qinghua; Wu, Tao; Tao, Ran; Fan, Jun

2017-12-01

Myocardial cell injury and cardiac myocyte apoptosis are associated with sepsis. Glutamine (Gln) has been reported to repair myocardial cell injury. The aim of this study was to explore the role of Gln on cardiac myocytes in a cecal ligation and puncture (CLP) model of sepsis in Wistar rats. Following induction of sepsis in a CLP rat model, viral encoding heat shock protein 90 (Hsp90) gene and Hsp90dsDNA were designed to express and knockdown Hsp90, respectively. Rat cardiac tissues were examined histologically, and apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling staining. The expression of B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein, Hsp90, p53 upregulated modulator of apoptosis, and p53 was measured by western blotting and real-time polymerase chain reaction. Caspase-3, caspase-8, and caspase-9 were detected by enzyme-linked immunosorbent assay. Rat cardiac myocyte damage induced by CLP was reduced by Gln treatment and Hsp90 overexpression, and these changes were reversed by Hsp90 knockdown. Bcl-2 expression, Bcl-2-associated X protein, p53, p53 upregulated modulator of apoptosis, caspase-8, caspase-9, and caspase-3 activities were significantly upregulated in the CLP model, which were reduced by Gln treatment and Hsp90 overexpression. Gln reduced apoptosis of cardiac myocytes in a rat model of sepsis, by promoting Hsp90 expression. Further studies are needed to determine the possible therapeutic action of Gln in sepsis in human tissue. Copyright © 2017 Elsevier Inc. All rights reserved.

1,3-Propanediol dehydrogenases in Lactobacillus reuteri: impact on central metabolism and 3-hydroxypropionaldehyde production.

Science.gov (United States)

Stevens, Marc J A; Vollenweider, Sabine; Meile, Leo; Lacroix, Christophe

2011-08-03

Lactobacillus reuteri metabolizes glycerol to 3-hydroxypropionaldehyde (3-HPA) and further to 1,3-propanediol (1,3-PDO), the latter step catalysed by a propanediol dehydrogenase (PDH). The last step in this pathway regenerates NAD+ and enables therefore the energetically more favourable production of acetate over ethanol during growth on glucose. A search throughout the genome of L. reuteri DSM 20016 revealed two putative PDHs encoded by ORFs lr_0030 and lr_1734. ORF lr_1734 is situated in the pdu operon encoding the glycerol conversion machinery and therefore likely involved in 1,3-PDO formation. ORF lr_0030 has not been associated with PDH-activity so far. To elucidate the role of these two PDHs, gene deletion mutant strains were constructed. Growth behaviour on glucose was comparable between the wild type and both mutant strains. However, on glucose + glycerol, the exponential growth rate of Δlr_0030 was lower compared to the wild type and the lr_1734 mutant. Furthermore, glycerol addition resulted in decreased ethanol production in the wild type and Δlr_1734, but not in Δlr_0030. PDH activity measurements using 3-HPA as a substrate revealed lower activity of Δlr_0030 extracts from exponential growing cells compared to wild type and Δlr_1734 extracts.During biotechnological 3-HPA production using non-growing cells, the ratio 3-HPA to 1,3-PDO was approximately 7 in the wild type and Δlr_0030, whereas this ratio was 12.5 in the mutant Δlr_1734. The enzyme encoded by lr_0030 plays a pivotal role in 3-HPA conversion in exponential growing L. reuteri cells. The enzyme encoded by lr_1734 is active during 3-HPA production by non-growing cells and this enzyme is a useful target to enhance 3-HPA production and minimize formation of the by-product 1,3-PDO.

Super gene alternation of magnetite and pyrite and the role of their alternation products in the fixation of uranium from the circulating media. Vol. 3

Energy Technology Data Exchange (ETDEWEB)

El-Gemmizi, M A [Nuclear Materials Authority, Cairo, (Egypt)

1996-03-01

In most of the Egyptian altered radioactive granites, highly magnetic heavy particles were found to be radioactive. They are a mixture of several iron oxide minerals which are products of super gene alternation of the preexisting hypo gene iron-bearing minerals especially magnetite and pyrite. The end products of this super gene alternation are mainly hydrated iron oxide minerals limonite and/or goethite. During the alternation, deformation and defects in the mineral structure took place, thereby promoting diffusion of the substitutional and interstitial ions (uranium) towards these sites. The mechanism of the alternation of the hypo gene iron-bearing minerals, magnetite and pyrite to form the secondary mineral hematite, limonite and goethite; and the role of these secondary minerals in fixing uranium from the circulating media, and as indicators to the radioactivity of the host rocks are discussed. 2 figs.

Super gene alternation of magnetite and pyrite and the role of their alternation products in the fixation of uranium from the circulating media. Vol. 3

International Nuclear Information System (INIS)

El-Gemmizi, M.A.

1996-01-01

In most of the Egyptian altered radioactive granites, highly magnetic heavy particles were found to be radioactive. They are a mixture of several iron oxide minerals which are products of super gene alternation of the preexisting hypo gene iron-bearing minerals especially magnetite and pyrite. The end products of this super gene alternation are mainly hydrated iron oxide minerals limonite and/or goethite. During the alternation, deformation and defects in the mineral structure took place, thereby promoting diffusion of the substitutional and interstitial ions (uranium) towards these sites. The mechanism of the alternation of the hypo gene iron-bearing minerals, magnetite and pyrite to form the secondary mineral hematite, limonite and goethite; and the role of these secondary minerals in fixing uranium from the circulating media, and as indicators to the radioactivity of the host rocks are discussed. 2 figs

STAT3 Target Genes Relevant to Human Cancers

International Nuclear Information System (INIS)

Carpenter, Richard L.; Lo, Hui-Wen

2014-01-01

Since its discovery, the STAT3 transcription factor has been extensively studied for its function as a transcriptional regulator and its role as a mediator of development, normal physiology, and pathology of many diseases, including cancers. These efforts have uncovered an array of genes that can be positively and negatively regulated by STAT3, alone and in cooperation with other transcription factors. Through regulating gene expression, STAT3 has been demonstrated to play a pivotal role in many cellular processes including oncogenesis, tumor growth and progression, and stemness. Interestingly, recent studies suggest that STAT3 may behave as a tumor suppressor by activating expression of genes known to inhibit tumorigenesis. Additional evidence suggested that STAT3 may elicit opposing effects depending on cellular context and tumor types. These mixed results signify the need for a deeper understanding of STAT3, including its upstream regulators, parallel transcription co-regulators, and downstream target genes. To help facilitate fulfilling this unmet need, this review will be primarily focused on STAT3 downstream target genes that have been validated to associate with tumorigenesis and/or malignant biology of human cancers

Improvement of oxytetracycline production mediated via cooperation of resistance genes in Streptomyces rimosus.

Science.gov (United States)

Yin, Shouliang; Wang, Xuefeng; Shi, Mingxin; Yuan, Fang; Wang, Huizhuan; Jia, Xiaole; Yuan, Fang; Sun, Jinliang; Liu, Tiejun; Yang, Keqian; Zhang, Yuxiu; Fan, Keqiang; Li, Zilong

2017-09-01

Increasing the self-resistance levels of Streptomyces is an effective strategy to improve the production of antibiotics. To increase the oxytetracycline (OTC) production in Streptomyces rimosus, we investigated the cooperative effect of three co-overexpressing OTC resistance genes: one gene encodes a ribosomal protection protein (otrA) and the other two express efflux proteins (otrB and otrC). Results indicated that combinational overexpression of otrA, otrB, and otrC (MKABC) exerted a synergetic effect. OTC production increased by 179% in the recombinant strain compared with that of the wild-type strain M4018. The resistance level to OTC was increased by approximately two-fold relative to the parental strain, thereby indicating that applying the cooperative effect of self-resistance genes is useful to improve OTC production. Furthermore, the previously identified cluster-situated activator OtcR was overexpressed in MKABC in constructing the recombinant strain MKRABC; such strain can produce OTC of approximately 7.49 g L -1 , which represents an increase of 19% in comparison with that of the OtcR-overexpressing strain alone. Our work showed that the cooperative overexpression of self-resistance genes is a promising strategy to enhance the antibiotics production in Streptomyces.

Biological evaluation of 177Lu-labeled DOTA-Ala(SO3H)-Aminooctanoyl-Gln-Trp-Ala-Val-N methyl Gly-His-Statine-Leu-NH2 for gastrin-releasing peptide receptor-positive prostate tumor targeting

International Nuclear Information System (INIS)

Lim, Jae Cheong; Cho, Eun Ha; Kim, Jin Joo; Choi, Sang Mu; Lee, So young; Nam, Sung Soo; Park, Ul Jae; Park, Soo Hyun

2015-01-01

Bombesin binds with selectivity and high affinity to a Gastrin-releasing peptide receptor (GRPR), which is highly overexpressed in prostate cancer cells. The present study describes the in vitro and in vivo biological characteristics of DOTA-Ala(SO 3 H)-Aminooctanoyl-Gln-Trp-Ala-Val-N methyl Gly-His-Statine-Leu-NH 2 (DOTA-sBBNA), an antagonist analogue of bombesin peptide for the targeting of GRPR. DOTA-sBBNA was synthesized and labeled with 177 Lu as previously published. A saturation assay on PC-3 human prostate cancer cells revealed that the Kd value of the radiolabeled peptide was 1.88 nM with a maximum binding capacity (Bmax) of 289.3 fmol/10 6 cells. The radio-peptide slowly internalized, and 24.4 ± 0.5% of the total binding was internalized in 4 hr. Biodistribution studies were conducted in healthy and PC-3 xenografted balb/c mice, which showed high uptake and retention of tumor-associated radioactivity in PC-3 xenografted mice. The tumor-to-blood ratio was 126.02 ± 9.36 at 1.5 hr p.i., and was increased to 216.33 ± 61.58 at 24 hr p.i., which means that the radiolabeled peptide was highly accumulated in a tumor and rapidly cleared from the blood pool. The GRPR is also over-expressed in Korean prostate cancer patients. These results suggest that this 177 Lu-labeled peptide has promising characteristics for application in nuclear medicine, namely for the diagnosis and treatment of GRPR over-expressing prostate tumors

Mutations in the LHX2 gene are not a frequent cause of micro/anophthalmia.

Science.gov (United States)

Desmaison, Annaïck; Vigouroux, Adeline; Rieubland, Claudine; Peres, Christine; Calvas, Patrick; Chassaing, Nicolas

2010-12-18

Microphthalmia and anophthalmia are at the severe end of the spectrum of abnormalities in ocular development. A few genes (orthodenticle homeobox 2 [OTX2], retina and anterior neural fold homeobox [RAX], SRY-box 2 [SOX2], CEH10 homeodomain-containing homolog [CHX10], and growth differentiation factor 6 [GDF6]) have been implicated mainly in isolated micro/anophthalmia but causative mutations of these genes explain less than a quarter of these developmental defects. The essential role of the LIM homeobox 2 (LHX2) transcription factor in early eye development has recently been documented. We postulated that mutations in this gene could lead to micro/anophthalmia, and thus performed molecular screening of its sequence in patients having micro/anophthalmia. Seventy patients having non-syndromic forms of colobomatous microphthalmia (n=25), isolated microphthalmia (n=18), or anophthalmia (n=17), and syndromic forms of micro/anophthalmia (n=10) were included in this study after negative molecular screening for OTX2, RAX, SOX2, and CHX10 mutations. Mutation screening of LHX2 was performed by direct sequencing of the coding sequences and intron/exon boundaries. Two heterozygous variants of unknown significance (c.128C>G [p.Pro43Arg]; c.776C>A [p.Pro259Gln]) were identified in LHX2 among the 70 patients. These variations were not identified in a panel of 100 control patients of mixed origins. The variation c.776C>A (p.Pro259Gln) was considered as non pathogenic by in silico analysis, while the variation c.128C>G (p.Pro43Arg) considered as deleterious by in silico analysis and was inherited from the asymptomatic father. Mutations in LHX2 do not represent a frequent cause of micro/anophthalmia.

Genome mining of Streptomyces scabrisporus NF3 reveals symbiotic features including genes related to plant interactions

Science.gov (United States)

Rodríguez-Luna, Stefany Daniela; Cruz Vázquez, Angélica Patricia; Jiménez Suárez, Verónica; Rodríguez-Sanoja, Romina; Alvarez-Buylla, Elena R.; Sánchez, Sergio

2018-01-01

Endophytic bacteria are wide-spread and associated with plant physiological benefits, yet their genomes and secondary metabolites remain largely unidentified. In this study, we explored the genome of the endophyte Streptomyces scabrisporus NF3 for discovery of potential novel molecules as well as genes and metabolites involved in host interactions. The complete genomes of seven Streptomyces and three other more distantly related bacteria were used to define the functional landscape of this unique microbe. The S. scabrisporus NF3 genome is larger than the average Streptomyces genome and not structured for an obligate endosymbiotic lifestyle; this and the fact that can grow in R2YE media implies that it could include a soil-living stage. The genome displays an enrichment of genes associated with amino acid production, protein secretion, secondary metabolite and antioxidants production and xenobiotic degradation, indicating that S. scabrisporus NF3 could contribute to the metabolic enrichment of soil microbial communities and of its hosts. Importantly, besides its metabolic advantages, the genome showed evidence for differential functional specificity and diversification of plant interaction molecules, including genes for the production of plant hormones, stress resistance molecules, chitinases, antibiotics and siderophores. Given the diversity of S. scabrisporus mechanisms for host upkeep, we propose that these strategies were necessary for its adaptation to plant hosts and to face changes in environmental conditions. PMID:29447216

Genome mining of Streptomyces scabrisporus NF3 reveals symbiotic features including genes related to plant interactions.

Directory of Open Access Journals (Sweden)

Corina Diana Ceapă

Full Text Available Endophytic bacteria are wide-spread and associated with plant physiological benefits, yet their genomes and secondary metabolites remain largely unidentified. In this study, we explored the genome of the endophyte Streptomyces scabrisporus NF3 for discovery of potential novel molecules as well as genes and metabolites involved in host interactions. The complete genomes of seven Streptomyces and three other more distantly related bacteria were used to define the functional landscape of this unique microbe. The S. scabrisporus NF3 genome is larger than the average Streptomyces genome and not structured for an obligate endosymbiotic lifestyle; this and the fact that can grow in R2YE media implies that it could include a soil-living stage. The genome displays an enrichment of genes associated with amino acid production, protein secretion, secondary metabolite and antioxidants production and xenobiotic degradation, indicating that S. scabrisporus NF3 could contribute to the metabolic enrichment of soil microbial communities and of its hosts. Importantly, besides its metabolic advantages, the genome showed evidence for differential functional specificity and diversification of plant interaction molecules, including genes for the production of plant hormones, stress resistance molecules, chitinases, antibiotics and siderophores. Given the diversity of S. scabrisporus mechanisms for host upkeep, we propose that these strategies were necessary for its adaptation to plant hosts and to face changes in environmental conditions.

Gene Cluster Responsible for Secretion of and Immunity to Multiple Bacteriocins, the NKR-5-3 Enterocins

Science.gov (United States)

Ishibashi, Naoki; Himeno, Kohei; Masuda, Yoshimitsu; Perez, Rodney Honrada; Iwatani, Shun; Wilaipun, Pongtep; Leelawatcharamas, Vichien; Nakayama, Jiro; Sonomoto, Kenji

2014-01-01

Enterococcus faecium NKR-5-3, isolated from Thai fermented fish, is characterized by the unique ability to produce five bacteriocins, namely, enterocins NKR-5-3A, -B, -C, -D, and -Z (Ent53A, Ent53B, Ent53C, Ent53D, and Ent53Z). Genetic analysis with a genome library revealed that the bacteriocin structural genes (enkA [ent53A], enkC [ent53C], enkD [ent53D], and enkZ [ent53Z]) that encode these peptides (except for Ent53B) are located in close proximity to each other. This NKR-5-3ACDZ (Ent53ACDZ) enterocin gene cluster (approximately 13 kb long) includes certain bacteriocin biosynthetic genes such as an ABC transporter gene (enkT), two immunity genes (enkIaz and enkIc), a response regulator (enkR), and a histidine protein kinase (enkK). Heterologous-expression studies of enkT and ΔenkT mutant strains showed that enkT is responsible for the secretion of Ent53A, Ent53C, Ent53D, and Ent53Z, suggesting that EnkT is a wide-range ABC transporter that contributes to the effective production of these bacteriocins. In addition, EnkIaz and EnkIc were found to confer self-immunity to the respective bacteriocins. Furthermore, bacteriocin induction assays performed with the ΔenkRK mutant strain showed that EnkR and EnkK are regulatory proteins responsible for bacteriocin production and that, together with Ent53D, they constitute a three-component regulatory system. Thus, the Ent53ACDZ gene cluster is essential for the biosynthesis and regulation of NKR-5-3 enterocins, and this is, to our knowledge, the first report that demonstrates the secretion of multiple bacteriocins by an ABC transporter. PMID:25149515

The SKP1-like gene family of Arabidopsis exhibits a high degree of differential gene expression and gene product interaction during development.

Directory of Open Access Journals (Sweden)

Mohammad H Dezfulian

Full Text Available The Arabidopsis thaliana genome encodes several families of polypeptides that are known or predicted to participate in the formation of the SCF-class of E3-ubiquitin ligase complexes. One such gene family encodes the Skp1-like class of polypeptide subunits, where 21 genes have been identified and are known to be expressed in Arabidopsis. Phylogenetic analysis based on deduced polypeptide sequence organizes the family of ASK proteins into 7 clades. The complexity of the ASK gene family, together with the close structural similarity among its members raises the prospect of significant functional redundancy among select paralogs. We have assessed the potential for functional redundancy within the ASK gene family by analyzing an expanded set of criteria that define redundancy with higher resolution. The criteria used include quantitative expression of locus-specific transcripts using qRT-PCR, assessment of the sub-cellular localization of individual ASK:YFP auto-fluorescent fusion proteins expressed in vivo as well as the in planta assessment of individual ASK-F-Box protein interactions using bimolecular fluorescent complementation techniques in combination with confocal imagery in live cells. The results indicate significant functional divergence of steady state transcript abundance and protein-protein interaction specificity involving ASK proteins in a pattern that is poorly predicted by sequence-based phylogeny. The information emerging from this and related studies will prove important for defining the functional intersection of expression, localization and gene product interaction that better predicts the formation of discrete SCF complexes, as a prelude to investigating their molecular mode of action.

Candidate genes for drought tolerance and improved productivity in ...

Indian Academy of Sciences (India)

Madhu

Improving drought tolerance and productivity is one of the most difficult tasks for ... Keywords. Candidate gene; mapping population; polymerase chain reaction; single marker analysis. .... ple and the mean value computed. 2.4 Isolation of DNA.

A novel hemoglobin variant found on the α1 chain: Hb KSVGH (HBA1: p.Lys57_Gly58insSerHisGlySerAlaGlnValLys).

Science.gov (United States)

Wang, Mei-Chun; Tsai, Kuo-Wang; Chu, Chih-Hsun; Yu, Ming-Sun; Lam, Hing-Chung

2015-01-01

Glycosylated hemoglobin (Hb A1C) is a crucial indicator for the long-term control and the diagnosis of diabetes. However, the presence of hemoglobin (Hb) variants may affect the measured value of Hb A1C and result in an abnormal graph trend and inconsistency between the clinical blood sugar test and Hb A1C values. In this study, laboratory data of 41,267 patients with diabetes were collected. The Hb A1C levels and the graph results were examined. We identified 74 cases containing abnormal Hb A1C graph trends. The conducted blood cell counts and capillary Hb electrophoresis were used to analyze Hb variants. We also determined gene variation for the Hb variants by a sequence approach. Fifteen different types of Hb variants were identified in this study. Among these, we found a novel variant in which the α1 subunit of Hb showed an insertion of 24 nucleotides (nts) between the 56th and 57th residues. We named this novel variant Hb Kaohsiung Veterans General Hospital (Hb KSVGH) (HBA1: p.Lys57_Gly58insSerHisGlySerAlaGlnValLys).

The identification of aluminium-resistance genes provides opportunities for enhancing crop production on acid soils.

Science.gov (United States)

Ryan, P R; Tyerman, S D; Sasaki, T; Furuichi, T; Yamamoto, Y; Zhang, W H; Delhaize, E

2011-01-01

Acid soils restrict plant production around the world. One of the major limitations to plant growth on acid soils is the prevalence of soluble aluminium (Al(3+)) ions which can inhibit root growth at micromolar concentrations. Species that show a natural resistance to Al(3+) toxicity perform better on acid soils. Our understanding of the physiology of Al(3+) resistance in important crop plants has increased greatly over the past 20 years, largely due to the application of genetics and molecular biology. Fourteen genes from seven different species are known to contribute to Al(3+) tolerance and resistance and several additional candidates have been identified. Some of these genes account for genotypic variation within species and others do not. One mechanism of resistance which has now been identified in a range of species relies on the efflux of organic anions such as malate and citrate from roots. The genes controlling this trait are members of the ALMT and MATE families which encode membrane proteins that facilitate organic anion efflux across the plasma membrane. Identification of these and other resistance genes provides opportunities for enhancing the Al(3+) resistance of plants by marker-assisted breeding and through biotechnology. Most attempts to enhance Al(3+) resistance in plants with genetic engineering have targeted genes that are induced by Al(3+) stress or that are likely to increase organic anion efflux. In the latter case, studies have either enhanced organic anion synthesis or increased organic anion transport across the plasma membrane. Recent developments in this area are summarized and the structure-function of the TaALMT1 protein from wheat is discussed.

Relationship between fumonisin production and FUM gene expression in Fusarium verticillioides under different environmental conditions.

Science.gov (United States)

Fanelli, Francesca; Iversen, Anita; Logrieco, Antonio F; Mulè, Giuseppina

2013-01-01

Fusarium verticillioides is the main source of fumonisins, a group of mycotoxins that can contaminate maize-based food and feed and cause diseases in humans and animals. The study of the effect of different environmental conditions on toxin production should provide information that can be used to develop strategies to minimize the risk. This study analysed the effect of temperature (15°C-35°C), water activity (a(w): 0.999-0.93), salinity (0-125 g l(-1) NaCl) and pH (5-8) on the growth and production of fumonisins B(1) (FB1), B(2) (FB2) and B(3) (FB3) and the expression of FUM1 and FUM21 in F. verticillioides. The highest growth rate was measured at 25°C, a(w) of 0.998-0.99 and 0-25 g l(-1) of NaCl. Optimal conditions for fumonisin production were 30°C, a(w) of 0.99, 25 g l(-1) of NaCl and pH 5; nevertheless, the strain showed a good adaptability and was able to produce moderate levels of fumonisins under a wide range of conditions. Gene expression mirrored fumonisin production profile under all conditions with the exception of temperature: FUM1 and FUM21 expression was highest at 15°C, while maximum fumonisin production was at 30°C. These data indicate that a post-transcriptional regulation mechanism could account for the different optimal temperatures for FUM gene expression and fumonisin production.

Deficiency in nucleotide excision repair family gene activity, especially ERCC3, is associated with non-pigmented hair fiber growth.

Directory of Open Access Journals (Sweden)

Mei Yu

Full Text Available We conducted a microarray study to discover gene expression patterns associated with a lack of melanogenesis in non-pigmented hair follicles (HF by microarray. Pigmented and non-pigmented HFs were collected and micro-dissected into the hair bulb (HB and the upper hair sheaths (HS including the bulge region. In comparison to pigmented HS and HBs, nucleotide excision repair (NER family genes ERCC1, ERCC2, ERCC3, ERCC4, ERCC5, ERCC6, XPA, NTPBP, HCNP, DDB2 and POLH exhibited statistically significantly lower expression in non- pigmented HS and HBs. Quantitative PCR verified microarray data and identified ERCC3 as highly differentially expressed. Immunohistochemistry confirmed ERCC3 expression in HF melanocytes. A reduction in ERCC3 by siRNA interference in human melanocytes in vitro reduced their tyrosinase production ability. Our results suggest that loss of NER gene function is associated with a loss of melanin production capacity. This may be due to reduced gene transcription and/or reduced DNA repair in melanocytes which may eventually lead to cell death. These results provide novel information with regard to melanogenesis and its regulation.

Coregulation of terpenoid pathway genes and prediction of isoprene production in Bacillus subtilis using transcriptomics

Energy Technology Data Exchange (ETDEWEB)

Hess, Becky M.; Xue, Junfeng; Markillie, Lye Meng; Taylor, Ronald C.; Wiley, H. S.; Ahring, Birgitte K.; Linggi, Bryan E.

2013-06-19

The isoprenoid pathway converts pyruvate to isoprene and related isoprenoid compounds in plants and some bacteria. Currently, this pathway is of great interest because of the critical role that isoprenoids play in basic cellular processes as well as the industrial value of metabolites such as isoprene. Although the regulation of several pathway genes has been described, there is a paucity of information regarding the system level regulation and control of the pathway. To address this limitation, we examined Bacillus subtilis grown under multiple conditions and then determined the relationship between altered isoprene production and the pattern of gene expression. We found that terpenoid genes appeared to fall into two distinct subsets with opposing correlations with respect to the amount of isoprene produced. The group whose expression levels positively correlated with isoprene production included dxs, the gene responsible for the commitment step in the pathway, as well as ispD, and two genes that participate in the mevalonate pathway, yhfS and pksG. The subset of terpenoid genes that inversely correlated with isoprene production included ispH, ispF, hepS, uppS, ispE, and dxr. A genome wide partial least squares regression model was created to identify other genes or pathways that contribute to isoprene production. This analysis showed that a subset of 213 regulated genes was sufficient to create a predictive model of isoprene production under different conditions and showed correlations at the transcriptional level. We conclude that gene expression levels alone are sufficiently informative about the metabolic state of a cell that produces increased isoprene and can be used to build a model which accurately predicts production of this secondary metabolite across many simulated environmental conditions.

Identification of the 14-3-3 gene family in Rafflesia cantleyi

Science.gov (United States)

Rosli, Khadijah; Wan, Kiew-Lian

2018-04-01

Rafflesia is known to be the largest flower in the world. Due to its size and appearance, it is considered to be very unique. Little is known about the molecular biology of this rare parasitic flowering plant as it is very difficult to locate and has a short life-span as a flower. Physiological activities in plants are regulated by signalling regulators such as the members of the 14-3-3 gene family. The number of members of this gene family varies in plants and there are thirteen known members in Arabidopsis thaliana. Their role is to bind to phosphorylated targets to complete signal transduction processes. Sequence comparison using BLAST of transcriptome data from three different Rafflesia cantleyi floral bud stages against the Swissprot database revealed 27 transcripts annotated as members of this gene family. All of the transcripts were expressed during floral bud stage 1 (S1) while 14 and four transcripts were expressed during floral bud stages 2 (S2) and 3 (S3), respectively. Significant downregulation was recorded for six and nine transcripts at S1 vs. S2 and S2 vs. S3 respectively. This gene family may play a critical role as signalling regulators during the development of Rafflesia floral bud.

Nano-Nutrition of Chicken Embryos. The Effect of in Ovo Administration of Diamond Nanoparticles and l-Glutamine on Molecular Responses in Chicken Embryo Pectoral Muscles

Directory of Open Access Journals (Sweden)

Marta Grodzik

2013-11-01

Full Text Available It has been demonstrated that the content of certain amino acids in eggs is not sufficient to fully support embryonic development. One possibility to supply the embryo with extra nutrients and energy is in ovo administration of nutrients. Nanoparticles of diamond are highly biocompatible non-toxic carbonic structures, and we hypothesized that bio-complexes of diamond nanoparticles with l-glutamine may affect molecular responses in breast muscle. The objective of the investigation was to evaluate the effect of diamond nanoparticle (ND and l-glutamine (Gln on expression of growth and differentiation factors of chicken embryo pectoral muscles. ND, Gln, and Gln/ND solutions (50 mg/L were injected into fertilized broiler chicken eggs at the beginning of embryogenesis. Muscle tissue was dissected at day 20 of incubation and analysed for gene expression of FGF2, VEGF-A, and MyoD1. ND and especially Gln/ND up-regulated expression of genes related to muscle cell proliferation (FGF2 and differentiation (MyoD1. Furthermore, the ratio between FGF2 and MyoD1 was highest in the Gln/ND group. At the end of embryogenesis, Gln/ND enhanced both proliferation and differentiation of pectoral muscle cells and differentiation dominated over proliferation. These preliminary results suggest that the bio-complex of glutamine and diamond nanoparticles may accelerate growth and maturation of muscle cells.

ArgR of Streptomyces coelicolor Is a Pleiotropic Transcriptional Regulator: Effect on the Transcriptome, Antibiotic Production, and Differentiation in Liquid Cultures

Science.gov (United States)

Botas, Alma; Pérez-Redondo, Rosario; Rodríguez-García, Antonio; Álvarez-Álvarez, Rubén; Yagüe, Paula; Manteca, Angel; Liras, Paloma

2018-01-01

ArgR is a well-characterized transcriptional repressor controlling the expression of arginine and pyrimidine biosynthetic genes in bacteria. In this work, the biological role of Streptomyces coelicolor ArgR was analyzed by comparing the transcriptomes of S. coelicolor ΔargR and its parental strain, S. coelicolor M145, at five different times over a 66-h period. The effect of S. coelicolor ArgR was more widespread than that of the orthologous protein of Escherichia coli, affecting the expression of 1544 genes along the microarray time series. This S. coelicolor regulator repressed the expression of arginine and pyrimidine biosynthetic genes, but it also modulated the expression of genes not previously described to be regulated by ArgR: genes involved in nitrogen metabolism and nitrate utilization; the act, red, and cpk genes for antibiotic production; genes for the synthesis of the osmotic stress protector ectoine; genes related to hydrophobic cover formation and sporulation (chaplins, rodlins, ramR, and whi genes); all the cwg genes encoding proteins for glycan cell wall biosynthesis; and genes involved in gas vesicle formation. Many of these genes contain ARG boxes for ArgR binding. ArgR binding to seven new ARG boxes, located upstream or near the ectA-ectB, afsS, afsR, glnR, and redH genes, was tested by DNA band-shift assays. These data and those of previously assayed fragments permitted the construction of an improved model of the ArgR binding site. Interestingly, the overexpression of sporulation genes observed in the ΔargR mutant in our culture conditions correlated with a sporulation-like process, an uncommon phenotype. PMID:29545785

Morphological regulation of Aspergillus niger to improve citric acid production by chsC gene silencing.

Science.gov (United States)

Sun, Xiaowen; Wu, Hefang; Zhao, Genhai; Li, Zhemin; Wu, Xihua; Liu, Hui; Zheng, Zhiming

2018-04-02

The mycelial morphology of Aspergillus niger, a major filamentous fungus used for citric acid production, is important for citric acid synthesis during submerged fermentation. To investigate the involvement of the chitin synthase gene, chsC, in morphogenesis and citric acid production in A. niger, an RNAi system was constructed to silence chsC and the morphological mutants were screened after transformation. The compactness of the mycelial pellets was obviously reduced in the morphological mutants, with lower proportion of dispersed mycelia. These morphological changes have caused a decrease in viscosity and subsequent improvement in oxygen and mass transfer efficiency, which may be conducive for citric acid accumulation. All the transformants exhibited improvements in citric acid production; in particular, chsC-3 showed 42.6% higher production than the original strain in the shake flask. Moreover, the high-yield strain chsC-3 exhibited excellent citric acid production potential in the scale-up process.The citric acid yield and the conversion rate of glucose of chsC-3 were both improved by 3.6%, when compared with that of the original strain in the stirred tank bioreactor.

Vasopressin Gene-Related Products in the Management of Breast Cancer

National Research Council Canada - National Science Library

North, William

1999-01-01

...), and this information coupled with an absence of vasopressin gene-related products from fibrocystic disease potentially provides us with a new screening test for distinguishing both breast cancer...

Impact of alg3 gene deletion on growth, development, pigment production, protein secretion, and functions of recombinant Trichoderma reesei cellobiohydrolases in Aspergillus niger.

Science.gov (United States)

Dai, Ziyu; Aryal, Uma K; Shukla, Anil; Qian, Wei-Jun; Smith, Richard D; Magnuson, Jon K; Adney, William S; Beckham, Gregg T; Brunecky, Roman; Himmel, Michael E; Decker, Stephen R; Ju, Xiaohui; Zhang, Xiao; Baker, Scott E

2013-12-01

Dolichyl-P-Man:Man(5)GlcNAc(2)-PP-dolichyl α-1,3-mannosyltransferase (also known as "asparagine-linked glycosylation 3", or ALG3) is involved in early N-linked glycan synthesis and thus is essential for formation of N-linked protein glycosylation. In this study, we examined the effects of alg3 gene deletion (alg3Δ) on growth, development, pigment production, protein secretion and recombinant Trichoderma reesei cellobiohydrolase (rCel7A) expressed in Aspergillus niger. The alg3Δ delayed spore germination in liquid cultures of complete medium (CM), potato dextrose (PD), minimal medium (MM) and CM with addition of cAMP (CM+cAMP), and resulted in significant reduction of hyphal growth on CM, potato dextrose agar (PDA), and CM+cAMP and spore production on CM. The alg3Δ also led to a significant accumulation of red pigment on both liquid and solid CM cultures. The relative abundances of 54 of the total 215 proteins identified in the secretome were significantly altered as a result of alg3Δ, 63% of which were secreted at higher levels in alg3Δ strain than the parent. The rCel7A expressed in the alg3Δ mutant was smaller in size than that expressed in both wild-type and parental strains, but still larger than T. reesei Cel7A. The circular dichroism (CD)-melt scans indicated that change in glycosylation of rCel7A does not appear to impact the secondary structure or folding. Enzyme assays of Cel7A and rCel7A on nanocrystalline cellulose and bleached kraft pulp demonstrated that the rCel7As have improved activities on hydrolyzing the nanocrystalline cellulose. Overall, the results suggest that alg3 is critical for growth, sporulation, pigment production, and protein secretion in A. niger, and demonstrate the feasibility of this alternative approach to evaluate the roles of N-linked glycosylation in glycoprotein secretion and function. Copyright © 2013 Elsevier Inc. All rights reserved.

Establishment of human sperm-specific voltage-dependent anion channel 3 recombinant vector for the production of a male contraceptive vaccine

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Asmarinah Asmarinah

2012-05-01

Full Text Available Background: The aim of this study was to construct a recombinant vector of human sperm specific VDAC3 gene for production of VDAC3 antibody, which is potential as male contraception vaccine.Methods: Target fragment sequence of VDAC3 gene was obtained through amplification of human sperm VDAC3 cDNA with primers covering exon 5 to exon 8. Its PCR product in size of 435 bp was cloned to the pET101/D-TOPO expression vector (5753 bp. E. coli bacteria were transformed with this vector. Cloning of VDAC3 fragment gene to the vector was confirmed by the using of XbaI restriction enzyme and PCR colony method with primers covering exons 5-8 of the human VDAC3 gene.Results: Alignment analysis of amplified fragment covering exon 5 to exon 8 of VDAC3 gene showed 94% homology to human VDAC3 gene from databank. After cloning to the expression vector and transformation to E. coli competent cells, twelve colonies could grow in culture media. Gel electrophoresis of sliced VDAC3 recombinant vector showed a single band in the size of 6181 bp in 8 colonies. After application of PCR colony and amplicon sequencing, the result showed a single band in the size of 435 bp and fragment sequence with 94% identity to human VDAC3 gene.Conclusion: The construction of human sperm specific VDAC3 gene recombinant vector was established in this study. In the future, this recombinant vector will be used to produce VDAC3 antibody for the development of a male contraception vaccine. (Med J Indones. 2012;21:61-5Keywords: Contraception, recombinant vector, sperm, VDAC3

Effect of 3,3',5-triiodothyronine and 3,5-diiodothyronine on progesterone production, cAMP synthesis, and mRNA expression of STAR, CYP11A1, and HSD3B genes in granulosa layer of chicken preovulatory follicles.

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Sechman, A; Pawlowska, K; Hrabia, A

2011-10-01

In vitro studies were performed to assess whether stimulatory effects of triiodothyronine (T3) on progesterone (P4) production in a granulosa layer (GL) of chicken preovulatory follicles are associated with 3',5'-cyclic adenosine monophosphate (cAMP) synthesis and mRNA expression of STAR protein, CYP11A1, and HSD3B. Effects of 3,5-diiodothyronine (3,5-T2) on steroidogenic function in these follicles were also investigated. The GL of F3 to F1 follicles was incubated in medium supplemented with T3 or 3,5-T2, LH, or forskolin (F), and a combination of each iodothyronine with LH or F. Levels of P4 and cAMP in culture media were determined by RIA. Expression of genes involved in P4 synthesis (ie, STAR protein, CYP11A1, and HSD3B) in the GL of F3 to F1 follicles incubated in medium with T3 or 3,5-T2 and their combination with LH was performed by real-time PCR. Triiodothyronine increased basal and LH- and F-stimulated P4 secretion by preovulatory follicles. The 3,5-T2 elevated P4 synthesis by F3, had no effect on F2 follicles, and diminished P4 production by the GL of F1 follicles. It had no effect on LH-stimulated P4 production; however, it augmented F-stimulated P4 production by F2 and F1 follicles. Although T3 did not affect basal and F-stimulated cAMP synthesis by the GL of preovulatory follicles, it increased LH-stimulated synthesis of this nucleotide. However, 3,5-T2 elevated F-stimulated cAMP synthesis in F3 and F2 follicles; it did not change basal and LH-stimulated cAMP production. Triiodothyronine decreased basal STAR and CYP11A1 mRNAs in F3 follicles, increased them in F1 follicles, and elevated HSD3B mRNA levels in F1 follicles. Triiodothyronine augmented LH-stimulated STAR, CYP11A1, and HSD3B mRNA levels in F2 and CYP11A1 in F1 follicles. However, T3 decreased LH-stimulated STAR and HSD3B mRNA levels in F1 follicles. The 3,5-T2 did not affect basal STAR and CYP11A1 mRNA expression in all investigated follicles; however, it decreased LH-stimulated STAR

Identification of Candidate Gene Variants in Korean MODY Families by Whole-Exome Sequencing.

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Shim, Ye Jee; Kim, Jung Eun; Hwang, Su-Kyeong; Choi, Bong Seok; Choi, Byung Ho; Cho, Eun-Mi; Jang, Kyoung Mi; Ko, Cheol Woo

2015-01-01

To date, 13 genes causing maturity-onset diabetes of the young (MODY) have been identified. However, there is a big discrepancy in the genetic locus between Asian and Caucasian patients with MODY. Thus, we conducted whole-exome sequencing in Korean MODY families to identify causative gene variants. Six MODY probands and their family members were included. Variants in the dbSNP135 and TIARA databases for Koreans and the variants with minor allele frequencies >0.5% of the 1000 Genomes database were excluded. We selected only the functional variants (gain of stop codon, frameshifts and nonsynonymous single-nucleotide variants) and conducted a case-control comparison in the family members. The selected variants were scanned for the previously introduced gene set implicated in glucose metabolism. Three variants c.620C>T:p.Thr207Ile in PTPRD, c.559C>G:p.Gln187Glu in SYT9, and c.1526T>G:p.Val509Gly in WFS1 were respectively identified in 3 families. We could not find any disease-causative alleles of known MODY 1-13 genes. Based on the predictive program, Thr207Ile in PTPRD was considered pathogenic. Whole-exome sequencing is a valuable method for the genetic diagnosis of MODY. Further evaluation is necessary about the role of PTPRD, SYT9 and WFS1 in normal insulin release from pancreatic beta cells. © 2015 S. Karger AG, Basel.

Data of evolutionary structure change: 2A3TA-2PPDC [Confc[Archive

Lifescience Database Archive (English)

Full Text Available 2A3TA-2PPDC 2A3T 2PPD A C --------LPRVKHTLVPPPFAHAHEQVAASGPVINEFE...EDTVKVMRTLTPTHVVFNGAVGALTGDKAMTAAVGEKVLIVHSQANRDTRPHLIGGHGDYVWATGKFNTPPDVDQETWFIP... GLN CA 428 2PPD C 2PPD...> 1 2PPD... C 2PPDC CAPPG-MVPWA

Expression of a mutated SPT15 gene in Saccharomyces cerevisiae enhances both cell growth and ethanol production in microaerobic batch, fed-batch, and simultaneous saccharification and fermentations.

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Seong, Yeong-Je; Park, Haeseong; Yang, Jungwoo; Kim, Soo-Jung; Choi, Wonja; Kim, Kyoung Heon; Park, Yong-Cheol

2017-05-01

The SPT15 gene encodes a Saccharomyces cerevisiae TATA-binding protein, which is able to globally control the transcription levels of various metabolic and regulatory genes. In this study, a SPT15 gene mutant (S42N, S78R, S163P, and I212N) was expressed in S. cerevisiae BY4741 (BSPT15-M3), of which effects on fermentative yeast properties were evaluated in a series of culture types. By applying different nitrogen sources and air supply conditions in batch culture, organic nitrogen sources and microaerobic condition were decided to be more favorable for both cell growth and ethanol production of the BSPT15-M3 strain than the control S. cerevisiae BY4741 strain expressing the SPT15 gene (BSPT15wt). Microaerobic fed-batch cultures of BSPT15-M3 with glucose shock in the presence of high ethanol content resulted in a 9.5-13.4% higher glucose consumption rate and ethanol productivity than those for the BSPT15wt strain. In addition, BSPT15-M3 showed 4.5 and 3.9% increases in ethanol productivity from cassava hydrolysates and corn starch in simultaneous saccharification and fermentation processes, respectively. It was concluded that overexpression of the mutated SPT15 gene would be a potent strategy to develop robust S. cerevisiae strains with enhanced cell growth and ethanol production abilities.

Production of the 2400 kb Duchenne muscular dystrophy (DMD) gene transcript; transcription time and cotranscriptional splicing

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Tennyson, C.N.; Worton, R.G. [Univ. of Toronto and the Hospital for Sick Children, Ontario (Canada)

1994-09-01

The largest known gene in any organism is the human DMD gene which has 79 exons that span 2400 kb. The extreme nature of the DMD gene raises questions concerning the time required for transcription and whether splicing begins before transcription is complete. DMD gene transcription is induced as cultured human myoblasts differentiate to form multinucleated myotubes, providing a system for studying the kinetics of transcription and splicing. Using quantitative RT-PCR, transcript accumulation was monitored from four different regions within the gene following induction of expression. By comparing the accumulation of transcripts from the 5{prime} and 3{prime} ends of the gene we have shown that approximately 12 hours are required to transcribe 1770 kb of the gene, extrapolating to a time of 16 hours for the transcription unit expressed in muscle. Comparison of accumulation profiles for spliced and total transcript demonstrated that transcripts are spliced at the 5{prime} end before transcription is complete, providing strong evidence for cotranscriptional splicing of DMD gene transcripts. Finally, the rate of transcript accumulation was reduced at the 3{prime} end of the gene relative to the 5{prime} end, perhaps due to premature termination of transcription complexes as they traverse this enormous transcription unit. The lag between transcription initiation and the appearance of complete transcripts could be important in limiting transcript production in dividing cells and to the timing of mRNA appearance in differentiating muscle.

A Two-Layer Gene Circuit for Decoupling Cell Growth from Metabolite Production.

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Lo, Tat-Ming; Chng, Si Hui; Teo, Wei Suong; Cho, Han-Saem; Chang, Matthew Wook

2016-08-01

We present a synthetic gene circuit for decoupling cell growth from metabolite production through autonomous regulation of enzymatic pathways by integrated modules that sense nutrient and substrate. The two-layer circuit allows Escherichia coli to selectively utilize target substrates in a mixed pool; channel metabolic resources to growth by delaying enzymatic conversion until nutrient depletion; and activate, terminate, and re-activate conversion upon substrate availability. We developed two versions of controller, both of which have glucose nutrient sensors but differ in their substrate-sensing modules. One controller is specific for hydroxycinnamic acid and the other for oleic acid. Our hydroxycinnamic acid controller lowered metabolic stress 2-fold and increased the growth rate 2-fold and productivity 5-fold, whereas our oleic acid controller lowered metabolic stress 2-fold and increased the growth rate 1.3-fold and productivity 2.4-fold. These results demonstrate the potential for engineering strategies that decouple growth and production to make bio-based production more economical and sustainable. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Detection of cfxA2, cfxA3, and cfxA6 genes in beta-lactamase producing oral anaerobes

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Buhle BINTA

2016-04-01

Full Text Available ABSTRACT Purpose The aim of this study was to identify β-lactamase-producing oral anaerobic bacteria and screen them for the presence of cfxA and BlaTEM genes that are responsible for β-lactamase production and resistance to β-lactam antibiotics. Material and Methods Periodontal pocket debris samples were collected from 48 patients with chronic periodontitis and anaerobically cultured on blood agar plates with and without β-lactam antibiotics. Presumptive β-lactamase-producing isolates were evaluated for definite β-lactamase production using the nitrocefin slide method and identified using the API Rapid 32A system. Antimicrobial susceptibility was performed using disc diffusion and microbroth dilution tests as described by CLSI Methods. Isolates were screened for the presence of the β-lactamase-TEM (BlaTEM and β-lactamase-cfxA genes using Polymerase Chain Reaction (PCR. Amplified PCR products were sequenced and the cfxA gene was characterized using Genbank databases. Results Seventy five percent of patients carried two species of β-lactamase-producing anaerobic bacteria that comprised 9.4% of the total number of cultivable bacteria. Fifty one percent of β-lactamase-producing strains mainly Prevotella, Porphyromonas, and Bacteroides carried the cfxA gene, whereas none of them carried blaTEM. Further characterization of the cfxA gene showed that 76.7% of these strains carried the cfxA2 gene, 14% carried cfxA3, and 9.3% carried cfxA6. The cfxA6 gene was present in three Prevotella spp. and in one Porphyromonas spp. Strains containing cfxA genes (56% were resistant to the β-lactam antibiotics. Conclusion This study indicates that there is a high prevalence of the cfxA gene in β-lactamase-producing anaerobic oral bacteria, which may lead to drug resistance and treatment failure.

1,3-Propanediol dehydrogenases in Lactobacillus reuteri: impact on central metabolism and 3-hydroxypropionaldehyde production

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Meile Leo

2011-08-01

Full Text Available Abstract Background Lactobacillus reuteri metabolizes glycerol to 3-hydroxypropionaldehyde (3-HPA and further to 1,3-propanediol (1,3-PDO, the latter step catalysed by a propanediol dehydrogenase (PDH. The last step in this pathway regenerates NAD+ and enables therefore the energetically more favourable production of acetate over ethanol during growth on glucose. Results A search throughout the genome of L. reuteri DSM 20016 revealed two putative PDHs encoded by ORFs lr_0030 and lr_1734. ORF lr_1734 is situated in the pdu operon encoding the glycerol conversion machinery and therefore likely involved in 1,3-PDO formation. ORF lr_0030 has not been associated with PDH-activity so far. To elucidate the role of these two PDHs, gene deletion mutant strains were constructed. Growth behaviour on glucose was comparable between the wild type and both mutant strains. However, on glucose + glycerol, the exponential growth rate of Δlr_0030 was lower compared to the wild type and the lr_1734 mutant. Furthermore, glycerol addition resulted in decreased ethanol production in the wild type and Δlr_1734, but not in Δlr_0030. PDH activity measurements using 3-HPA as a substrate revealed lower activity of Δlr_0030 extracts from exponential growing cells compared to wild type and Δlr_1734 extracts. During biotechnological 3-HPA production using non-growing cells, the ratio 3-HPA to 1,3-PDO was approximately 7 in the wild type and Δlr_0030, whereas this ratio was 12.5 in the mutant Δlr_1734. Conclusion The enzyme encoded by lr_0030 plays a pivotal role in 3-HPA conversion in exponential growing L. reuteri cells. The enzyme encoded by lr_1734 is active during 3-HPA production by non-growing cells and this enzyme is a useful target to enhance 3-HPA production and minimize formation of the by-product 1,3-PDO.

Combination of traditional mutation and metabolic engineering to enhance ansamitocin P-3 production in Actinosynnema pretiosum.

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Du, Zhi-Qiang; Zhang, Yuan; Qian, Zhi-Gang; Xiao, Han; Zhong, Jian-Jiang

2017-12-01

Ansamitocin P-3 (AP-3) is a maytansinoid with its most compelling antitumor activity, however, the low production titer of AP-3 greatly restricts its wide commercial application. In this work, a combinatorial approach including random mutation and metabolic engineering was conducted to enhance AP-3 biosynthesis in Actinosynnema pretiosum. First, a mutant strain M was isolated by N-methyl-N'-nitro-N-nitrosoguanidine mutation, which could produce AP-3 almost threefold that of wild type (WT) in 48 deep-well plates. Then, by overexpressing key biosynthetic genes asmUdpg and asm13-17 in the M strain, a further 60% increase of AP-3 production in 250-ml shake flasks was achieved in the engineered strain M-asmUdpg:asm13-17 compared to the M strain, and its maximum AP-3 production reached 582.7 mg/L, which is the highest as ever reported. Both the gene transcription levels and intracellular intermediate concentrations in AP-3 biosynthesis pathway were significantly increased in the M and M-asmUdpg:asm13-17 during fermentation compared to the WT. The good fermentation performance of the engineered strain was also confirmed in a lab-scale bioreactor. This work demonstrated that combination of random mutation and metabolic engineering could promote AP-3 biosynthesis and might be helpful for increasing the production of other industrially important secondary metabolites. © 2017 Wiley Periodicals, Inc.

Transcriptome and Gene Ontology (GO) Enrichment Analysis Reveals Genes Involved in Biotin Metabolism That Affect L-Lysine Production in Corynebacterium glutamicum.

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Kim, Hong-Il; Kim, Jong-Hyeon; Park, Young-Jin

2016-03-09

Corynebacterium glutamicum is widely used for amino acid production. In the present study, 543 genes showed a significant change in their mRNA expression levels in L-lysine-producing C. glutamicum ATCC21300 than that in the wild-type C. glutamicum ATCC13032. Among these 543 differentially expressed genes (DEGs), 28 genes were up- or downregulated. In addition, 454 DEGs were functionally enriched and categorized based on BLAST sequence homologies and gene ontology (GO) annotations using the Blast2GO software. Interestingly, NCgl0071 (bioB, encoding biotin synthase) was expressed at levels ~20-fold higher in the L-lysine-producing ATCC21300 strain than that in the wild-type ATCC13032 strain. Five other genes involved in biotin metabolism or transport--NCgl2515 (bioA, encoding adenosylmethionine-8-amino-7-oxononanoate aminotransferase), NCgl2516 (bioD, encoding dithiobiotin synthetase), NCgl1883, NCgl1884, and NCgl1885--were also expressed at significantly higher levels in the L-lysine-producing ATCC21300 strain than that in the wild-type ATCC13032 strain, which we determined using both next-generation RNA sequencing and quantitative real-time PCR analysis. When we disrupted the bioB gene in C. glutamicum ATCC21300, L-lysine production decreased by approximately 76%, and the three genes involved in biotin transport (NCgl1883, NCgl1884, and NCgl1885) were significantly downregulated. These results will be helpful to improve our understanding of C. glutamicum for industrial amino acid production.

Alterations in expression of senescence marker protein-30 gene by 3,3',5-triiodo-L-thyronine (T3).

Science.gov (United States)

Sar, Pranati; Rath, Bandita; Subudhi, Umakanta; Chainy, Gagan Bihari Nityananda; Supakar, Prakash Chandra

2007-09-01

Thyroid hormone (T3) is essential for normal development, differentiation, and metabolic balance of the body. A toxic dose of T(3) in animals increases the basal metabolic rate and reactive oxygen species production, resulting more oxidative stress through Ca(2+) influx to cytoplasm. Senescence Marker Protein-30 (SMP30) is preferentially expressed in the liver and protects cells against various injuries by enhancement of Ca(2+) efflux to either extra cellular space or intraorganellar spaces through membrane Ca(2+) pump activity. In this paper we report an alteration in the level of SMP30 gene expression using RT-PCR and western blot analysis in T(3) treated female Wistar rats. The results indicate that there is an induction of SMP30 expression during early hours of T(3 )treatment and it declines in severe hyperthyroidism. Therefore, we speculate that SMP30 is regulated by T(3) and might play a protective role in hyperthyroidism.

CFH Y402H polymorphism and the complement activation product C5a: effects on NF-κB activation and inflammasome gene regulation.

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Cao, Sijia; Wang, Jay Ching Chieh; Gao, Jiangyuan; Wong, Matthew; To, Elliott; White, Valerie A; Cui, Jing Z; Matsubara, Joanne A

2016-05-01

The Y402H polymorphism in the complement factor H (CFH) gene is an important risk factor for age-related macular degeneration (AMD). Complement activation products and proinflammatory cytokines are associated with this polymorphism at the systemic level, but less is known of the associations in the outer retina of the genotyped eye. Here we investigate complement activation products and their role in nuclear factor (NF)-κB activation and gene expression of the NLRP3 inflammasome pathway. Postmortem donor eyes were genotyped for the CFH Y402H polymorphism and assessed for complement C3a, C5a, interleukin (IL)-18 and tumour necrosis factor (TNF)-α. ARPE19 cells were stimulated basolaterally with C5a or TNF-α in polarised cultures. NF-κB activation was assessed with a reporter cell line. Gene expression of inflammasome-related (NLRP3, caspase-1, IL-1β and IL-18) and classic inflammatory (IL-6 and IL-8) genes was studied. The distribution of inflammasome products, IL-1β and IL-18, was studied in postmortem donor eyes with AMD pathologies. Eyes with the homozygous at-risk variant demonstrated higher levels of C5a, IL-18 and TNF-α in Bruch's membrane and choroid. C5a promoted NF-κB activation and upregulation of IL-18 in polarised ARPE19. TNF-α promoted NF-κB activation and gene expression of caspase-1, IL-1β, IL-18, IL-6 and IL-8, but downregulated NLRP3. In eyes with geographic atrophy, strong immunoreactivity was observed for inflammasome products IL-1β and IL-18 compared with age-matched controls. The at-risk polymorphism of the CFH Y402H may contribute to AMD disease process through increased complement and NF-κB activation, and the upregulation of IL-18, a product of inflammasome activation. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

The Aspergillus flavus Homeobox Gene, hbx1, Is Required for Development and Aflatoxin Production

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Jeffrey W. Cary

2017-10-01

Full Text Available Homeobox proteins, a class of well conserved transcription factors, regulate the expression of targeted genes, especially those involved in development. In filamentous fungi, homeobox genes are required for normal conidiogenesis and fruiting body formation. In the present study, we identified eight homeobox (hbx genes in the aflatoxin-producing ascomycete, Aspergillus flavus, and determined their respective role in growth, conidiation and sclerotial production. Disruption of seven of the eight genes had little to no effect on fungal growth and development. However, disruption of the homeobox gene AFLA_069100, designated as hbx1, in two morphologically different A. flavus strains, CA14 and AF70, resulted in complete loss of production of conidia and sclerotia as well as aflatoxins B1 and B2, cyclopiazonic acid and aflatrem. Microscopic examination showed that the Δhbx1 mutants did not produce conidiophores. The inability of Δhbx1 mutants to produce conidia was related to downregulation of brlA (bristle and abaA (abacus, regulatory genes for conidiophore development. These mutants also had significant downregulation of the aflatoxin pathway biosynthetic genes aflC, aflD, aflM and the cluster-specific regulatory gene, aflR. Our results demonstrate that hbx1 not only plays a significant role in controlling A. flavus development but is also critical for the production of secondary metabolites, such as aflatoxins.

New insights about antibiotic production by Pseudomonas aeruginosa: a gene expression analysis

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Gionco, Bárbara; Tavares, Eliandro R.; de Oliveira, Admilton G.; Yamada-Ogatta, Sueli F.; do Carmo, Anderson O.; Pereira, Ulisses de Pádua; Chideroli, Roberta T.; Simionato, Ane S.; Navarro, Miguel O. P.; Chryssafidis, Andreas L.; Andrade, Galdino

2017-09-01

The bacterial resistance for antibiotics is one of the most important problems in public health and only a small number of new products are in development. Antagonistic microorganisms from soil are a promising source of new candidate molecules. Products of secondary metabolism confer adaptive advantages for their producer, in the competition for nutrients in the microbial community. The biosynthesis process of compounds with antibiotic activity is the key to optimize their production and the transcriptomic study of microorganisms is of great benefit for the discovery of these metabolic pathways. Pseudomonas aeruginosa LV strain growing in the presence of copper chloride produces a bioactive organometallic compound, which has a potent antimicrobial activity against various microorganisms. The objective of this study was to verify overexpressed genes and evaluate their relation to the organometallic biosynthesis in this microorganism. P. aeruginosa LV strain was cultured in presence and absence of copper chloride. Two methods were used for transcriptomic analysis, genome reference-guided assembly and de novo assembly. The genome referenced analysis identified nine upregulated genes when bacteria were exposed to copper chloride, while the De Novo Assembly identified twelve upregulated genes. Nineteen genes can be related to an increased microbial metabolism for the extrusion process of exceeding intracellular copper. Two important genes are related to the biosynthesis of phenazine and tetrapyrroles compounds, which can be involved in the bioremediation of intracellular copper and biosynthesis of the organometallic compound. Additional studies are being carried out to further prove the function of the described genes and relate them to the biosynthetic pathway of the organometallic compound.

Data of evolutionary structure change: 1BS7A-3CPMA [Confc[Archive

Lifescience Database Archive (English)

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Data of evolutionary structure change: 1BS5A-3CPMA [Confc[Archive

Lifescience Database Archive (English)

Full Text Available 1BS5A-3CPMA 1BS5 3CPM A A -----SVLQVLHIPDERLRKVAKPVEEVNAEIQRIVDDM...line> 3CPM A 3CPMA THR CA 218 GLY CA 252 ILE CA 234 3CPM... A 3CPMA IKKYSDKLVPFILE CA 320 ARG CA 317 GLN CA 243 3CPM

Whole-Genome Microarray and Gene Deletion Studies Reveal Regulation of the Polyhydroxyalkanoate Production Cycle by the Stringent Response in Ralstonia eutropha H16

Energy Technology Data Exchange (ETDEWEB)

Brigham, CJ; Speth, DR; Rha, C; Sinskey, AJ

2012-10-22

Poly(3-hydroxybutyrate) (PHB) production and mobilization in Ralstonia eutropha are well studied, but in only a few instances has PHB production been explored in relation to other cellular processes. We examined the global gene expression of wild-type R. eutropha throughout the PHB cycle: growth on fructose, PHB production using fructose following ammonium depletion, and PHB utilization in the absence of exogenous carbon after ammonium was resupplied. Our results confirm or lend support to previously reported results regarding the expression of PHB-related genes and enzymes. Additionally, genes for many different cellular processes, such as DNA replication, cell division, and translation, are selectively repressed during PHB production. In contrast, the expression levels of genes under the control of the alternative sigma factor sigma(54) increase sharply during PHB production and are repressed again during PHB utilization. Global gene regulation during PHB production is strongly reminiscent of the gene expression pattern observed during the stringent response in other species. Furthermore, a ppGpp synthase deletion mutant did not show an accumulation of PHB, and the chemical induction of the stringent response with DL-norvaline caused an increased accumulation of PHB in the presence of ammonium. These results indicate that the stringent response is required for PHB accumulation in R. eutropha, helping to elucidate a thus-far-unknown physiological basis for this process.

The R2R3-MYB–Like Regulatory Factor EOBI, Acting Downstream of EOBII, Regulates Scent Production by Activating ODO1 and Structural Scent-Related Genes in Petunia[C][W

Science.gov (United States)

Spitzer-Rimon, Ben; Farhi, Moran; Albo, Boaz; Cna’ani, Alon; Ben Zvi, Michal Moyal; Masci, Tania; Edelbaum, Orit; Yu, Yixun; Shklarman, Elena; Ovadis, Marianna; Vainstein, Alexander

2012-01-01

Flower scent is a highly dynamic trait, under developmental, spatial, and diurnal regulation. The mechanism governing scent production is only beginning to be unraveled. In petunia (Petunia hybrida), EMISSION OF BENZENOIDS II (EOBII) controls transcription of both the shikimate pathway-regulating MYB factor ODORANT1 (ODO1) and phenylpropanoid scent-related structural genes. A promoter-activation screen identified an R2R3-MYB–like regulatory factor of phenylpropanoid volatile biosynthesis acting downstream of EOBII, designated EOBI. EOBI silencing led to downregulation of ODO1 and numerous structural scent-related genes from both the shikimate and phenylpropanoid pathways. The ability of EOBI to directly activate ODO1, as revealed by electrophoretic mobility shift assay and yeast one-hybrid analysis, place EOBI upstream of ODO1 in regulating substrate availability for volatile biosynthesis. Interestingly, ODO1-silenced transgenic petunia flowers accumulated higher EOBI transcript levels than controls, suggesting a complex feedback loop between these regulatory factors. The accumulation pattern of EOBI transcript relative to EOBII and ODO1, and the effect of up/downregulation of EOBII on transcript levels of EOBI and ODO1, further support these factors' hierarchical relationships. The dependence of scent production on EOBI expression and its direct interaction with both regulatory and structural genes provide evidence for EOBI’s wide-ranging involvement in the production of floral volatiles. PMID:23275577

The YJR127C/ZMS1 gene product is involved in glycerol-based respiratory growth of the yeast Saccharomyces cerevisiae.

Science.gov (United States)

Lu, Lin; Roberts, George G; Oszust, Cynthia; Hudson, Alan P

2005-10-01

A putative yeast mitochondrial upstream activating sequence (UAS) was used in a one-hybrid screening procedure that identified the YJR127C ORF on chromosome X. This gene was previously designated ZMS1 and is listed as a transcription factor on the SGD website. Real time RT-PCR assays showed that expression of YJR127C/ZMS1 was glucose-repressible, and a deletion mutant for the gene showed a growth defect on glycerol-based but not on glucose- or ethanol-based medium. Real time RT-PCR analyses identified severely attenuated transcript levels from GUT1 and GUT2 to be the source of that growth defect, the products of GUT1 and GUT2 are required for glycerol utilization. mRNA levels from a large group of mitochondria- and respiration-related nuclear genes also were shown to be attenuated in the deletion mutant. Importantly, transcript levels from the mitochondrial OLI1 gene, which has an associated organellar UAS, were attenuated in the DeltaYJR127C mutant during glycerol-based growth, but those from COX3 (OXI2), which lacks an associated mitochondrial UAS, were not. Transcriptome analysis of the glycerol-grown deletion mutant showed that genes in several metabolic and other categories are affected by loss of this gene product, including protein transport, signal transduction, and others. Thus, the product of YJR127C/ZMS1 is involved in transcriptional control for genes in both cellular genetic compartments, many of which specify products required for glycerol-based growth, respiration, and other functions.

A second pectin lyase gene (pel2) from Aspergillus oryzae KBN616: its sequence analysis and overexpression, and characterization of the gene products.

Science.gov (United States)

Kitamoto, N; Yoshino-Yasuda, S; Ohmiya, K; Tsukagoshi, N

2001-01-01

A second pectin lyase gene, designated pel2, was isolated from a shoyu koji mold Aspergillus oryzae KBN616 and characterized. The structural gene comprised 1306 bp with three introns. The ORF encoded 375 amino acids with a signal peptide of 19 amino acids. The deduced amino acid sequence showed high similarity to those of A. oryzae Pel1, Aspergillus niger pectin lyases and Glomerella cingulata Pn1A. The pel2 gene was overexpressed under the control of the promoter of the A. oryzae TEF1 gene for purification and enzymatic characterization of its gene product. The gene product exhibited two molecular masses of 48 and 44 kDa due to different degrees of glycosylation. Both proteins had the same pH optimum of 6.0 and temperature optimum of 50 degrees C.

Data of evolutionary structure change: 2A3TA-2PPDA [Confc[Archive

Lifescience Database Archive (English)

Full Text Available 2A3TA-2PPDA 2A3T 2PPD A A --------LPRVKHTLVPPPFAHAHEQVAASGPVINEFE...EDTVKVMRTLTPTHVVFNGAVGALTGDKAMTAAVGEKVLIVHSQANRDTRPHLIGGHGDYVWATGKFNTPPDVDQETWFIP... GLN CA 428 2PPD A 2PPD...indel> 1 2PPD A 2PPDA CAPPG-MVPWA

Data of evolutionary structure change: 2A3TA-2PPDB [Confc[Archive

Lifescience Database Archive (English)

Full Text Available 2A3TA-2PPDB 2A3T 2PPD A B --------LPRVKHTLVPPPFAHAHEQVAASGPVINEFE...YEDTVKVMRTLTPTHVVFNGAVGALTGDKAMTAAVGEKVLIVHSQANRDTRPHLIGGHGDYVWATGKFNTPPDVDQETWFI...ne> GLN CA 428 2PPD B 2PPD...ax> 1 2PPD... B 2PPDB CAPPG-MVPWA

Further enhanced production of heterologous proteins by double-gene disruption (ΔAosedD ΔAovps10) in a hyper-producing mutant of Aspergillus oryzae.

Science.gov (United States)

Zhu, Lin; Maruyama, Jun-ichi; Kitamoto, Katsuhiko

2013-07-01

The filamentous fungus Aspergillus oryzae is used as one of the most favored hosts for heterologous protein production due to its ability to secrete large amounts of proteins into the culture medium. We previously generated a hyper-producing mutant strain of A. oryzae, AUT1, which produced 3.2- and 2.6-fold higher levels of bovine chymosin (CHY) and human lysozyme (HLY), respectively, compared with the wild-type strain. However, further enhancement of heterologous protein production by multiple gene disruption is difficult because of the low gene-targeting efficiency in strain AUT1. Here, we disrupted the ligD gene, which is involved in nonhomologous recombination, and the pyrG gene to create uridine/uracil auxotrophy in strain AUT1, to generate a hyper-producing mutant applicable to pyrG marker recycling with highly efficient gene targeting. We generated single and double disruptants of the tripeptidyl peptidase gene AosedD and vacuolar sorting receptor gene Aovps10 in the hyper-producing mutant background, and found that all disruptants showed significant increases in heterologous protein production. Particularly, double disruption of the Aovps10 and AosedD genes increased the production levels of CHY and HLY by 1.6- and 2.1-fold, respectively, compared with the parental strain. Thus, we successfully generated a fungal host for further enhancing the heterologous protein production ability by combining mutational and molecular breeding techniques.

Major genes and QTL influencing wool production and quality: a review

Directory of Open Access Journals (Sweden)

Purvis Ian

2005-12-01

Full Text Available Abstract The opportunity exists to utilise our knowledge of major genes that influence the economically important traits in wool sheep. Genes with Mendelian inheritance have been identified for many important traits in wool sheep. Of particular importance are genes influencing pigmentation, wool quality and the keratin proteins, the latter of which are important for the morphology of the wool fibre. Gene mapping studies have identified some chromosomal regions associated with variation in wool quality and production traits. The challenge now is to build on this knowledge base in a cost-effective way to deliver molecular tools that facilitate enhanced genetic improvement programs for wool sheep.

Association studies of OGG1, XRCC1, XRCC2 and XRCC3 polymorphisms with differentiated thyroid cancer

Energy Technology Data Exchange (ETDEWEB)

Garcia-Quispes, Wilser-Andres; Perez-Machado, Giselle; Akdi, Abdelmounaim [Grup de Mutagenesi, Departament de Genetica i de Microbiologia, Universitat Autonoma de Barcelona, Bellaterra (Spain); Pastor, Susana [Grup de Mutagenesi, Departament de Genetica i de Microbiologia, Universitat Autonoma de Barcelona, Bellaterra (Spain); CIBER Epidemiologia y Salud Publica, Instituto de Salud Carlos III (Spain); Galofre, Pere [Servei de Medicina Nuclear, Hospitals Universitaris Vall d' Hebron, Barcelona (Spain); Biarnes, Fina [Unitat d' Endocrinologia, Hospital Josep Trueta, Girona (Spain); Castell, Joan [Servei de Medicina Nuclear, Hospitals Universitaris Vall d' Hebron, Barcelona (Spain); Velazquez, Antonia [Grup de Mutagenesi, Departament de Genetica i de Microbiologia, Universitat Autonoma de Barcelona, Bellaterra (Spain); CIBER Epidemiologia y Salud Publica, Instituto de Salud Carlos III (Spain); Marcos, Ricard, E-mail: ricard.marcos@uab.es [Grup de Mutagenesi, Departament de Genetica i de Microbiologia, Universitat Autonoma de Barcelona, Bellaterra (Spain); CIBER Epidemiologia y Salud Publica, Instituto de Salud Carlos III (Spain)

2011-05-10

The role of the DNA repair genes OGG1, XRCC1, XRCC2 and XRCC3 on differentiated thyroid cancer (DTC) susceptibility was examined in 881 individuals (402 DTC and 479 controls). DNA repair genes were proposed as candidate genes, since the current data indicate that exposure to ionizing radiation is the only established factor in the development of thyroid cancer, especially when it occurs in early stages of life. We have genotyped DNA repair genes involved in base excision repair (BER) (OGG1, Ser326Cys; XRCC1, Arg280His and Arg399Gln), and homologous recombination repair (HRR) (XRCC2, Arg188His and XRCC3, ISV-14G). Genotyping was carried out using the iPLEX (Sequenom) technique. Multivariate logistic regression analyses were performed in a case-control study design. From all the studied polymorphism, only a positive association (OR = 1.58, 95% CI 1.05-2.46, P = 0.027) was obtained for XRCC1 (Arg280His). No associations were observed for the other polymorphisms. No effects of the histopathological type of tumor were found when the DTC patients were stratified according to the type of tumor. It must be emphasized that this study include the greater patients group, among the few studies carried out until now determining the role of DNA repair genes in thyroid cancer susceptibility.

Association studies of OGG1, XRCC1, XRCC2 and XRCC3 polymorphisms with differentiated thyroid cancer

International Nuclear Information System (INIS)

Garcia-Quispes, Wilser-Andres; Perez-Machado, Giselle; Akdi, Abdelmounaim; Pastor, Susana; Galofre, Pere; Biarnes, Fina; Castell, Joan; Velazquez, Antonia; Marcos, Ricard

2011-01-01

The role of the DNA repair genes OGG1, XRCC1, XRCC2 and XRCC3 on differentiated thyroid cancer (DTC) susceptibility was examined in 881 individuals (402 DTC and 479 controls). DNA repair genes were proposed as candidate genes, since the current data indicate that exposure to ionizing radiation is the only established factor in the development of thyroid cancer, especially when it occurs in early stages of life. We have genotyped DNA repair genes involved in base excision repair (BER) (OGG1, Ser326Cys; XRCC1, Arg280His and Arg399Gln), and homologous recombination repair (HRR) (XRCC2, Arg188His and XRCC3, ISV-14G). Genotyping was carried out using the iPLEX (Sequenom) technique. Multivariate logistic regression analyses were performed in a case-control study design. From all the studied polymorphism, only a positive association (OR = 1.58, 95% CI 1.05-2.46, P = 0.027) was obtained for XRCC1 (Arg280His). No associations were observed for the other polymorphisms. No effects of the histopathological type of tumor were found when the DTC patients were stratified according to the type of tumor. It must be emphasized that this study include the greater patients group, among the few studies carried out until now determining the role of DNA repair genes in thyroid cancer susceptibility.

5' Analysis of the soybean leghaemoglobin lbc(3) gene

DEFF Research Database (Denmark)

Stougaard, J; Sandal, N N; Grøn, A

1987-01-01

The soybean leghaemoglobin lbc(3) gene promoter was analysed in transgenic Lotus corniculatus plants. Hybrid-promoter constructions and 5' deletions were studied using chimeric genes composed of the various promoters, the chloramphenicol acetyltransferase (CAT) coding sequence and the lbc(3) 3...

Mutation in the peroxin-coding gene PEX22 contributing to high malate production in Saccharomyces cerevisiae.

Science.gov (United States)

Negoro, Hiroaki; Sakamoto, Mitsuru; Kotaka, Atsushi; Matsumura, Kengo; Hata, Yoji

2018-02-01

Saccharomyces cerevisiae produces organic acids such as succinate, acetate, and malate during alcoholic fermentation. Since malate contributes to the pleasant taste of sake (a Japanese alcoholic beverage), various methods for breeding high-malate-producing yeast strains have been developed. Here, a high-malate-producing yeast strain F-701H was isolated. This mutant was sensitive to dimethyl succinate (DMS) and harbored a nonsense mutation in the peroxin gene PEX22, which was identified as the cause of high malate production by comparative genome analysis. This mutation, which appeared to cause Pex22p dysfunction, was sufficient to confer increased malate productivity and DMS sensitivity to yeast cells. Next, we investigated the mechanism by which this mutation led to high malate production in yeast cells. Peroxins, such as Pex22p, maintain peroxisomal biogenesis. Analysis of 29 PEX disruptants revealed an increased malate production by deletion of the genes encoding peroxins responsible for importing proteins (containing peroxisomal targeting signal 1, PTS1) into the peroxisomal matrix, and those responsible for the assembly of peroxins themselves in the peroxisomal membrane. A defect in peroxisomal malate dehydrogenase (Mdh3p), harboring endogenous PTS1, inhibited the high malate-producing phenotype in the PEX22 mutant. Moreover, Mdh3p, which was normally sorted to the peroxisomal matrix, was potentially mislocalized to the cytosol in the PEX22 mutant. This suggested that an increase in malate production resulted from the mislocalization of Mdh3p from the peroxisome to the cytoplasm due to the loss of peroxin-mediated transportation. Thus, the present study revealed a novel mechanism for organic acid productions in yeast during sake brewing. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Co-Transcriptomes of Initial Interactions In Vitro between Streptococcus Pneumoniae and Human Pleural Mesothelial Cells.

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Claire J Heath

Full Text Available Streptococcus pneumoniae (Spn is a major causative organism of empyema, an inflammatory condition occurring in the pleural sac. In this study, we used human and Spn cDNA microarrays to characterize the transcriptional responses occurring during initial contact between Spn and a human pleural mesothelial cell line (PMC in vitro. Using stringent filtering criteria, 42 and 23 Spn genes were up-and down-regulated respectively. In particular, genes encoding factors potentially involved in metabolic processes and Spn adherence to eukaryotic cells were up-regulated e.g. glnQ, glnA, aliA, psaB, lytB and nox. After Spn initial contact, 870 human genes were differentially regulated and the largest numbers of significant gene expression changes were found in canonical pathways for eukaryotic initiation factor 2 signaling (60 genes out of 171, oxidative phosphorylation (32/103, mitochondrial dysfunction (37/164, eIF4 and p70S6K signaling (28/142, mTOR signaling (27/182, NRF2-mediated oxidative stress response (20/177, epithelial adherens junction remodeling (11/66 and ubiquitination (22/254. The cellular response appeared to be directed towards host cell survival and defense. Spn did not activate NF-kB or phosphorylate p38 MAPK or induce cytokine production from PMC. Moreover, Spn infection of TNF-α pre-stimulated PMC inhibited production of IL-6 and IL-8 secretion by >50% (p<0.01. In summary, this descriptive study provides datasets and a platform for examining further the molecular mechanisms underlying the pathogenesis of empyema.

Studies of the associations between functional beta2-adrenergic receptor variants and obesity, hypertension and type 2 diabetes in 7,808 white subjects

DEFF Research Database (Denmark)

Gjesing, A P; Andersen, G; Burgdorf, K S

2007-01-01

Functional and common Arg16Gly and Gln27Glu polymorphisms have been identified in ADRB2, the gene encoding the beta2-adrenergic receptor. These variants have previously been examined for association with obesity, hypertension and diabetes with inconclusive results.......Functional and common Arg16Gly and Gln27Glu polymorphisms have been identified in ADRB2, the gene encoding the beta2-adrenergic receptor. These variants have previously been examined for association with obesity, hypertension and diabetes with inconclusive results....

The evolution of milk casein genes from tooth genes before the origin of mammals.

Science.gov (United States)

Kawasaki, Kazuhiko; Lafont, Anne-Gaelle; Sire, Jean-Yves

2011-07-01

Caseins are among cardinal proteins that evolved in the lineage leading to mammals. In milk, caseins and calcium phosphate (CaP) form a huge complex called casein micelle. By forming the micelle, milk maintains high CaP concentrations, which help altricial mammalian neonates to grow bone and teeth. Two types of caseins are known. Ca-sensitive caseins (α(s)- and β-caseins) bind Ca but precipitate at high Ca concentrations, whereas Ca-insensitive casein (κ-casein) does not usually interact with Ca but instead stabilizes the micelle. Thus, it is thought that these two types of caseins are both necessary for stable micelle formation. Both types of caseins show high substitution rates, which make it difficult to elucidate the evolution of caseins. Yet, recent studies have revealed that all casein genes belong to the secretory calcium-binding phosphoprotein (SCPP) gene family that arose by gene duplication. In the present study, we investigated exon-intron structures and phylogenetic distributions of casein and other SCPP genes, particularly the odontogenic ameloblast-associated (ODAM) gene, the SCPP-Pro-Gln-rich 1 (SCPPPQ1) gene, and the follicular dendritic cell secreted peptide (FDCSP) gene. The results suggest that contemporary Ca-sensitive casein genes arose from a putative common ancestor, which we refer to as CSN1/2. The six putative exons comprising CSN1/2 are all found in SCPPPQ1, although ODAM also shares four of these exons. By contrast, the five exons of the Ca-insensitive casein gene are all reminiscent of FDCSP. The phylogenetic distribution of these genes suggests that both SCPPPQ1 and FDCSP arose from ODAM. We thus argue that all casein genes evolved from ODAM via two different pathways; Ca-sensitive casein genes likely originated directly from SCPPPQ1, whereas the Ca-insensitive casein genes directly differentiated from FDCSP. Further, expression of ODAM, SCPPPQ1, and FDCSP was detected in dental tissues, supporting the idea that both types of caseins

Regulatory Oversight of Cell and Gene Therapy Products in Canada.

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Ridgway, Anthony; Agbanyo, Francisca; Wang, Jian; Rosu-Myles, Michael

2015-01-01

Health Canada regulates gene therapy products and many cell therapy products as biological drugs under the Canadian Food and Drugs Act and its attendant regulations. Cellular products that meet certain criteria, including minimal manipulation and homologous use, may be subjected to a standards-based approach under the Safety of Human Cells, Tissues and Organs for Transplantation Regulations. The manufacture and clinical testing of cell and gene therapy products (CGTPs) presents many challenges beyond those for protein biologics. Cells cannot be subjected to pathogen removal or inactivation procedures and must frequently be administered shortly after final formulation. Viral vector design and manufacturing control are critically important to overall product quality and linked to safety and efficacy in patients through concerns such as replication competence, vector integration, and vector shedding. In addition, for many CGTPs, the value of nonclinical studies is largely limited to providing proof of concept, and the first meaningful data relating to appropriate dosing, safety parameters, and validity of surrogate or true determinants of efficacy must come from carefully designed clinical trials in patients. Addressing these numerous challenges requires application of various risk mitigation strategies and meeting regulatory expectations specifically adapted to the product types. Regulatory cooperation and harmonisation at an international level are essential for progress in the development and commercialisation of these products. However, particularly in the area of cell therapy, new regulatory paradigms may be needed to harness the benefits of clinical progress in situations where the resources and motivation to pursue a typical drug product approval pathway may be lacking.

Development of Jerusalem artichoke resource for efficient one-step fermentation of poly-(γ-glutamic acid) using a novel strain Bacillus amyloliquefaciens NX-2S.

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Qiu, Yibin; Sha, Yuanyuan; Zhang, Yatao; Xu, Zongqi; Li, Sha; Lei, Peng; Xu, Zheng; Feng, Xiaohai; Xu, Hong

2017-09-01

This study aimed to develop non-food fermentation for the cost-effective production of poly-(γ-glutamic acid) (γ-PGA) using a novel strain of Bacillus amyloliquefaciens NX-2S. The new isolate assimilated inulin more efficiently than other carbohydrates from Jerusalem artichoke, without hydrolytic treatment. To investigate the effect of inulin on γ-PGA production, the transcript levels of γ-PGA synthetase genes (pgsB, pgsC, pgsA), regulatory genes (comA, degQ, degS), and the glutamic acid biosynthesis gene (glnA) were analyzed; inulin addition upregulated these key genes. Without exogenous glutamate, strain NX-2S could produce 6.85±0.22g/L of γ-PGA during fermentation. Exogenous glutamate greatly enhances the γ-PGA yield (39.4±0.38g/L) and productivity (0.43±0.05g/L/h) in batch fermentation. Our study revealed a potential method of non-food fermentation to produce high-value products. Copyright © 2017. Published by Elsevier Ltd.

Clinical significance of productive immunoglobulin heavy chain gene rearrangements in childhood acute lymphoblastic leukemia.

Science.gov (United States)

Katsibardi, Katerina; Braoudaki, Maria; Papathanasiou, Chrissa; Karamolegou, Kalliopi; Tzortzatou-Stathopoulou, Fotini

2011-09-01

We analyzed the CDR3 region of 80 children with B-cell acute lymphoblastic leukemia (B-ALL) using the ImMunoGeneTics Information System and JOINSOLVER. In total, 108 IGH@ rearrangements were analyzed. Most of them (75.3%) were non-productive. IGHV@ segments proximal to IGHD-IGHJ@ were preferentially rearranged (45.3%). Increased utilization of IGHV3 segments IGHV3-13 (11.3%) and IGHV3-15 (9.3%), IGHD3 (30.5%), and IGHJ4 (34%) was noted. In pro-B ALL more frequent were IGHV3-11 (33.3%) and IGHV6-1 (33.3%), IGHD2-21 (50%), IGHJ4 (50%), and IGHJ6 (50%) segments. Shorter CDR3 length was observed in IGHV@6, IGHD7, and IGHJ1 segments, whereas increased CDR3 length was related to IGHV3, IGHD2, and IGHJ4 segments. Increased risk of relapse was found in patients with productive sequences. Specifically, the relapse-free survival rate at 5 years in patients with productive sequences at diagnosis was 75% (standard error [SE] ±9%), whereas in patients with non-productive sequences it was 97% (SE ±1.92%) (p-value =0.0264). Monoclonality and oligoclonality were identified in 81.2% and 18.75% cases at diagnosis, respectively. Sequence analysis revealed IGHV@ to IGHDJ joining only in 6.6% cases with oligoclonality. The majority (75%) of relapsed patients had monoclonal IGH@ rearrangements. The preferential utilization of IGHV@ segments proximal to IGHDJ depended on their location on the IGHV@ locus. Molecular mechanisms occurring during IGH@ rearrangement might play an essential role in childhood ALL prognosis. In our study, the productivity of the rearranged sequences at diagnosis proved to be a significant prognostic factor.

Polymorphisms in Toll-like receptors 2 and 4 genes and their expression in chronic suppurative otitis media.

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Jotic, Ana; Jesic, Snezana; Zivkovic, Maja; Tomanovic, Nada; Kuveljic, Jovana; Stankovic, Aleksandra

2015-12-01

Toll-like receptors (TLRs) have a prominent role in inducing innate immune response. It has been suggested that regulation of TLRs is involved in the pathogenesis of chronic otitis media. TLR 2 and TLR 4 polymorphisms were connected with susceptibility to acute otitis and chronic otitis with effusion. The objective of this study was to establish expression of TLR 2 and 4 on middle ear mucosa in different types of chronic suppurative otitis media (CSOM), and the influence of gene polymorphisms TLR 2 Arg753Gln and TLR 4 Thr399Ile and Asp299Gly to susceptibility to CSOM. Middle ear mucosa and full blood samples were obtained from 85 patients with chronic suppurative otitis media with and without cholesteatoma. Control group for mucosal TLR expression consisted of 71 samples of middle ear mucosa taken from patients with otosclerosis, and control group for DNA polymorphism consisted of 100 full blood samples in healthy subjects. DNA polymorphism detection was done with restriction fragment length polymorphism in RT PCR. Expression of TLR 2 and 4 was determined with immunohistochemical staining. TLR 2 and TLR 4 expression on the middle ear mucosa was not influenced by age of the patients with chronic otitis media. Incidence of TLR 2 Arg753Gln polymorphism was significantly higher in patients with chronic otitis media, compared to control group. Significant association between TLR 2 Arg753Gln polymorphism and different types of mucosal changes in patients with chronic otitis media was established. TLR 2 and 4 expression on experimental group mucosa was significantly different compared to control group, where there was no expression (p=0.000). Strong dependence of TLR 2 and TLR 4 expression on middle ear mucosa with different mucosal changes and immunohistochemical activity after staining was detected. Certain polymorphisms in TLR genes could be indicative for susceptibility to chronic otitis media. Expression of TLR 2 and 4 on middle ear mucosa was more dependable on

Aromatic Polyketide GTRI-02 is a Previously Unidentified Product of the act Gene Cluster in Streptomyces coelicolor A3(2).

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Wu, Changsheng; Ichinose, Koji; Choi, Young Hae; van Wezel, Gilles P

2017-07-18

The biosynthesis of aromatic polyketides derived from type II polyketide synthases (PKSs) is complex, and it is not uncommon that highly similar gene clusters give rise to diverse structural architectures. The act biosynthetic gene cluster (BGC) of the model actinomycete Streptomyces coelicolor A3(2) is an archetypal type II PKS. Here we show that the act BGC also specifies the aromatic polyketide GTRI-02 (1) and propose a mechanism for the biogenesis of its 3,4-dihydronaphthalen-1(2H)-one backbone. Polyketide 1 was also produced by Streptomyces sp. MBT76 after activation of the act-like qin gene cluster by overexpression of the pathway-specific activator. Mining of this strain also identified dehydroxy-GTRI-02 (2), which most likely originated from dehydration of 1 during the isolation process. This work shows that even extensively studied model gene clusters such as act of S. coelicolor can still produce new chemistry, offering new perspectives for drug discovery. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Analysis of the structural genes encoding M-factor in the fission yeast Schizosaccharomyces pombe: identification of a third gene, mfm3

DEFF Research Database (Denmark)

Kjaerulff, S; Davey, William John; Nielsen, O

1994-01-01

We previously identified two genes, mfm1 and mfm2, with the potential to encode the M-factor mating pheromone of the fission yeast Schizosaccharomyces pombe (J. Davey, EMBO J. 11:951-960, 1992), but further analysis revealed that a mutant strain lacking both genes still produced active M-factor. ......We previously identified two genes, mfm1 and mfm2, with the potential to encode the M-factor mating pheromone of the fission yeast Schizosaccharomyces pombe (J. Davey, EMBO J. 11:951-960, 1992), but further analysis revealed that a mutant strain lacking both genes still produced active M...... that is not rescued by addition of exogenous M-factor. A mutational analysis reveals that all three mfm genes contribute to the production of M-factor. Their transcription is limited to M cells and requires the mat1-Mc and ste11 gene products. Each gene is induced when the cells are starved of nitrogen and further...

Exocrine pancreas glutamate secretion help to sustain enterocyte nutritional needs under protein restriction.

Science.gov (United States)

Araya, S; Kuster, E; Gluch, D; Mariotta, L; Lutz, C; Reding, T V; Graf, R; Verrey, F; Camargo, S M R

2018-04-01

Glutamine (Gln) is the most concentrated amino acid in blood and considered conditionally essential. Its requirement is increased during physiological stress, such as malnutrition or illness, despite its production by muscle and other organs. In the malnourished state, Gln has been suggested to have a trophic effect on the exocrine pancreas and small intestine. However, the Gln transport capacity, the functional relationship of these two organs, and the potential role of the Gln-glutamate (Glu) cycle are unknown. We observed that pancreatic acinar cells express lower levels of Glu than Gln transporters. Consistent with this expression pattern, the rate of Glu influx into acinar cells was approximately sixfold lower than that of Gln. During protein restriction, acinar cell glutaminase expression was increased and Gln accumulation was maintained. Moreover, Glu secretion by acinar cells into pancreatic juice and thus into the lumen of the small intestine was maintained. In the intestinal lumen, Glu absorption was preserved and Glu dehydrogenase expression was augmented, potentially providing the substrates for increasing energy production via the TCA cycle. Our findings suggest that one mechanism by which Gln exerts a positive effect on exocrine pancreas and small intestine involves the Gln metabolism in acinar cells and the secretion of Glu into the small intestine lumen. The exocrine pancreas acinar cells not only avidly accumulate Gln but metabolize Gln to generate energy and to synthesize Glu for secretion in the pancreatic juice. Secreted Glu is suggested to play an important role during malnourishment in sustaining small intestinal homeostasis. NEW & NOTEWORTHY Glutamine (Gln) has been suggested to have a trophic effect on exocrine pancreas and small intestine in malnourished states, but the mechanism is unknown. In this study, we suggest that this trophic effect derives from an interorgan relationship between exocrine pancreas and small intestine for Gln

New Insights about Antibiotic Production by Pseudomonas aeruginosa: A Gene Expression Analysis

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Bárbara Gionco

2017-09-01

Full Text Available The bacterial resistance for antibiotics is one of the most important problems in public health and only a small number of new products are in development. Antagonistic microorganisms from soil are a promising source of new candidate molecules. Products of secondary metabolism confer adaptive advantages for their producer, in the competition for nutrients in the microbial community. The biosynthesis process of compounds with antibiotic activity is the key to optimize their production and the transcriptomic study of microorganisms is of great benefit for the discovery of these metabolic pathways. Pseudomonas aeruginosa LV strain growing in the presence of copper chloride produces a bioactive organometallic compound, which has a potent antimicrobial activity against various microorganisms. The objective of this study was to verify overexpressed genes and evaluate their relation to the organometallic biosynthesis in this microorganism. P. aeruginosa LV strain was cultured in presence and absence of copper chloride. Two methods were used for transcriptomic analysis, genome reference-guided assembly and de novo assembly. The genome referenced analysis identified nine upregulated genes when bacteria were exposed to copper chloride, while the De Novo Assembly identified 12 upregulated genes. Nineteen genes can be related to an increased microbial metabolism for the extrusion process of exceeding intracellular copper. Two important genes are related to the biosynthesis of phenazine and tetrapyrroles compounds, which can be involved in the bioremediation of intracellular copper and we suggesting that may involve in the biosynthesis of the organometallic compound. Additional studies are being carried out to further prove the function of the described genes and relate them to the biosynthetic pathway of the organometallic compound.

Characterization of foot-and-mouth disease virus gene products with antisera against bacterially synthesized fusion proteins

International Nuclear Information System (INIS)

Strebel, K.; Beck, E.; Strohmaier, K.; Schaller, H.

1986-01-01

Defined segments of the cloned foot-and-mouth disease virus genome corresponding to all parts of the coding region were expressed in Escherichia coli as fusions to the N-terminal part of the MS2-polymerase gene under the control of the inducible λPL promoter. All constructs yielded large amounts of proteins, which were purified and used to raise sequence-specific antisera in rabbits. These antisera were used to identify the corresponding viral gene products in 35 S-labeled extracts from foot-and-mouth disease virus-infected BHK cells. This allowed us to locate unequivocally all mature foot-and-mouth disease virus gene products in the nucleotide sequence, to identify precursor-product relationships, and to detect several foot-and mouth disease virus gene products not previously identified in vivo or in vitro

Modulation of steroidogenic gene expression and hormone production of H295R cells by pharmaceuticals and other environmentally active compounds

International Nuclear Information System (INIS)

Gracia, Tannia; Hilscherova, Klara; Jones, Paul D.; Newsted, John L.; Higley, Eric B.; Zhang, Xiaowei; Hecker, Markus; Murphy, Margaret B.; Yu, Richard M.K.; Lam, Paul K.S.; Wu, Rudolf S.S.; Giesy, John P.

2007-01-01

The H295R cell bioassay was used to evaluate the potential endocrine disrupting effects of 18 of the most commonly used pharmaceuticals in the United States. Exposures for 48 h with single pharmaceuticals and binary mixtures were conducted; the expression of five steroidogenic genes, 3βHSD2, CYP11β1, CYP11β2, CYP17 and CYP19, was quantified by Q-RT-PCR. Production of the steroid hormones estradiol (E2), testosterone (T) and progesterone (P) was also evaluated. Antibiotics were shown to modulate gene expression and hormone production. Amoxicillin up-regulated the expression of CYP11β2 and CYP19 by more than 2-fold and induced estradiol production up to almost 3-fold. Erythromycin significantly increased CYP11β2 expression and the production of P and E2 by 3.5- and 2.4-fold, respectively, while production of T was significantly decreased. The β-blocker salbutamol caused the greatest induction of CYP17, more than 13-fold, and significantly decreased E2 production. The binary mixture of cyproterone and salbutamol significantly down-regulated expression of CYP19, while a mixture of ethynylestradiol and trenbolone, increased E2 production 3.7-fold. Estradiol production was significantly affected by changes in concentrations of trenbolone, cyproterone, and ethynylestradiol. Exposures with individual pharmaceuticals showed the possible secondary effects that drugs may exert on steroid production. Results from binary mixture exposures suggested the possible type of interactions that may occur between drugs and the joint effects product of such interactions. Dose-response results indicated that although two chemicals may share a common mechanism of action the concentration effects observed may be significantly different

Symbiotic Burkholderia Species Show Diverse Arrangements of nif/fix and nod Genes and Lack Typical High-Affinity Cytochrome cbb3 Oxidase Genes.

Science.gov (United States)

De Meyer, Sofie E; Briscoe, Leah; Martínez-Hidalgo, Pilar; Agapakis, Christina M; de-Los Santos, Paulina Estrada; Seshadri, Rekha; Reeve, Wayne; Weinstock, George; O'Hara, Graham; Howieson, John G; Hirsch, Ann M

2016-08-01

Genome analysis of fourteen mimosoid and four papilionoid beta-rhizobia together with fourteen reference alpha-rhizobia for both nodulation (nod) and nitrogen-fixing (nif/fix) genes has shown phylogenetic congruence between 16S rRNA/MLSA (combined 16S rRNA gene sequencing and multilocus sequence analysis) and nif/fix genes, indicating a free-living diazotrophic ancestry of the beta-rhizobia. However, deeper genomic analysis revealed a complex symbiosis acquisition history in the beta-rhizobia that clearly separates the mimosoid and papilionoid nodulating groups. Mimosoid-nodulating beta-rhizobia have nod genes tightly clustered in the nodBCIJHASU operon, whereas papilionoid-nodulating Burkholderia have nodUSDABC and nodIJ genes, although their arrangement is not canonical because the nod genes are subdivided by the insertion of nif and other genes. Furthermore, the papilionoid Burkholderia spp. contain duplications of several nod and nif genes. The Burkholderia nifHDKEN and fixABC genes are very closely related to those found in free-living diazotrophs. In contrast, nifA is highly divergent between both groups, but the papilionoid species nifA is more similar to alpha-rhizobia nifA than to other groups. Surprisingly, for all Burkholderia, the fixNOQP and fixGHIS genes required for cbb3 cytochrome oxidase production and assembly are missing. In contrast, symbiotic Cupriavidus strains have fixNOQPGHIS genes, revealing a divergence in the evolution of two distinct electron transport chains required for nitrogen fixation within the beta-rhizobia.

Nitrous oxide production and mRNA expression analysis of nitrifying and denitrifying bacterial genes under floodwater disappearance and fertilizer application.

Science.gov (United States)

Riya, Shohei; Takeuchi, Yuki; Zhou, Sheng; Terada, Akihiko; Hosomi, Masaaki

2017-06-01

A pulse of nitrous oxide (N 2 O) emission has been observed following the disappearance of floodwater by drainage. However, its mechanism is not well understood. We conducted a column study to clarify the mechanism for N 2 O production during floodwater disappearance by using a microsensor and determining the bacterial gene expression. An increase in N 2 O flux was observed following floodwater disappearance after the addition of NH 4 + , with a corresponding increase in the concentrations of NO 3 - and dissolved N 2 O in the oxic and anoxic soil layers, respectively. The transcription level of the bacterial amoA mRNA did not change, while that of nirK mRNA increased sharply after an hour of floodwater disappearance. An additional anoxic soil slurry experiment demonstrated that the addition of NO 3 - induced the expression of nirK gene and caused a concomitant increase in N 2 O production. These findings suggest that NO 3 - production in the oxic layers is important as it provides a substrate and induces the synthesis of denitrification enzymes in the anoxic layer during N 2 O production.

Increasing Avermectin Production in Streptomyces avermitilis by Manipulating the Expression of a Novel TetR-Family Regulator and Its Target Gene Product.

Science.gov (United States)

Liu, Wenshuai; Zhang, Qinling; Guo, Jia; Chen, Zhi; Li, Jilun; Wen, Ying

2015-08-01

Avermectins produced by Streptomyces avermitilis are commercially important anthelmintic agents. The detailed regulatory mechanisms of avermectin biosynthesis remain unclear. Here, we identified SAV3619, a TetR-family transcriptional regulator designated AveT, to be an activator for both avermectin production and morphological differentiation in S. avermitilis. AveT was shown to indirectly stimulate avermectin production by affecting transcription of the cluster-situated activator gene aveR. AveT directly repressed transcription of its own gene (aveT), adjacent gene pepD2 (sav_3620), sav_7490 (designated aveM), and sav_7491 by binding to an 18-bp perfect palindromic sequence (CGAAACGKTKYCGTTTCG, where K is T or G and Y is T or C and where the underlining indicates inverted repeats) within their promoter regions. aveM (which encodes a putative transmembrane efflux protein belonging to the major facilitator superfamily [MFS]), the important target gene of AveT, had a striking negative effect on avermectin production and morphological differentiation. Overexpression of aveT and deletion of aveM in wild-type and industrial strains of S. avermitilis led to clear increases in the levels of avermectin production. In vitro gel-shift assays suggested that C-5-O-B1, the late pathway precursor of avermectin B1, acts as an AveT ligand. Taken together, our findings indicate positive-feedback regulation of aveT expression and avermectin production by a late pathway intermediate and provide the basis for an efficient strategy to increase avermectin production in S. avermitilis by manipulation of AveT and its target gene product, AveM. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Relationship between Gene Body DNA Methylation and Intragenic H3K9me3 and H3K36me3 Chromatin Marks

OpenAIRE

Hahn, Maria A.; Wu, Xiwei; Li, Arthur X.; Hahn, Torsten; Pfeifer, Gerd P.

2011-01-01

To elucidate the relationship between intragenic DNA methylation and chromatin marks, we performed epigenetic profiling of chromosome 19 in human bronchial epithelial cells (HBEC) and in the colorectal cancer cell line HCT116 as well as its counterpart with double knockout of DNMT1 and DNMT3B (HCT116-DKO). Analysis of H3K36me3 profiles indicated that this intragenic mark of active genes is associated with two categories of genes: (i) genes with low CpG density and H3K9me3 in the gene body or ...

Isoepoxydon dehydrogenase (idh) gene expression in relation to patulin production by Penicillium expansum under different temperature and atmosphere.

Science.gov (United States)

De Clercq, N; Vlaemynck, G; Van Pamel, E; Van Weyenberg, S; Herman, L; Devlieghere, F; De Meulenaer, B; Van Coillie, E

2016-03-02

Penicillium expansum growth and patulin production occur mainly at post-harvest stage during the long-term storage of apples. Low temperature in combination with reduced oxygen concentrations is commonly applied as a control strategy to extend apple shelf life and supply the market throughout the year. Our in vitro study investigated the effect of temperature and atmosphere on expression of the idh gene in relation to the patulin production by P. expansum. The idh gene encodes the isoepoxydon dehydrogenase enzyme, a key enzyme in the patulin biosynthesis pathway. First, a reverse transcription real-time PCR (RT-qPCR) method was optimized to measure accurately the P. expansum idh mRNA levels relative to the mRNA levels of three reference genes (18S, β-tubulin, calmodulin), taking into account important parameters such as PCR inhibition and multiple reference gene stability. Subsequently, two P. expansum field isolates and one reference strain were grown on apple puree agar medium (APAM) under three conditions of temperature and atmosphere: 20 °C - air, 4 °C - air and 4 °C - controlled atmosphere (CA; 3% O2). When P. expansum strains reached a 0.5 and 2.0 cm colony diameter, idh expression and patulin concentrations were determined by means of the developed RT-qPCR and an HPLC-UV method, respectively. The in vitro study showed a clear reduction in patulin production and down-regulation of the idh gene expression when P. expansum was grown under 4 °C - CA. The results suggest that stress (low temperature and oxygen level) caused a delay of the fungal metabolism rather than a complete inhibition of toxin biosynthesis. A good correlation was found between the idh expression and patulin production, corroborating that temperature and atmosphere affected patulin production by acting at the transcriptional level of the idh gene. Finally, a reliable RT-qPCR can be considered as an alternative tool to investigate the effect of control strategies on the toxin formation in

redD and actII-ORF4, Pathway-Specific Regulatory Genes for Antibiotic Production in Streptomyces coelicolor A3(2), Are Transcribed In Vitro by an RNA Polymerase Holoenzyme Containing σhrdD

NARCIS (Netherlands)

Fujii, T.; Gramajo, H.C.; Takano, E.; Bibb, M.J.

1996-01-01

redD and actII-ORF4, regulatory genes required for synthesis of the antibiotics undecylprodigiosin and actinorhodin by Streptomyces coelicolor A3(2), were transcribed in vitro by an RNA polymerase holoenzyme containing σhrdD. Disruption of hrdD had no effect on antibiotic production, indicating that

Cell surface expression system for the display of heterologous gene products using chimeric flagellin fusions of bacillus halodurans isolate

CSIR Research Space (South Africa)

Du Plessis, A

2006-10-01

Full Text Available system for the display of heterologous gene products using chimeric flagellin fusions of a Bacillus halodurans isolate Slide 2 © CSIR 2006 www.csir.co.za Bacillus halodurans Alk 36 xrhombus Ability to over-produce cell... for functionality of the His-tag for metal binding. Slide 13 © CSIR 2006 www.csir.co.za PAGE gel showing over-production of chimeric poly-His flagellin proteins 66.2 kDa 45.0 kDa 31.0 kDa 1. LMW ladder 2. NC3 3. NHisC3 4. NC6 5...

Identification and localization of a gene that specifies production of Escherichia coli DNA topoisomerase I

International Nuclear Information System (INIS)

Trucksis, M.; Depew, R.E.

1981-01-01

A gene that specifies production of Escherichia coli DNA topoisomerase I (ω protein) was identified with the aid of a radioimmunoassay for this protein. E. coli DNA topoisomerase I was produced by Salmonella typhimurium merodiploids that harbored E. coli plasmid F' 123, but not by strains that lost this plasmid. Analysis of strains with spontaneous deletions of F' 123 showed that the gene, topA, required for production of the E. coli ω protein was between the trp operon and the cysB gene. Deletions that eliminated topA also eliminated the supX gene. We suggest that topA is the structural gene of E. coli DNA topoisomerase I and that topA is identical to supX

Disruption of ten protease genes in the filamentous fungus Aspergillus oryzae highly improves production of heterologous proteins.

Science.gov (United States)

Yoon, Jaewoo; Maruyama, Jun-ichi; Kitamoto, Katsuhiko

2011-02-01

Proteolytic degradation by secreted proteases into the culture medium is one of the significant problems to be solved in heterologous protein production by filamentous fungi including Aspergillus oryzae. Double (tppA, and pepE) and quintuple (tppA, pepE, nptB, dppIV, and dppV) disruption of protease genes enhanced human lysozyme (HLY) and bovine chymosin (CHY) production by A. oryzae. In this study, we used a quintuple protease gene disruptant and performed successive rounds of disruption for five additional protease genes (alpA, pepA, AopepAa, AopepAd, and cpI), which were previously investigated by DNA microarray analyses for their expression. Gene disruption was performed by pyrG marker recycling with a highly efficient gene-targeting background (∆ligD) as previously reported. As a result, the maximum yields of recombinant CHY and HLY produced by a decuple protease gene disruptant were approximately 30% and 35%, respectively, higher than those produced by a quintuple protease gene disruptant. Thus, we successfully constructed a decuple protease gene disruptant possessing highly improved capability of heterologous protein production. This is the first report on decuple protease gene disruption that improved the levels of heterologous protein production by the filamentous fungus A. oryzae.

Single gene insertion drives bioalcohol production by a thermophilic archaeon

Energy Technology Data Exchange (ETDEWEB)

Basen, M; Schut, GJ; Nguyen, DM; Lipscomb, GL; Benn, RA; Prybol, CJ; Vaccaro, BJ; Poole, FL; Kelly, RM; Adams, MWW

2014-12-09

Bioethanol production is achieved by only two metabolic pathways and only at moderate temperatures. Herein a fundamentally different synthetic pathway for bioalcohol production at 70 degrees C was constructed by insertion of the gene for bacterial alcohol dehydrogenase (AdhA) into the archaeon Pyrococcus furiosus. The engineered strain converted glucose to ethanol via acetate and acetaldehyde, catalyzed by the host-encoded aldehyde ferredoxin oxidoreductase (AOR) and heterologously expressed AdhA, in an energy-conserving, redox-balanced pathway. Furthermore, the AOR/AdhA pathway also converted exogenously added aliphatic and aromatic carboxylic acids to the corresponding alcohol using glucose, pyruvate, and/or hydrogen as the source of reductant. By heterologous coexpression of a membrane-bound carbon monoxide dehydrogenase, CO was used as a reductant for converting carboxylic acids to alcohols. Redirecting the fermentative metabolism of P. furiosus through strategic insertion of foreign genes creates unprecedented opportunities for thermophilic bioalcohol production. Moreover, the AOR/AdhA pathway is a potentially game-changing strategy for syngas fermentation, especially in combination with carbon chain elongation pathways.

Methodological issues in detecting gene-gene interactions in breast cancer susceptibility: a population-based study in Ontario

Directory of Open Access Journals (Sweden)

Onay Venus

2007-08-01

Full Text Available Abstract Background There is growing evidence that gene-gene interactions are ubiquitous in determining the susceptibility to common human diseases. The investigation of such gene-gene interactions presents new statistical challenges for studies with relatively small sample sizes as the number of potential interactions in the genome can be large. Breast cancer provides a useful paradigm to study genetically complex diseases because commonly occurring single nucleotide polymorphisms (SNPs may additively or synergistically disturb the system-wide communication of the cellular processes leading to cancer development. Methods In this study, we systematically studied SNP-SNP interactions among 19 SNPs from 18 key genes involved in major cancer pathways in a sample of 398 breast cancer cases and 372 controls from Ontario. We discuss the methodological issues associated with the detection of SNP-SNP interactions in this dataset by applying and comparing three commonly used methods: the logistic regression model, classification and regression trees (CART, and the multifactor dimensionality reduction (MDR method. Results Our analyses show evidence for several simple (two-way and complex (multi-way SNP-SNP interactions associated with breast cancer. For example, all three methods identified XPD-[Lys751Gln]*IL10-[G(-1082A] as the most significant two-way interaction. CART and MDR identified the same critical SNPs participating in complex interactions. Our results suggest that the use of multiple statistical approaches (or an integrated approach rather than a single methodology could be the best strategy to elucidate complex gene interactions that have generally very different patterns. Conclusion The strategy used here has the potential to identify complex biological relationships among breast cancer genes and processes. This will lead to the discovery of novel biological information, which will improve breast cancer risk management.

Diversity, distribution and quantification of antibiotic resistance genes in goat and lamb slaughterhouse surfaces and meat products.

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Leyre Lavilla Lerma

Full Text Available The distribution and quantification of tetracycline, sulfonamide and beta-lactam resistance genes were assessed in slaughterhouse zones throughout meat chain production and the meat products; this study represents the first to report quantitatively monitor antibiotic resistance genes (ARG in goat and lamb slaughterhouse using a culture independent approach, since most studies focused on individual bacterial species and their specific resistance types. Quantitative PCR (qPCR revealed a high prevalence of tetracycline resistance genes tetA and tetB in almost all slaughterhouse zones. Sulfonamide resistance genes were largely distributed, while beta-lactam resistance genes were less predominant. Statistical analysis revealed that resistant bacteria, in most cases, were spread by the same route in almost all slaughterhouse zones, except for tetB, blaCTX and blaTEM genes, which occurred in few zones as isolated 'hot spots.' The sum of all analyzed ARG indicated that slaughterhouse surfaces and end products act as reservoirs of ARG, mainly tet genes, which were more prevalent in slaughtering room (SR, cutting room (CR and commercial meat products (MP. Resistance gene patterns suggest they were disseminated throughout slaughterhouse zones being also detected in commercial meat products, with significant correlations between different sampling zones/end products and total resistance in SR, CR and white room (WR zones, and also refrigerator 4 (F4 and MP were observed. Strategically controlling key zones in slaughterhouse (SR, CR and WR by adequate disinfection methods could strategically reduce the risks of ARG transmission and minimize the issues of food safety and environment contamination.

Diversity, distribution and quantification of antibiotic resistance genes in goat and lamb slaughterhouse surfaces and meat products.

Science.gov (United States)

Lavilla Lerma, Leyre; Benomar, Nabil; Knapp, Charles W; Correa Galeote, David; Gálvez, Antonio; Abriouel, Hikmate

2014-01-01

The distribution and quantification of tetracycline, sulfonamide and beta-lactam resistance genes were assessed in slaughterhouse zones throughout meat chain production and the meat products; this study represents the first to report quantitatively monitor antibiotic resistance genes (ARG) in goat and lamb slaughterhouse using a culture independent approach, since most studies focused on individual bacterial species and their specific resistance types. Quantitative PCR (qPCR) revealed a high prevalence of tetracycline resistance genes tetA and tetB in almost all slaughterhouse zones. Sulfonamide resistance genes were largely distributed, while beta-lactam resistance genes were less predominant. Statistical analysis revealed that resistant bacteria, in most cases, were spread by the same route in almost all slaughterhouse zones, except for tetB, blaCTX and blaTEM genes, which occurred in few zones as isolated 'hot spots.' The sum of all analyzed ARG indicated that slaughterhouse surfaces and end products act as reservoirs of ARG, mainly tet genes, which were more prevalent in slaughtering room (SR), cutting room (CR) and commercial meat products (MP). Resistance gene patterns suggest they were disseminated throughout slaughterhouse zones being also detected in commercial meat products, with significant correlations between different sampling zones/end products and total resistance in SR, CR and white room (WR) zones, and also refrigerator 4 (F4) and MP were observed. Strategically controlling key zones in slaughterhouse (SR, CR and WR) by adequate disinfection methods could strategically reduce the risks of ARG transmission and minimize the issues of food safety and environment contamination.

Functional and bioinformatics analysis of an exopolysaccharide-related gene (epsN) from Lactobacillus kefiranofaciens ZW3.

Science.gov (United States)

Wang, Jingrui; Tang, Wei; Zheng, Yongna; Xing, Zhuqing; Wang, Yanping

2016-09-01

A novel lactic acid bacteria strain Lactobacillus kefiranofaciens ZW3 exhibited the characteristics of high production of exopolysaccharide (EPS). The epsN gene, located in the eps gene cluster of this strain, is associated with EPS biosynthesis. Bioinformatics analysis of this gene was performed. The conserved domain analysis showed that the EpsN protein contained MATE-Wzx-like domains. Then the epsN gene was amplified to construct the recombinant expression vector pMG36e-epsN. The results showed that the EPS yields of the recombinants were significantly improved. By determining the yields of EPS and intracellular polysaccharide, it was considered that epsN gene could play its Wzx flippase role in the EPS biosynthesis. This is the first time to prove the effect of EpsN on L. kefiranofaciens EPS biosynthesis and further prove its functional property.

The NtAMI1 gene functions in cell division of tobacco BY-2 cells in the presence of indole-3-acetamide.

Science.gov (United States)

Nemoto, Keiichirou; Hara, Masamitsu; Suzuki, Masashi; Seki, Hikaru; Muranaka, Toshiya; Mano, Yoshihiro

2009-01-22

Tobacco (Nicotiana tabacum) Bright Yellow-2 (BY-2) cells can be grown in medium containing indole-3-acetamide (IAM). Based on this finding, the NtAMI1 gene, whose product is functionally equivalent to the AtAMI1 gene of Arabidopsis thaliana and the aux2 gene of Agrobacterium rhizogenes, was isolated from BY-2 cells. Overexpression of the NtAMI1 gene allowed BY-2 cells to proliferate at lower concentrations of IAM, whereas suppression of the NtAMI1 gene by RNA interference (RNAi) caused severe growth inhibition in the medium containing IAM. These results suggest that IAM is incorporated into plant cells and converted to the auxin, indole-3-acetic acid, by NtAMI1.

A shortest-path graph kernel for estimating gene product semantic similarity

Directory of Open Access Journals (Sweden)

Alvarez Marco A

2011-07-01

Full Text Available Abstract Background Existing methods for calculating semantic similarity between gene products using the Gene Ontology (GO often rely on external resources, which are not part of the ontology. Consequently, changes in these external resources like biased term distribution caused by shifting of hot research topics, will affect the calculation of semantic similarity. One way to avoid this problem is to use semantic methods that are "intrinsic" to the ontology, i.e. independent of external knowledge. Results We present a shortest-path graph kernel (spgk method that relies exclusively on the GO and its structure. In spgk, a gene product is represented by an induced subgraph of the GO, which consists of all the GO terms annotating it. Then a shortest-path graph kernel is used to compute the similarity between two graphs. In a comprehensive evaluation using a benchmark dataset, spgk compares favorably with other methods that depend on external resources. Compared with simUI, a method that is also intrinsic to GO, spgk achieves slightly better results on the benchmark dataset. Statistical tests show that the improvement is significant when the resolution and EC similarity correlation coefficient are used to measure the performance, but is insignificant when the Pfam similarity correlation coefficient is used. Conclusions Spgk uses a graph kernel method in polynomial time to exploit the structure of the GO to calculate semantic similarity between gene products. It provides an alternative to both methods that use external resources and "intrinsic" methods with comparable performance.

Beta 2 adrenergic receptor polymorphisms, at codons 16 and 27, and bronchodilator responses in adult Venezuelan asthmatic patients.

Science.gov (United States)

Larocca, Nancy; Moreno, Dolores; Garmendia, Jenny Valentina; Velasquez, Olga; Martin-Rojo, Joana; Talamo, Carlos; Garcia, Alexis; De Sanctis, Juan Bautista

2013-12-01

One of the gene polymorphisms often studied in asthmatic patients is the β2 adrenergic receptor (ADRβ2). Even though in the Venezuelan Mestizo population there is a high incidence of asthma, there are no direct reports of ADRβ2 gene polymorphism, and treatment response. The aim of this study was to assess, in this population, the gene frequency of ADRβ2 polymorphisms at codons 16 Arg/Gly and 27 Gln/Glu, allergen sensitization, and its relationship to bronchodilator response. Purified genomic DNA was obtained form 105 Mestizo asthmatic and 100 Mestizo healthy individuals from Venezuela. The two polymorphisms were assessed by PCR-RFLP. Patient sensitization to aeroallergens and their response to bronchodilatation were correlated. Significant differences between patients and controls were recorded in: 1) the prevalence of Arg/Arg at codon 16 (28.6% in patients vs. 47% in controls, P<0.01), 2) the frequency of heterozygotes Arg/Gly (55% in patients vs. 35% in controls, P<0.01). Conversely, no differences in polymorphism frequencies were found at codon 27. The haplotypes Arg/Gly-Gln/Gln were more common in patients than controls (P <0.01), whereas the Arg/Arg-Gln/Glu combination prevailed in the control group (P<0.01). The Arg/Gly and Gln/Glu genotypes were associated with better responses after salbutamol. The asthmatic homozygotes Arg/Arg have higher sensitivity to aeroallergens. The difference in Arg/Arg frequency between groups suggests that this could be a protective genotype although the asthmatic group had a higher sensitivity to aeroallergens. The asthmatic heterozygotes had better bronchodilator responses than the homozygotes.

Use of endophytic diazotrophic bacteria as a vector to express the cry3A gene from Bacillus thuringiensis

Directory of Open Access Journals (Sweden)

Salles Joana Falcão

2000-01-01

Full Text Available The goal of this study was to evaluate the potential of endophytic diazotrophic bacteria as a vector to express a cry gene from Bacillus thuringiensis, envisaging the control of pests that attack sugarcane plants. The endophytic nitrogen-fixing bacteria Gluconacetobacter diazotrophicus strain BR11281 and Herbaspirillum seropedicae strain BR11335 were used as models. The cry3A gene was transferred by conjugation using a suicide plasmid and the recombinant strains were selected by their ability to fix nitrogen in semi-solid N-free medium. The presence of the cry gene was detected by Southern-blot using an internal fragment of 1.0 kb as a probe. The production of delta-endotoxin by the recombinant H. seropedicae strain was detected by dot blot while for G. diazotrophicus the Western-blot technique was used. In both cases, a specific antibody raised against the B. thuringiensis toxin was applied. The delta-endotoxin production showed by the G. diazotrophicus recombinant strain was dependent on the nitrogen fixing conditions since the cry3A gene was fused to a nif promoter. In the case of H. seropedicae the delta-endotoxin expression was not affected by the promoter (rhi used. These results suggest that endophytic diazotrophic bacteria can be used as vectors to express entomopathogenic genes envisaging control of sugarcane pests.

Biallelic loss-of-function variants in DOCK3 cause muscle hypotonia, ataxia, and intellectual disability.

Science.gov (United States)

Helbig, K L; Mroske, C; Moorthy, D; Sajan, S A; Velinov, M

2017-10-01

DOCK3 encodes the dedicator of cytokinesis 3 protein, a member of the DOCK180 family of proteins that are characterized by guanine-nucleotide exchange factor activity. DOCK3 is expressed exclusively in the central nervous system and plays an important role in axonal outgrowth and cytoskeleton reorganization. Dock3 knockout mice exhibit motor deficiencies with abnormal ataxic gait and impaired learning. We report 2 siblings with biallelic loss-of-function variants in DOCK3. Diagnostic whole-exome sequencing (WES) and chromosomal microarray were performed on a proband with severe developmental disability, hypotonia, and ataxic gait. Testing was also performed on the proband's similarly affected brother. A paternally inherited 458 kb deletion in chromosomal region 3p21.2 disrupting the DOCK3 gene was identified in both affected siblings. WES identified a nonsense variant c.382C>G (p.Gln128*) in the DOCK3 gene (NM_004947) on the maternal allele in both siblings. Common features in both affected individuals include severe developmental disability, ataxic gait, and severe hypotonia, which recapitulates the Dock3 knockout mouse phenotype. We show that complete DOCK3 deficiency in humans leads to developmental disability with significant hypotonia and gait ataxia, probably due to abnormal axonal development. © 2017 The Authors. Clinical Genetics published by John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Molecular screening of the ghrelin gene in Italian obese children: the Leu72Met variant is associated with an earlier onset of obesity.

Science.gov (United States)

Miraglia del Giudice, E; Santoro, N; Cirillo, G; Raimondo, P; Grandone, A; D'Aniello, A; Di Nardo, M; Perrone, L

2004-03-01

To test whether ghrelin variants could play a role in modulating some aspects of the obese phenotype during childhood. We screened the ghrelin gene in 300 Italian obese children and adolescents (mean age 10.5+/-3.2 y; range 4-19 y) and 200 controls by using the single-strand conformation polymorphism and the restriction fragment length polymoprhism analysis. No mutations were detected with the exception of two previously described polymorphisms, Arg51Gln and Leu72Met. For both variations, allelic frequencies were similar between patients and controls. Interestingly, we showed that the Leu72Met polymorphism was associated with differences in the age at obesity onset. Patients with the Met72 allele became obese earlier than homozygous patients for the wild Leu72 allele. The logrank test comparing the plots of the complement of Kaplan-Meier estimates between the two groups of patients was statistically significant (Pghrelin variations cause the obesity due to single-gene mutations. The Leu72Met polymorphism of the ghrelin gene seems to play a role in anticipating the onset of obesity among children suggesting, therefore, that ghrelin may be involved in the pathophysiology of human adiposity.