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1.
Gene ; 760: 145018, 2020 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-32758580

RESUMO

Protein turnover is a process that is regulated by several factors and can lead to muscle hypertrophy or atrophy. The purpose of the present study was to determine the effects of ß-hydroxy-ß-methylbutyrate free acid (HMB-FA) and eccentric resistance exercise on variables related to protein turnover in rats. Thirty-two male rats were randomly assigned into four groups of eight, including control, control-HMB, exercise, and exercise-HMB. Animals in HMB groups received 340 mg/kg/day for two weeks. Animals in the exercise groups performed one session of eccentric resistance exercise consisting of eight repetitions descending from a ladder with a slope of 80 degree, with an extra load of two times body weight (100% 1RM). Twenty-four hours after the exercise session, triceps brachii muscle and serum were collected for further analysis. Exercise and HMB-FA induced lower muscle myostatin and higher muscle Fibronectin type III domain containing 5 (FNDC5), P70-S6 kinase 1 gene expression, as well as higher serum irisin and IGF-1 concentrations. Exercise alone induced higher caspase-3 and caspase-8 gene expression while HMB-FA alone induced lower caspase 3 gene expression. HMB-FA supplement increased the effect of exercise on muscle FNDC5, myostatin, and P70-S6 kinase 1 gene expression. The interaction of exercise and HMBFA resulted in an additive effect, increasing serum irisin and IGF-1 concentrations. In conclusion, a 2-week HMB-FA supplementation paired with acute eccentric resistance exercise can positively affect some genes related to muscle protein turnover.


Assuntos
Proteínas Musculares/efeitos dos fármacos , Valeratos/farmacologia , Animais , Suplementos Nutricionais , Fibronectinas/efeitos dos fármacos , Fibronectinas/metabolismo , Genes Reguladores/genética , Fator de Crescimento Insulin-Like I/metabolismo , Masculino , Proteínas Musculares/metabolismo , Músculo Esquelético/metabolismo , Miostatina/efeitos dos fármacos , Miostatina/metabolismo , Condicionamento Físico Animal/métodos , Ratos , Ratos Sprague-Dawley , Treinamento de Resistência/métodos , Proteínas Quinases S6 Ribossômicas 70-kDa/efeitos dos fármacos , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo
2.
Zhongguo Shi Yan Xue Ye Xue Za Zhi ; 28(4): 1414-1418, 2020 Aug.
Artigo em Chinês | MEDLINE | ID: mdl-32798436

RESUMO

Acute myeloid leukemia(AML)is a myelopoietic stem/progenitor malignant disease. The exact etiology of this leukemia remains unclear, thus it is important to explore the pathogenesis of AML and to discover the new diagnostic markers and therapeutic targets. The long non coding RNA (lnc RNA) is a class of RNA molecules with transcripts over 200 nucleotides in eukaryotic cells which almost don't possess the ability to code proteins, but can regulate the expression of other genes at transcriptional and post-transcriptional levels, thereby participate in occurrence and development of varied tumors. Of late years, along with the deepening of study, the lncRNA roles played in the AML have been reported and confirmed. In this review, the relationships between the IncRNA (UCA1, ANRIL, H19, HOTAIR, CCAT1, ZFAS1, LINC00152, HOXA-A52, NEAT1, TUG1, IRAIN1, PANDAR, LINC00899, SNHG5, and KCNQ1OT1) and AML is summarized briefly, so as to provide the potential basis for the clinical diagnosis and therapy of AML.


Assuntos
Leucemia Mieloide Aguda/genética , RNA Longo não Codificante , Genes Reguladores , Humanos , Prognóstico
3.
BMC Bioinformatics ; 21(1): 318, 2020 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-32690031

RESUMO

BACKGROUND: Gene Regulatory Networks (GRNs) have been previously studied by using Boolean/multi-state logics. While the gene expression values are usually scaled into the range [0, 1], these GRN inference methods apply a threshold to discretize the data, resulting in missing information. Most of studies apply fuzzy logics to infer the logical gene-gene interactions from continuous data. However, all these approaches require an a priori known network structure. RESULTS: Here, by introducing a new probabilistic logic for continuous data, we propose a novel logic-based approach (called the LogicNet) for the simultaneous reconstruction of the GRN structure and identification of the logics among the regulatory genes, from the continuous gene expression data. In contrast to the previous approaches, the LogicNet does not require an a priori known network structure to infer the logics. The proposed probabilistic logic is superior to the existing fuzzy logics and is more relevant to the biological contexts than the fuzzy logics. The performance of the LogicNet is superior to that of several Mutual Information-based and regression-based tools for reconstructing GRNs. CONCLUSIONS: The LogicNet reconstructs GRNs and logic functions without requiring prior knowledge of the network structure. Moreover, in another application, the LogicNet can be applied for logic function detection from the known regulatory genes-target interactions. We also conclude that computational modeling of the logical interactions among the regulatory genes significantly improves the GRN reconstruction accuracy.


Assuntos
Algoritmos , Biologia Computacional/métodos , Escherichia coli/genética , Lógica Fuzzy , Redes Reguladoras de Genes , Genes Reguladores , Modelos Genéticos , Simulação por Computador , Escherichia coli/metabolismo , Perfilação da Expressão Gênica
4.
Arch Microbiol ; 202(9): 2453-2459, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32607723

RESUMO

Determinant genes controlling biofilm formation in a plant commensal bacterium, Pseudomonas protegens Pf-5, were identified by transposon mutagenesis. Comprehensive screening of 7500 transposon-inserted mutants led to the isolation of four mutants exhibiting decreased and five mutants exhibiting increased biofilm formation. Mutations in the genes encoding MFS drug resistance transporter, LapA adhesive protein, RetS sensor histidine kinase/response regulator, and HecA adhesin/hemagglutinin led to decreased biofilm formation, indicating that these genes are necessary for biofilm formation in Pf-5. The mutants exhibiting increased biofilm formation had transposon insertions in the genes coding for an outer membrane protein, a GGDEF domain-containing protein, AraC transcriptional regulator, non-ribosomal peptide synthetase OfaB, and the intergenic region of a DNA-binding protein and the Aer aerotaxis receptor, suggesting that these genes are negative regulators of biofilm formation. Some of these mutants also showed altered swimming and swarming motilities, and a negative correlation between biofilm formation and swarming motility was observed. Thus, sessile-motile lifestyle is regulated by divergent regulatory genes in Pf-5.


Assuntos
Proteínas de Bactérias/genética , Biofilmes , Pseudomonas/genética , Adesinas Bacterianas/genética , Adesinas Bacterianas/metabolismo , Genes Reguladores , Histidina Quinase/genética , Mutação
5.
PLoS One ; 15(5): e0232986, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32407419

RESUMO

Here we focus on the highly conserved MYB-bHLH-WD repeat (MBW) transcriptional complex model in eggplant, which is pivotal in the transcriptional regulation of the anthocyanin biosynthetic pathway. Through a genome-wide approach performed on the recently released Eggplant Genome (cv. 67/3) previously identified, and reconfirmed by us, members belonging to the MBW complex (SmelANT1, SmelAN2, SmelJAF13, SmelAN1) were functionally characterized. Furthermore, a regulatory R3 MYB type repressor (SmelMYBL1), never reported before, was identified and characterized as well. Through a qPCR approach, we revealed specific transcriptional patterns of candidate genes in different plant tissue/organs at two stages of fruit development. Two strategies were adopted for investigating the interactions of bHLH partners (SmelAN1, SmelJAF13) with MYB counterparts (SmelANT1, SmelAN2 and SmelMYBL1): Yeast Two Hybrid (Y2H) and Bimolecular Fluorescent Complementation (BiFC) in A. thaliana mesophylls protoplast. Agro-infiltration experiments highlighted that N. benthamiana leaves transiently expressing SmelANT1 and SmelAN2 showed an anthocyanin-pigmented phenotype, while their co-expression with SmelMYBL1 prevented anthocyanin accumulation. Our results suggest that SmelMYBL1 may inhibits the MBW complex via the competition with MYB activators for bHLH binding site, although this hypothesis requires further elucidation.


Assuntos
Antocianinas/biossíntese , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Solanum melongena/genética , Solanum melongena/metabolismo , Sequência de Aminoácidos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação da Expressão Gênica de Plantas , Genes de Plantas , Genes Reguladores , Família Multigênica , Filogenia , Plantas Geneticamente Modificadas , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Tabaco/genética , Tabaco/metabolismo
6.
Nucleic Acids Res ; 48(11): e62, 2020 06 19.
Artigo em Inglês | MEDLINE | ID: mdl-32329779

RESUMO

Recent advances in genomic technologies have generated data on large-scale protein-DNA interactions and open chromatin regions for many eukaryotic species. How to identify condition-specific functions of transcription factors using these data has become a major challenge in genomic research. To solve this problem, we have developed a method called ConSReg, which provides a novel approach to integrate regulatory genomic data into predictive machine learning models of key regulatory genes. Using Arabidopsis as a model system, we tested our approach to identify regulatory genes in data sets from single cell gene expression and from abiotic stress treatments. Our results showed that ConSReg accurately predicted transcription factors that regulate differentially expressed genes with an average auROC of 0.84, which is 23.5-25% better than enrichment-based approaches. To further validate the performance of ConSReg, we analyzed an independent data set related to plant nitrogen responses. ConSReg provided better rankings of the correct transcription factors in 61.7% of cases, which is three times better than other plant tools. We applied ConSReg to Arabidopsis single cell RNA-seq data, successfully identifying candidate regulatory genes that control cell wall formation. Our methods provide a new approach to define candidate regulatory genes using integrated genomic data in plants.


Assuntos
Arabidopsis/genética , Redes Reguladoras de Genes , Genes de Plantas/genética , Genes Reguladores , Aprendizado de Máquina , Proteínas de Arabidopsis , Conjuntos de Dados como Assunto , Aprendizado Profundo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Regiões Promotoras Genéticas/genética , RNA-Seq , Análise de Célula Única , Estresse Fisiológico/genética , Fatores de Transcrição
7.
Microbiol Immunol ; 64(6): 469-475, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32227523

RESUMO

We performed RNA sequencing on Bordetella pertussis, the causative agent of whooping cough, and identified nine novel small RNAs (sRNAs) that were transcribed during the bacterial colonization of murine tracheas. Among them, four sRNAs were more strongly expressed in vivo than in vitro. Moreover, the expression of eight sRNAs was not regulated by the BvgAS two-component system, which is the master regulator for the expression of genes contributing to the bacterial infection. The present results suggest a BvgAS-independent gene regulatory system involving the sRNAs that is active during B. pertussis infection.


Assuntos
Bordetella pertussis , RNA Bacteriano/genética , Pequeno RNA não Traduzido/genética , Traqueia/microbiologia , Coqueluche/microbiologia , Animais , Proteínas de Bactérias/genética , Bordetella pertussis/genética , Bordetella pertussis/patogenicidade , Regulação Bacteriana da Expressão Gênica/genética , Genes Reguladores/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Traqueia/patologia , Fatores de Transcrição/genética , Virulência/genética , Fatores de Virulência de Bordetella/genética
8.
PLoS One ; 15(3): e0229843, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32150571

RESUMO

Seasonal phenomena in plants are primarily affected by day length and temperature. The shoot transcriptomes of trees grown in the field and a controlled-environment chamber were compared to characterize genes that control annual rhythms and the effects of day length- and temperature-regulated genes in the gymnosperm Japanese cedar (Cryptomeria japonica D. Don), which exhibits seasonally indeterminate growth. Annual transcriptome dynamics were clearly demonstrated by principal component analysis using microarray data obtained under field-grown conditions. Analysis of microarray data from trees grown in a controlled chamber identified 2,314 targets exhibiting significantly different expression patterns under short-day (SD) and long-day conditions, and 2,045 targets exhibited significantly different expression patterns at 15°C (LT; low temperature) versus 25°C. Interestingly, although growth was suppressed under both SD and LT conditions, approximately 80% of the SD- and LT-regulated targets differed, suggesting that each factor plays a unique role in the annual cycle. The top 1,000 up-regulated targets in the growth/dormant period in the field coincided with more than 50% of the SD- and LT-regulated targets, and gene co-expression network analysis of the annual transcriptome indicated a close relationship between the SD- and LT-regulated targets. These results indicate that the respective effects of day length and temperature interact to control annual transcriptome dynamics. Well-known upstream genes of signaling pathways responsive to environmental conditions, such as the core clock (LHY/CjLHYb and CCA1/CjLHYa) and PEBP family (MFT) genes, exhibited unique expression patterns in Japanese cedar compared with previous reports in other species, suggesting that these genes control differences in seasonal regulation mechanisms between species. The results of this study provide new insights into seasonal regulation of transcription in Japanese cedar.


Assuntos
Cryptomeria/genética , Regulação da Expressão Gênica de Plantas , Estações do Ano , Temperatura , Transcriptoma , Cycadopsida/genética , Genes de Plantas/genética , Genes Reguladores , Árvores/genética , Árvores/metabolismo
9.
Nat Microbiol ; 5(5): 679-687, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32203410

RESUMO

CRISPR-Cas systems are adaptive immune systems that protect bacteria from bacteriophage (phage) infection1. To provide immunity, RNA-guided protein surveillance complexes recognize foreign nucleic acids, triggering their destruction by Cas nucleases2. While the essential requirements for immune activity are well understood, the physiological cues that regulate CRISPR-Cas expression are not. Here, a forward genetic screen identifies a two-component system (KinB-AlgB), previously characterized in the regulation of Pseudomonas aeruginosa alginate biosynthesis3,4, as a regulator of the expression and activity of the P. aeruginosa Type I-F CRISPR-Cas system. Downstream of KinB-AlgB, activators of alginate production AlgU (a σE orthologue) and AlgR repress CRISPR-Cas activity during planktonic and surface-associated growth5. AmrZ, another alginate regulator6, is triggered to repress CRISPR-Cas immunity upon surface association. Pseudomonas phages and plasmids have taken advantage of this regulatory scheme and carry hijacked homologs of AmrZ that repress CRISPR-Cas expression and activity. This suggests that while CRISPR-Cas regulation may be important to limit self-toxicity, endogenous repressive pathways represent a vulnerability for parasite manipulation.


Assuntos
Alginatos/metabolismo , Bactérias/metabolismo , Bactérias/virologia , Bacteriófagos/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Bactérias/genética , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sequência de Bases , Sistemas CRISPR-Cas , Proteínas de Ligação a DNA/genética , Regulação Bacteriana da Expressão Gênica , Genes Bacterianos/genética , Genes Reguladores/genética , Imunidade , Fagos de Pseudomonas/genética , Pseudomonas aeruginosa/metabolismo , Fatores de Transcrição , Transcrição Genética
10.
Gene ; 739: 144508, 2020 May 20.
Artigo em Inglês | MEDLINE | ID: mdl-32097695

RESUMO

Pesticides exposure can have harmful effects on human health. The liver is the most common organ of pesticides toxicity due to its major metabolic activity. The molecular mechanism of pesticides effect is complex and is controlled by gene regulatory networks. All components of regulatory networks are controlled by transcription factors and other regulatory elements. Therefore, identification of key regulators through system biology approaches and high-throughput techniques can help to provide comprehensive insights into molecular mechanisms of the pesticide effect. In the current study, a microarray data-set was used to potentially identify molecular mechanisms that regulate gene expression profile of rat hepatocyte cell lines in response to pesticides exposure. Results showed that the number of differentially expressed genes (DEGs) and differentially expressed transcription factors (DE-TFs) were dramatically different among pesticides tested. Results also revealed 205 common DEGs and 11 DE-TFs among pesticides tested. Additionally, we found that six DE-TFs (CREB1, CTNNB1, PPARG, SP1, SRF and STAT3) had the highest number of interactions with other DEGs and acted as the key regulatory genes. The results of this study revealed regulator genes that have the key functions in response to pesticides toxicity in rat liver, which can provide the basis for future studies. Furthermore, these regulatory genes can be used as toxicity biomarkers to improve diagnosis and prognosis.


Assuntos
Biologia Computacional , Redes Reguladoras de Genes/efeitos dos fármacos , Genes Reguladores/genética , Praguicidas/toxicidade , Fatores de Transcrição/genética , Transcriptoma/efeitos dos fármacos , Animais , Linhagem Celular , Perfilação da Expressão Gênica , Hepatócitos/efeitos dos fármacos , Fígado/efeitos dos fármacos , Especificidade de Órgãos , Ratos
11.
PLoS One ; 15(2): e0229351, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32078641

RESUMO

Polycystic ovary syndrome (PCOS) affects around 10% of young women, with adverse consequences on fertility and cardiometabolic outcomes. PCOS appears to result from a genetic predisposition interacting with developmental events during fetal or perinatal life. We hypothesised that PCOS candidate genes might be expressed in the fetal ovary when the stroma develops; mechanistically linking the genetics, fetal origins and adult ovarian phenotype of PCOS. In bovine fetal ovaries (n = 37) of 18 PCOS candidate genes only SUMO1P1 was not expressed. Three patterns of expression were observed: early gestation (FBN3, GATA4, HMGA2, TOX3, DENND1A, LHCGR and FSHB), late gestation (INSR, FSHR, and LHCGR) and throughout gestation (THADA, ERBB4, RAD50, C8H9orf3, YAP1, RAB5B, SUOX and KRR1). A splice variant of FSHB exon 3 was also detected early in the bovine ovaries, but exon 2 was not detected. Three other genes, likely to be related to the PCOS aetiology (AMH, AR and TGFB1I1), were also expressed late in gestation. Significantly within each of the three gene groups, the mRNA levels of many genes were highly correlated with each other, despite, in some instances, being expressed in different cell types. TGFß is a well-known stimulator of stromal cell replication and collagen synthesis and TGFß treatment of cultured fetal ovarian stromal cells inhibited the expression of INSR, AR, C8H9orf3 and RAD50 and stimulated the expression of TGFB1I1. In human ovaries (n = 15, < 150 days gestation) many of the same genes as in bovine (FBN3, GATA4, HMGA2, FSHR, DENND1A and LHCGR but not TOX3 or FSHB) were expressed and correlated with each other. With so many relationships between PCOS candidate genes during development of the fetal ovary, including TGFß and androgen signalling, we suggest that future studies should determine if perturbations of these genes in the fetal ovary can lead to PCOS in later life.


Assuntos
Biomarcadores/análise , Desenvolvimento Fetal/genética , Regulação da Expressão Gênica no Desenvolvimento , Ovário/patologia , Síndrome do Ovário Policístico/patologia , Polimorfismo de Nucleotídeo Único , Adulto , Animais , Bovinos , Feminino , Genes Reguladores , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Genótipo , Humanos , Ovário/metabolismo , Síndrome do Ovário Policístico/genética , Gravidez , Células Estromais/metabolismo , Células Estromais/patologia
12.
Appl Environ Microbiol ; 86(9)2020 04 17.
Artigo em Inglês | MEDLINE | ID: mdl-32086301

RESUMO

The four regulatory genes fscR1 to fscR4 in Streptomyces sp. strain FR-008 form a genetic arrangement that is widely distributed in macrolide-producing bacteria. Our previous work has demonstrated that fscR1 and fscR4 are critical for production of the polyene antibiotic candicidin. In this study, we further characterized the roles of the other two regulatory genes, fscR2 and fscR3, focusing on the relationship between these four regulatory genes. Disruption of a single or multiple regulatory genes did not affect bacterial growth, but transcription of genes in the candicidin biosynthetic gene cluster decreased, and candicidin production was abolished, indicating a critical role for each of the four regulatory genes, including fscR2 and fscR3, in candicidin biosynthesis. We found that fscR1 to fscR4, although differentially expressed throughout the growth phase, displayed similar temporal expression patterns, with an abrupt increase in the early exponential phase, coincident with initial detection of antibiotic production in the same phase. Our data suggest that the four regulatory genes fscR1 to fscR4 have various degrees of control over structural genes in the biosynthetic cluster under the conditions examined. Extensive transcriptional analysis indicated that complex regulation exists between these four regulatory genes, forming a regulatory network, with fscR1 and fscR4 functioning at a lower level. Comprehensive cross-complementation analysis indicates that functional complementation is restricted among the four regulators and unidirectional, with fscR1 complementing the loss of fscR3 or -4 and fscR4 complementing loss of fscR2 Our study provides more insights into the roles of, and the regulatory network formed by, these four regulatory genes controlling production of an important pharmaceutical compound.IMPORTANCE The regulation of antibiotic biosynthesis by Streptomyces species is complex, especially for biosynthetic gene clusters with multiple regulatory genes. The biosynthetic gene cluster for the polyene antibiotic candicidin contains four consecutive regulatory genes, which encode regulatory proteins from different families and which form a subcluster within the larger biosynthetic gene cluster in Streptomyces sp. FR-008. Syntenic arrangements of these regulatory genes are widely distributed in polyene gene clusters, such as the amphotericin and nystatin gene clusters, suggesting a conserved regulatory mechanism controlling production of these clinically important medicines. However, the relationships between these multiple regulatory genes are unknown. In this study, we determined that each of these four regulatory genes is critical for candicidin production. Additionally, using transcriptional analyses, bioassays, high-performance liquid chromatography (HPLC) analysis, and genetic cross-complementation, we showed that FscR1 to FscR4 comprise a hierarchical regulatory network that controls candicidin production and is likely representative of how expression of other polyene biosynthetic gene clusters is controlled.


Assuntos
Antibacterianos/biossíntese , Proteínas de Bactérias/metabolismo , Candicidina/biossíntese , Regulação Bacteriana da Expressão Gênica , Streptomyces/metabolismo , Fatores de Transcrição/metabolismo , Proteínas de Bactérias/genética , Diterpenos , Genes Bacterianos , Genes Reguladores , Streptomyces/genética , Fatores de Transcrição/genética
13.
Genes (Basel) ; 11(1)2020 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-31936116

RESUMO

Heat stress affects the physiology and production performance of Chinese Holstein dairy cows. As such, the selection of heat tolerance in cows and elucidating its underlying mechanisms are vital to the dairy industry. This study aimed to investigate the heat tolerance associated genes and molecular mechanisms in Chinese Holstein dairy cows using a high-throughput sequencing approach and bioinformatics analysis. Heat-induced physiological indicators and milk yield changes were assessed to determine heat tolerance levels in Chinese Holstein dairy cows by Principal Component Analysis method following Membership Function Value Analysis. Results indicated that rectal temperature (RT), respiratory rate (RR), and decline in milk production were significantly lower (p < 0.05) in heat tolerant (HT) cows while plasma levels of heat shock protein (HSP: HSP70, HSP90), and cortisol were significantly higher (p < 0.05) when compared to non-heat tolerant (NHT) Chinese Holstein dairy cows. By applying RNA-Seq analysis, we identified 200 (81 down-regulated and 119 up-regulated) significantly (|log2fold change| ≥ 1.4 and p ≤ 0.05) differentially expressed genes (DEGs) in HT versus NHT Chinese Holstein dairy cows. In addition, 14 of which were involved in protein-protein interaction (PPI) network. Importantly, several hub genes (OAS2, MX2, IFIT5 and TGFB2) were significantly enriched in immune effector process. These findings might be helpful to expedite the understanding for the mechanism of heat tolerance in Chinese Holstein dairy cows.


Assuntos
Bovinos/genética , Genes Reguladores/genética , Termotolerância/genética , Animais , China , Indústria de Laticínios/métodos , Perfilação da Expressão Gênica/métodos , Transtornos de Estresse por Calor/genética , Proteínas de Choque Térmico , Resposta ao Choque Térmico/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Temperatura Alta , Transcriptoma/genética
14.
BMC Genomics ; 21(1): 42, 2020 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-31931708

RESUMO

BACKGROUND: Rockweeds are among the most important foundation species of temperate rocky littoral shores. In the Baltic Sea, the rockweed Fucus vesiculosus is distributed along a decreasing salinity gradient from the North Atlantic entrance to the low-salinity regions in the north-eastern margins, thus, demonstrating a remarkable tolerance to hyposalinity. The underlying mechanisms for this tolerance are still poorly understood. Here, we exposed F. vesiculosus from two range-margin populations to the hyposaline (2.5 PSU - practical salinity unit) conditions that are projected to occur in the region by the end of this century as a result of climate change. We used transcriptome analysis (RNA-seq) to determine the gene expression patterns associated with hyposalinity acclimation, and examined the variation in these patterns between the sampled populations. RESULTS: Hyposalinity induced different responses in the two populations: in one, only 26 genes were differentially expressed between salinity treatments, while the other population demonstrated up- or downregulation in 3072 genes. In the latter population, the projected future hyposalinity induced an acute response in terms of antioxidant production. Genes associated with membrane composition and structure were also heavily involved, with the upregulation of fatty acid and actin production, and the downregulation of ion channels and alginate pathways. Changes in gene expression patterns clearly indicated an inhibition of the photosynthetic machinery, with a consequent downregulation of carbohydrate production. Simultaneously, energy consumption increased, as revealed by the upregulation of genes associated with respiration and ATP synthesis. Overall, the genes that demonstrated the largest increase in expression were ribosomal proteins involved in translation pathways. The fixation rate of SNP:s was higher within genes responding to hyposalinity than elsewhere in the transcriptome. CONCLUSIONS: The high fixation rate in the genes coding for salinity acclimation mechanisms implies strong selection for them. The among-population differentiation that we observed in the transcriptomic response to hyposalinity stress suggests that populations of F. vesiculosus may differ in their tolerance to future desalination, possibly as a result of local adaptation to salinity conditions within the Baltic Sea. These results emphasise the importance of considering interspecific genetic variation when evaluating the consequences of environmental change.


Assuntos
Adaptação Biológica/genética , Fucus/fisiologia , Regulação da Expressão Gênica , Genes Reguladores , Salinidade , Alga Marinha/fisiologia , Biologia Computacional/métodos , Perfilação da Expressão Gênica , Ontologia Genética , Variação Genética , Anotação de Sequência Molecular , Transcriptoma
15.
Nat Med ; 26(2): 200-206, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31988463

RESUMO

Chronic granulomatous disease (CGD) is a rare inherited disorder of phagocytic cells1,2. We report the initial results of nine severely affected X-linked CGD (X-CGD) patients who received ex vivo autologous CD34+ hematopoietic stem and progenitor cell-based lentiviral gene therapy following myeloablative conditioning in first-in-human studies (trial registry nos. NCT02234934 and NCT01855685). The primary objectives were to assess the safety and evaluate the efficacy and stability of biochemical and functional reconstitution in the progeny of engrafted cells at 12 months. The secondary objectives included the evaluation of augmented immunity against bacterial and fungal infection, as well as assessment of hematopoietic stem cell transduction and engraftment. Two enrolled patients died within 3 months of treatment from pre-existing comorbidities. At 12 months, six of the seven surviving patients demonstrated stable vector copy numbers (0.4-1.8 copies per neutrophil) and the persistence of 16-46% oxidase-positive neutrophils. There was no molecular evidence of either clonal dysregulation or transgene silencing. Surviving patients have had no new CGD-related infections, and six have been able to discontinue CGD-related antibiotic prophylaxis. The primary objective was met in six of the nine patients at 12 months follow-up, suggesting that autologous gene therapy is a promising approach for CGD patients.


Assuntos
Cromossomos Humanos X , Terapia Genética/métodos , Doença Granulomatosa Crônica/genética , Lentivirus/genética , Adolescente , Antígenos CD34/genética , Criança , Pré-Escolar , Comorbidade , Inativação Gênica , Genes Reguladores , Vetores Genéticos , Doença Granulomatosa Crônica/terapia , Células-Tronco Hematopoéticas/citologia , Humanos , Masculino , NADPH Oxidases/genética , Neutrófilos/metabolismo , Segurança do Paciente , Regiões Promotoras Genéticas , Condicionamento Pré-Transplante , Resultado do Tratamento , Reino Unido , Estados Unidos , Adulto Jovem
16.
mBio ; 10(6)2019 11 12.
Artigo em Inglês | MEDLINE | ID: mdl-31719176

RESUMO

Transcription of bacterial genes is controlled by the coordinated action of cis- and trans-acting regulators. The activity and mode of action of these regulators can reflect different requirements for gene products in different environments. A well-studied example is the regulatory function that integrates the environmental availability of glucose and lactose to control the Escherichia coli lac operon. Most studies of lac operon regulation have focused on a few closely related strains. To determine the range of natural variation in lac regulatory function, we introduced a reporter construct into 23 diverse E. coli strains and measured expression with combinations of inducer concentrations. We found a wide range of regulatory functions. Several functions were similar to the one observed in a reference lab strain, whereas others depended weakly on the presence of cAMP. Some characteristics of the regulatory function were explained by the genetic relatedness of strains, indicating that differences varied on relatively short time scales. The regulatory characteristics explained by genetic relatedness were among those that best predicted the initial growth of strains following transition to a lactose environment, suggesting a role for selection. Finally, we transferred the lac operon, with the lacI regulatory gene, from five natural isolate strains into a reference lab strain. The regulatory function of these hybrid strains revealed the effect of local and global regulatory elements in controlling expression. Together, this work demonstrates that regulatory functions can be varied within a species and that there is variation within a species to best match a function to particular environments.IMPORTANCE The lac operon of Escherichia coli is a classic model for studying gene regulation. This study has uncovered features such as the environmental input logic controlling gene expression, as well as gene expression bistability and hysteresis. Most lac operon studies have focused on a few lab strains, and it is not known how generally those findings apply to the diversity of E. coli strains. We examined the environmental dependence of lac gene regulation in 20 natural isolates of E. coli and found a wide range of regulatory responses. By transferring lac genes from natural isolate strains into a common reference strain, we found that regulation depends on both the lac genes themselves and on the broader genetic background, indicating potential for still-greater regulatory diversity following horizontal gene transfer. Our results reveal that there is substantial natural variation in the regulation of the lac operon and indicate that this variation can be ecologically meaningful.


Assuntos
Escherichia coli/classificação , Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica , Patrimônio Genético , Variação Genética , Óperon Lac , Escherichia coli/isolamento & purificação , Evolução Molecular , Genes Bacterianos , Genes Reguladores , Mutação , Fenótipo , Filogenia , Polimorfismo Genético
17.
BMC Genomics ; 20(1): 767, 2019 Oct 22.
Artigo em Inglês | MEDLINE | ID: mdl-31640553

RESUMO

BACKGROUND: Escherichia coli C forms more robust biofilms than other laboratory strains. Biofilm formation and cell aggregation under a high shear force depend on temperature and salt concentrations. It is the last of five E. coli strains (C, K12, B, W, Crooks) designated as safe for laboratory purposes whose genome has not been sequenced. RESULTS: Here we present the complete genomic sequence of this strain in which we utilized both long-read PacBio-based sequencing and high resolution optical mapping to confirm a large inversion in comparison to the other laboratory strains. Notably, DNA sequence comparison revealed the absence of several genes thought to be involved in biofilm formation, including antigen 43, waaSBOJYZUL for lipopolysaccharide (LPS) synthesis, and cpsB for curli synthesis. The first main difference we identified that likely affects biofilm formation is the presence of an IS3-like insertion sequence in front of the carbon storage regulator csrA gene. This insertion is located 86 bp upstream of the csrA start codon inside the - 35 region of P4 promoter and blocks the transcription from the sigma32 and sigma70 promoters P1-P3 located further upstream. The second is the presence of an IS5/IS1182 in front of the csgD gene. And finally, E. coli C encodes an additional sigma70 subunit driven by the same IS3-like insertion sequence. Promoter analyses using GFP gene fusions provided insights into understanding this regulatory pathway in E. coli. CONCLUSIONS: Biofilms are crucial for bacterial survival, adaptation, and dissemination in natural, industrial, and medical environments. Most laboratory strains of E. coli grown for decades in vitro have evolved and lost their ability to form biofilm, while environmental isolates that can cause infections and diseases are not safe to work with. Here, we show that the historic laboratory strain of E. coli C produces a robust biofilm and can be used as a model organism for multicellular bacterial research. Furthermore, we ascertained the full genomic sequence of this classic strain, which provides for a base level of characterization and makes it useful for many biofilm-based applications.


Assuntos
Biofilmes/crescimento & desenvolvimento , Escherichia coli/genética , Genoma Bacteriano/genética , Aderência Bacteriana/genética , Mapeamento Cromossômico , Escherichia coli/crescimento & desenvolvimento , Proteínas de Escherichia coli/genética , Regulação Bacteriana da Expressão Gênica , Genes Reguladores/genética , Regiões Promotoras Genéticas , Estresse Salino/genética , Inversão de Sequência , Temperatura , Fatores de Transcrição/genética
18.
Environ Microbiol Rep ; 11(6): 784-796, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31621205

RESUMO

Amino acids are vital components in cell metabolism. Leucine is a regulatory factor that generates significant impact on protein synthesis/turnover, modulates diverse cellular signalling pathways and participates in oxidative processes and immune responses. Here, we identified and characterized the functions of a leucine-associated Zn2 Cys6 -type transcription factor, MoLeu3. Disruption of MoLEU3 resulted in significantly reduced pathogenicity in barley and rice. Quantitative RT-PCR showed that the expression levels of the putative leucine biosynthesis-related genes, MoLEU1, MoLEU2 and MoLEU4 were downregulated in the ΔMoleu3 mutant. We used high-throughput gene knockout method to generate the null mutants of MoLEU1, MoLEU2 and MoLEU4 respectively. The ΔMoleu1, ΔMoleu2 and ΔMoleu4 mutants are leucine auxotroph and showed similar phenotypic characterizations, including reduced conidiation, delayed mobilization and degradation of glycogen and lipid droplets, limited appressorium-mediated penetration, and restricted invasive hyphae growth within host cells. Collectively, MoLEU1, MoLEU2, and MoLEU4 regulated by MoLEU3 play crucial roles in fungal development and infectious processes through modulation of leucine biosynthesis in Magnaporthe oryzae.


Assuntos
Proteínas Fúngicas/genética , Genes Reguladores , Leucina/biossíntese , Magnaporthe/crescimento & desenvolvimento , Magnaporthe/metabolismo , Redes e Vias Metabólicas/genética , Fatores de Virulência/metabolismo , Proteínas Fúngicas/metabolismo , Deleção de Genes , Regulação Fúngica da Expressão Gênica , Hordeum , Magnaporthe/genética , Magnaporthe/patogenicidade , Oryza , Doenças das Plantas/microbiologia , Virulência
19.
Elife ; 82019 09 23.
Artigo em Inglês | MEDLINE | ID: mdl-31545165

RESUMO

Enhancers are the primary DNA regulatory elements that confer cell type specificity of gene expression. Recent studies characterizing individual enhancers have revealed their potential to direct heterologous gene expression in a highly cell-type-specific manner. However, it has not yet been possible to systematically identify and test the function of enhancers for each of the many cell types in an organism. We have developed PESCA, a scalable and generalizable method that leverages ATAC- and single-cell RNA-sequencing protocols, to characterize cell-type-specific enhancers that should enable genetic access and perturbation of gene function across mammalian cell types. Focusing on the highly heterogeneous mammalian cerebral cortex, we apply PESCA to find enhancers and generate viral reagents capable of accessing and manipulating a subset of somatostatin-expressing cortical interneurons with high specificity. This study demonstrates the utility of this platform for developing new cell-type-specific viral reagents, with significant implications for both basic and translational research.


Assuntos
Regulação da Expressão Gênica/efeitos dos fármacos , Biologia Molecular/métodos , Neurônios/efeitos dos fármacos , Neurofisiologia/métodos , Proteínas Recombinantes/biossíntese , Somatostatina/metabolismo , Vírus/genética , Animais , Animais Geneticamente Modificados , Córtex Cerebral/fisiologia , Genes Reguladores , Vetores Genéticos , Interneurônios/fisiologia , Camundongos , Proteínas Recombinantes/genética
20.
BMC Plant Biol ; 19(1): 356, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31416414

RESUMO

BACKGROUND: Drought is a major environmental constraint to plant growth, development and productivity. Compared with most willows that are generally susceptible to drought, the desert willow Salix psammophila has extraordinary adaptation to drought stress. However, its molecular basis of drought tolerance is still largely unknown. RESULTS: During polyethylene glycol 6000 (PEG 6000)-simulated drought stress, we found that the osmotic adjustment substances were accumulated and the antioxidant enzyme activities were enhanced in S. psammophila roots. A total of 8172 differentially expressed genes were identified in roots of S. psammophila through RNA-Sequencing. Based on K-means clustering, their expression patterns were classified into nine clusters, which were enriched in several stress-related processes including transcriptional regulation, response to various stresses, cell death, etc. Moreover, 672 transcription factors from 45 gene families were differentially expressed under drought stress. Furthermore, a weighted gene co-expression network was constructed, and eight genes were identified as hub genes. We demonstrated the function of two hub genes, magnesium-dependent phosphatase 1 (SpMDP1) and SpWRKY33, through overexpression in Arabidopsis thaliana. Overexpression of the two hub genes enhanced the drought tolerance in transgenic plants, suggesting that the identification of candidate drought tolerance genes in this study was highly efficient and credible. CONCLUSIONS: Our study analyzed the physiological and molecular responses to drought stress in S. psammophila, and these results contribute to dissect the mechanism of drought tolerance of S. psammophila and facilitate identification of critical genes involved in drought tolerance for willow breeding.


Assuntos
Secas , Regulação da Expressão Gênica de Plantas/fisiologia , Genoma de Planta/fisiologia , Proteínas de Plantas/genética , Salix/fisiologia , Fatores de Transcrição/genética , Transcriptoma/fisiologia , Arabidopsis/genética , Arabidopsis/fisiologia , Genes Reguladores/fisiologia , Proteínas de Plantas/metabolismo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/fisiologia , Salix/genética , Fatores de Transcrição/metabolismo
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