Mapping of genotype-phenotype diversity amongst clinical isolates of Mycobacterium tuberculosis by RNA-seq
Ontology highlight
ABSTRACT:
ORGANISM(S): Mycobacterium tuberculosis
SUBMITTER: Graham Rose
PROVIDER: E-MTAB-1446 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:To construct a regulatory map of the genome of the human pathogen, Mycobacterium tuberculosis, we applied two complementary high-resolution approaches, strand-specific RNA-seq and ChIP-seq, to survey the global transcriptome and monitor genome-wide dynamics of RNA polymerase (RNAP) and NusA. ChIP-seq revealed that RNAP and the antiterminator, NusA, occurred ubiquitously with the NusA profile mirroring RNAP distribution in both the exponential and stationary phases of growth. Generally, promoter-proximal peaks for RNAP and NusA were observed, followed by a decrease in signal strength reflecting transcriptional polarity. Differential binding of RNAP and NusA in the two growth conditions correlated with transcriptional activity as reflected by RNA abundance. Indeed, a significant association between expression levels and the presence of NusA throughout the gene body was detected, confirming the peculiar transcription-promoting role of NusA. Integration of the datasets pinpointed transcriptional units, mapped promoters and uncovered new anti-sense and non-coding transcripts. Highly expressed transcriptional units were situated mainly on the leading strand, despite the relatively unbiased distribution of genes throughout the genome, thus helping the replicative and transcriptional complexes to align. ChIP-Seq of RNAP and NusA in Mtb H37Rv at exponential and stationary phase cultures. Each experiment was performed in duplicate. Input DNA (No IP) was used as a control.
2012-12-01 | E-GEOD-40845 | biostudies-arrayexpress
Project description:This SuperSeries is composed of the following subset Series: GSE11873: Bacillus subtilis 168 cells: wild-type vs. ccpN (non polar) GSE11876: Bacillus subtilis 168 cells: wild-type vs. (ccpN-yqfL) double mutant Refer to individual Series
2008-07-01 | E-GEOD-11937 | biostudies-arrayexpress
Project description:Global transcriptional profiling of Bacillus subtilis cells comparing wild-type to a ccpN (yqzB) non polar mutant. Abstract of associated publication (article accepted): The transcriptional regulator CcpN of Bacillus subtilis has been recently characterized as a repressor of two gluconeogenic genes, gapB and pckA, and of a small non-coding regulatory RNA, sr1, involved in arginine catabolism. Deletion of ccpN impairs growth on glucose and strongly alters the distribution of intracellular fluxes, rerouting the main glucose catabolism from glycolysis to the pentose phosphate (PP) pathway. Using transcriptome analysis, we show that during growth on glucose, gapB and pckA are the only protein-coding genes directly repressed by CcpN. By quantifying intracellular fluxes in deletion mutants, we demonstrate that derepression of pckA under glycolytic condition causes the growth defect observed in the ccpN mutant due to extensive futile cycling through the pyruvate carboxylase, phosphoenolpyruvate carboxykinase, and pyruvate kinase. Beyond ATP dissipation via this cycle, PckA activity causes a drain on tricarboxylic acid cycle intermediates, which we show to be the main reason for the reduced growth of a ccpN mutant. The high flux through the PP pathway in the ccpN mutant is modulated by the flux through the alternative glyceraldehyde-3-phosphate dehydrogenases, GapA and GapB. Strongly increased concentrations of intermediates in upper glycolysis indicate that GapB overexpression causes a metabolic jamming of this pathway and, consequently, increases the relative flux through the PP pathway. In contrast, derepression of sr1, the third known target of CcpN, plays only a marginal role in ccpN mutant phenotypes. Two-condition experiment, wt vs. ccpN mutant. 4 experiments were performed different days with 3 independent RNA preparations (from separate cultures) and 3 independent sets of macroarrays: Experiment 1 (samples wt1 and ccpN1): total RNA extract1 / membranes set A Expereince 2 (samples wt2 and ccpN2): total RNA extract 2 / membranes set A Experiment 3 (samples wt2 and ccpN2): total RNA extract 3 / membranes set B Experiment 4 (samples wt2 and ccpN2): total RNA extract 3 / membranes set C
2008-06-26 | E-GEOD-11873 | biostudies-arrayexpress
Project description:Global transcriptional profiling of Bacillus subtilis cells comparing wild-type to a (ccpN-yqfL) double mutant. Abstract of the associated publication (article accepted): The transcriptional regulator CcpN of Bacillus subtilis has been recently characterized as a repressor of two gluconeogenic genes, gapB and pckA, and of a small non-coding regulatory RNA, sr1, involved in arginine catabolism. Deletion of ccpN impairs growth on glucose and strongly alters the distribution of intracellular fluxes, rerouting the main glucose catabolism from glycolysis to the pentose phosphate (PP) pathway. Using transcriptome analysis, we show that during growth on glucose, gapB and pckA are the only protein-coding genes directly repressed by CcpN. By quantifying intracellular fluxes in deletion mutants, we demonstrate that derepression of pckA under glycolytic condition causes the growth defect observed in the ccpN mutant due to extensive futile cycling through the pyruvate carboxylase, phosphoenolpyruvate carboxykinase, and pyruvate kinase. Beyond ATP dissipation via this cycle, PckA activity causes a drain on tricarboxylic acid cycle intermediates, which we show to be the main reason for the reduced growth of a ccpN mutant. The high flux through the PP pathway in the ccpN mutant is modulated by the flux through the alternative glyceraldehyde-3-phosphate dehydrogenases, GapA and GapB. Strongly increased concentrations of intermediates in upper glycolysis indicate that GapB overexpression causes a metabolic jamming of this pathway and, consequently, increases the relative flux through the PP pathway. In contrast, derepression of sr1, the third known target of CcpN, plays only a marginal role in ccpN mutant phenotypes. Two-condition experiment: wt vs. (ccpN-yqfL) double mutant. 2 totally independent comparisons were performed different days with 2 independent RNA preparations and 2 sets of macroarrays.
2008-07-01 | E-GEOD-11876 | biostudies-arrayexpress
Project description:New tools for studying bacterial transcripts at the single nucleotide level offer an unparalleled opportunity to understand the bacterial transcriptome. For the model pathogen Salmonella enterica serovar Typhimurium, it is necessary to identify the regulatory inputs for all RNA transcripts, including small RNAs (sRNAs) and coding genes. Here, we use RNA-seq to define the transcriptomes of mutants lacking 18 global regulatory systems that, among other functions, modulate the expression of the SPI1 and SPI2 Type Three secretion systems in S. Typhimurium strain 4/74. We directly compared the roles of the major regulators of transcription, and reported the effects of the regulatory mutations on expression of sRNAs. We also use this method to describe the impact of the RNA chaperone Hfq upon the steady state levels of 280 sRNA transcripts. Transcriptome analysis of S. Typhimurium 4/74 using RNA isolated wild-type and mutants grown under infection-relevant conditions.
2016-08-09 | E-GEOD-79314 | biostudies-arrayexpress
Project description:To construct a regulatory map of the genome of the human pathogen, Mycobacterium tuberculosis, we applied two complementary high-resolution approaches, strand-specific RNA-seq and ChIP-seq, to survey the global transcriptome and monitor genome-wide dynamics of RNA polymerase (RNAP) and NusA. ChIP-seq revealed that RNAP and the antiterminator, NusA, occurred ubiquitously with the NusA profile mirroring RNAP distribution in both the exponential and stationary phases of growth. Generally, promoter-proximal peaks for RNAP and NusA were observed, followed by a decrease in signal strength reflecting transcriptional polarity. Differential binding of RNAP and NusA in the two growth conditions correlated with transcriptional activity as reflected by RNA abundance. Indeed, a significant association between expression levels and the presence of NusA throughout the gene body was detected, confirming the peculiar transcription-promoting role of NusA. Integration of the datasets pinpointed transcriptional units, mapped promoters and uncovered new anti-sense and non-coding transcripts. Highly expressed transcriptional units were situated mainly on the leading strand, despite the relatively unbiased distribution of genes throughout the genome, thus helping the replicative and transcriptional complexes to align. RNA-Seq in exponential and stationary phase cultures
2012-12-01 | E-GEOD-40846 | biostudies-arrayexpress
Project description:In a previous study we adopted an integrated transcriptomic and proteomic approach to determine the physiological response of E. coli O157:H7 Sakai during exponential phase growth under steady-state conditions relevant to low temperature and water activity conditions experienced during meat carcass chilling in cold air (Kocharunchitt et al., 2012). The findings of that study provide a baseline of knowledge of the physiology of this pathogen, with the response of E. coli O157:H7 to steady-state conditions of combined cold and osmotic stress. To provide an insight into the genetic systems enabling this organism to adapt to growth at low temperature, we extended the aforementioned study to investigate the growth kinetics of E. coli O157:H7 Sakai during abrupt temperature downshift from 35 degrees C to 14 degrees C and, examined time-dependent global alterations in its genome expression upon cold shock from 35 degrees C to 14 degrees C. The genome-wide expression response of E. coli was analysed by both cDNA microarray (transcriptome response) and 2D-LC/MS/MS analysis (proteome response). Differences in gene and protein expression patterns in E. coli before and after cold shock were analysed through quantitative and comparative analysis of time series changes in both mRNA and proteins levels.
2014-10-18 | E-MTAB-2010 | biostudies-arrayexpress
Project description:In this study transcriptional start sites (TSS) for the phytopathogen Dickeya dadantii 3937 were determined based on differential RNA-seq (dRNA-seq) as previously described (Sharma and Vogel, 2014). RNAs from E-MTAB-541 (Jiang et al. 2015) were pooled into four samples S1 to S4 reflecting a variety of environmental conditions. Each of them resulted in a combination of stress conditions (pH, NaCl, H2O2) and growth conditions: exponential phase with stress (S1), exponential phase without stress (S2), stationary phase with stress (S3), stationary phase without stress (S4). Those samples were then supplied to Vertis Biotechnologie AG for TEX treatment and Illumina sequencing. Briefly, ribosomal RNA molecules were depleted from the total RNA samples using the Ribo-Zero rRNA Removal Kit for bacteria (Epicentre). From the rRNA depleted RNA samples, the small RNA fractions <200 nt were separated from the large fractions using the RNeasy MinElute Cleanup Kit (Qiagen). TSS cDNA libraries were generated from the rRNA depleted large RNA fractions. First, the samples were fragmented using RNase III. Samples were then poly(A)-tailed using poly(A) polymerase, and split into two halves. One half was treated with Terminator exonuclease (+TEX, Epicentre), while the other one was left untreated (-TEX). 5'PPP structures were then converted into 5'P ends using RNA 5' Polyphosphatase (5'PP, Epicentre). Afterwards, an RNA adapter was ligated to the 5'P of the RNA. First-strand cDNA synthesis was performed using an oligo(dT)-adapter primer and the M-MLV reverse transcriptase. The resulting cDNAs were PCR-amplified using a high fidelity DNA polymerase, and purified using the Agencourt AMPure XP kit (Beckman Coulter Genomics). The cDNA pools were then sequenced on an Illumina NextSeq 500 system using 60 bp read length and a single-end, strand-specific protocol. Sequencing reads were trimmed, and both polyA and adapters removed.
2022-01-01 | E-MTAB-9075 | biostudies-arrayexpress
Project description:The M-NM-1-proteobacterium Sinorhizobium fredii NGR234 has an exceptionally wide host range as it forms nitrogen-fixing nodules with more legumes than any other known microsymbiont. Within its 6.9 Mbp genome it encodes two N-acyl-homoserine-lactone synthase genes (i.e. traI and ngrI) involved in the biosynthesis of two distinct autoinducer I-type molecules. Here we report on the construction of a NGR234-M-NM-^TtraI and a NGR234-M-NM-^TngrI mutant and the genome wide RNA-seq-based transcriptome analysis in the parent strain and in the two constructed mutants. A high-resolution RNA-seq analysis of early stationary phase cultures in the background of NGR234-M-NM-^TtraI suggested that up to 316 genes (4.9% of all predicted genes) were differentially expressed in the mutant vs. the parent strain. Similarly, in the background of NGR234-M-NM-^TngrI 466 differentially regulated genes (7.3% of all predicted genes) were identified. A considerable overlap in the gene expression pattern was uncovered between both mutants resulting in a common set of 186 genes which were almost identically regulated. Co-regulated genes that were linked to the ngrI- and the traI-dependent autoinducer regulatory circuits included 53 flagella biosynthesis genes and genes linked to EPS succinoglycan biosynthesis. Among the genes and ORFs that were differentially regulated in NGR234-M-NM-^TtraI were those linked to replication of the pNGR234a symbiotic plasmid and cytochrome c-related genes. In the NGR234-M-NM-^TngrI mutant biotin and pyrroloquinoline quinone (PQQ) biosynthesis genes were differentially expressed as well as the entire cluster of 21 genes linked to assembly of the NGR234 type three secretion system II (T3SS-II). We also discovered that genes responsible for octopine catabolism in NGR234 are strongly repressed in the presence of high levels of N-acyl-homoserine-lactones (AHLs). Surprisingly, only few truly symbiosis-related genes were identified that appeared to be quorum sensing regulated. Together with nodulation assays, the RNAseq-based findings suggested that QS-dependent gene regulation appears to be of higher relevance during non-symbiotic growth rather than for life within root nodules. In total 12 samples were analyzed (six different treatments with two independent samples), treatment A: NGR234 wild type strain in stationary phase, treatment B: NGR234 traI-mutant strain in stationary phase, treatment C: NGR234 ngrI-mutant strain in stationary phase, treatment D: NGR234 wild type strain in exponential phase, treatment E: NGR234 wild type supplemented with 0.05 M-BM-5M AHLs in exponential phase, treatment F: NGR234 wild type supplemented with 50 M-BM-5M AHLs in exponential phase
2014-07-18 | E-GEOD-54381 | biostudies-arrayexpress
Project description:In a previous study we adopted an integrated transcriptomic and proteomic approach to determine the physiological response of E. coli O157:H7 Sakai during exponential phase growth under steady-state conditions relevant to low temperature and water activity conditions experienced during meat carcass chilling in cold air (Kocharunchitt et al., 2012). The findings of that study provide a baseline of knowledge of the physiology of this pathogen, with the response of E. coli O157:H7 to steady-state conditions of cold and osmotic stress. To provide an insight into the genetic systems enabling this organism to adapt to growth at low water activity, we extended the aforementioned study to investigate the growth kinetics of E. coli O157:H7 Sakai during abrupt water activity downshift from 0.993 to 0.967 and, examined time-dependent global alterations in its genome expression upon water activity downshift from 0.993 to 0.967. The genome-wide expression response of E. coli was analysed by both cDNA microarray (transcriptome response) and 2D-LC/MS/MS analysis (proteome response). Differences in gene and protein expression patterns in E. coli before and after water activity downshift were analysed through quantitative and comparative analysis of time series changes in both mRNA and proteins levels.
2014-10-18 | E-MTAB-2011 | biostudies-arrayexpress