Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

High-resolution transcriptome and genome-wide dynamics of RNA polymerase and NusA in Mycobacterium tuberculosis (RNA-seq)

ABSTRACT: 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

ORGANISM(S): Mycobacterium tuberculosis

SUBMITTER: Swapna Uplekar

PROVIDER: E-GEOD-40846 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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High-resolution transcriptome and genome-wide dynamics of RNA polymerase and NusA in Mycobacterium tuberculosis.

Uplekar Swapna S Rougemont Jacques J Cole Stewart T ST Sala Claudia C

Nucleic acids research 20121207 2

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, to survey the global transcriptome, and ChIP-seq, to monitor the genome-wide dynamics of RNA polymerase (RNAP) and the anti-terminator NusA. Although NusA does not bind directly to DNA, but rather to RNAP and/or to the nascent transcript, we demonstrate that NusA interacts with RNAP ubiquitously throughout the chromosome, ...[more]

PMID: 23222129

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Project description:The PhoPR two-component system is essential for virulence in Mycobacterium tuberculosis where it controls expression of approximately 2% of the genes, including those for the ESX-1 secretion apparatus, a major virulence determinant. Mutations in phoP lead to compromised production of pathogen-specific cell wall components and attenuation both ex vivo and in vivo. Using antibodies against the native protein in ChIP-seq experiments (chromatin immunoprecipitation followed by high-throughput sequencing) we demonstrated that PhoP binds to at least 35 loci on the M. tuberculosis genome. The PhoP regulon comprises several transcriptional regulators as well as genes for polyketide synthases and PE/PPE proteins. Integration of ChIP-seq results with high-resolution transcriptomic analysis (RNA-seq) revealed that PhoP controls 30 genes directly, whilst regulatory cascades are responsible for signal amplification and downstream effects. The most prominent site of PhoP regulation was located in the intergenic region between rv2395 and PE_PGRS41, where the mcr7 gene codes for a small non-coding RNA (ncRNA). Northern blot experiments confirmed the absence of Mcr7 in the M. tuberculosis phoP mutant as well as low-level expression of the ncRNA in M. tuberculosis complex members other than M. tuberculosis. By means of genetic and proteomic analyses we demonstrated that Mcr7 modulates translation of the tatC mRNA thereby impacting the activity of the Twin Arginine Translocation (Tat) protein secretion apparatus. As a result, secretion of the immunodominant Ag85 complex and the beta-lactamase BlaC is affected, among others. Mcr7, the first ncRNA of M. tuberculosis whose function has been established, therefore represents a missing link between the PhoPR two-component system and the downstream functions necessary for successful infection of the host. RNA-Seq: M. tuberculosis H37Rv (GC1237) cultures grown in vitro to exponential phase. One wild type sample plus one isogenic phoP mutant sample

Dataset Information

High-resolution transcriptome and genome-wide dynamics of RNA polymerase and NusA in Mycobacterium tuberculosis (RNA-seq)

Publications

High-resolution transcriptome and genome-wide dynamics of RNA polymerase and NusA in Mycobacterium tuberculosis.

Similar Datasets

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