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Rho directs widespread termination of intragenic and stable RNA transcription

ABSTRACT: The transcription termination factor Rho is a global regulator of RNA polymerase (RNAP). Although individual Rho-dependent terminators have been studied extensively, less is known about the sites of RNAP regulation by Rho on a genome-wide scale. Using chromatin immunoprecipitation and microarrays (ChIP-chip), we examined changes in the distribution of Escherichia coli RNAP in response to the Rho-specific inhibitor bicyclomycin (BCM). We found ~200 Rho-terminated loci that were divided evenly into two classes: intergenic (at the ends of genes) and intragenic (within genes). The intergenic class contained noncoding RNAs such as small RNAs (sRNAs) and transfer RNAs (tRNAs), establishing a previously unappreciated role of Rho in termination of stable RNA synthesis. The intragenic class of terminators included a novel set of short antisense transcripts, as judged by a shift in the distribution of RNAP in BCM-treated cells that was opposite to the direction of the corresponding gene. These Rho-terminated antisense transcripts point to a novel role of noncoding transcription in E. coli gene regulation that may resemble the ubiquitous noncoding transcription recently found to play myriad roles in eukaryotic gene regulation.

ORGANISM(S): Escherichia coli Escherichia coli str. K-12 substr. MG1655

PROVIDER: GSE16562 | GEO | 2009/06/23

SECONDARY ACCESSION(S): PRJNA116199

REPOSITORIES: GEO

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Rho directs widespread termination of intragenic and stable RNA transcription

Project description:The transcription termination factor Rho is a global regulator of RNA polymerase (RNAP). Although individual Rho-dependent terminators have been studied extensively, less is known about the sites of RNAP regulation by Rho on a genome-wide scale. Using chromatin immunoprecipitation and microarrays (ChIP-chip), we examined changes in the distribution of Escherichia coli RNAP in response to the Rho-specific inhibitor bicyclomycin (BCM). We found ~200 Rho-terminated loci that were divided evenly into two classes: intergenic (at the ends of genes) and intragenic (within genes). The intergenic class contained noncoding RNAs such as small RNAs (sRNAs) and transfer RNAs (tRNAs), establishing a previously unappreciated role of Rho in termination of stable RNA synthesis. The intragenic class of terminators included a novel set of short antisense transcripts, as judged by a shift in the distribution of RNAP in BCM-treated cells that was opposite to the direction of the corresponding gene. These Rho-terminated antisense transcripts point to a novel role of noncoding transcription in E. coli gene regulation that may resemble the ubiquitous noncoding transcription recently found to play myriad roles in eukaryotic gene regulation. Chromatin immunoprecipitation (ChIP) experiments were performed using antibodies against RNA polymerase (Beta or Beta' subunit) in cells treated with 20ug/ml bicyclomycin or untreated cells. Differentially labeled ChIP DNA and genomic DNA were competitively hybridized to an E. coli K-12 MG1655 tiling array with overlapping probes at ~12bp spacing across the entire genome. The series contains 4 datasets.

2010-05-18 | E-GEOD-16562 | biostudies-arrayexpress

Rho and NusG suppress pervasive antisense transcription in Escherichia coli [ChIP-chip].

Project description:Despite the prevalence of antisense transcripts in bacterial transcriptomes, little is known about how their synthesis is controlled. We report that a major function of the Escherichia coli termination factor Rho and its co-factor NusG is suppression of ubiquitous antisense transcription genome-wide. Rho binds C-rich unstructured nascent RNA (high C/G ratio) prior to its ATP-dependent dissociation of transcription complexes. NusG is required for efficient termination at minority subsets (~20%) of both antisense and sense Rho-dependent terminators with lower C/G ratio sequences. In contrast, a widely studied nusA deletion proposed to compromise Rho-dependent termination had no effect on antisense or sense Rho-dependent terminators in vivo. Global co-localization of the nucleoid-associated protein H-NS with Rho-dependent terminators and genetic interactions between hns and rho suggest that H-NS aids Rho in suppression of antisense transcription. The combined actions of Rho, NusG, and H-NS appear to be analogous to the Sen1-Nrd1-Nab3 and nucleosome systems that suppress antisense transcription in eukaryotes. Chromatin immunoprecipitation (ChIP) experiments were performed using antibodies against RNA polymerase (RNAP; Beta subunit) in wild-type cells or cells deleted for hns, nusG, or a partial deletion of nusA. Differentially labeled ChIP DNA and genomic DNA were competitively hybridized to an E. coli K-12 MG1655 tiling array with overlapping probes at ~12bp spacing across the entire genome. The series contains 12 datasets.

2012-11-01 | E-GEOD-41936 | biostudies-arrayexpress

Identification of bicyclomycin-sensitive transcripts in Vibrio cholerae

Project description:In this study we used BCM, inhibitor of Rho activity, to determine Rho-dependent transcription termination sites in V. cholerae.

2020-05-07 | GSE144478 | GEO

High-resolution RNA 3’-ends mapping of bacterial rho-dependent transcripts

Project description:Transcription termination in bacteria can occur either via Rho-dependent or independent (intrinsic) mechanisms. Intrinsic terminators are composed of a stem-loop RNA structure followed by a uridine stretch and are known to terminate in a precise manner. In contrast, Rho-dependent terminators have more loosely defined characteristics and are thought to terminate in a diffuse manner. While transcripts ending in an intrinsic terminator are protected from 3’-5’ exonuclease digestion due to the stem-loop structure of the terminator, it remains unclear what protects Rho-dependent transcripts from being degraded. In this study, we mapped the exact steady-state RNA 3’ ends of hundreds of E. coli genes terminated either by Rho-dependent or independent mechanisms. We found that transcripts generated from Rho-dependent termination have precise 3’-ends at steady state. These termini were localized immediately downstream of energetically stable stem-loop structures, which were not followed by uridine rich sequences. We provide evidence that these structures protect Rho-dependent transcripts from 3’-5’ exonucleases such as PNPase and RNase II, and present data localizing the Rho-utilization (rut) sites immediately downstream of these protective structures. This study represents the first extensive in-vivo map of exact RNA 3’-ends of Rho-dependent transcripts in E. coli.

2018-04-04 | GSE109766 | GEO

Rho and NusG suppress pervasive antisense transcription in Escherichia coli [RNA-seq].

Project description:Despite the prevalence of antisense transcripts in bacterial transcriptomes, little is known about how their synthesis is controlled. We report that a major function of the Escherichia coli termination factor Rho and its co-factor NusG is suppression of ubiquitous antisense transcription genome-wide. Rho binds C-rich unstructured nascent RNA (high C/G ratio) prior to its ATP-dependent dissociation of transcription complexes. NusG is required for efficient termination at minority subsets (~20%) of both antisense and sense Rho-dependent terminators with lower C/G ratio sequences. In contrast, a widely studied nusA deletion proposed to compromise Rho-dependent termination had no effect on antisense or sense Rho-dependent terminators in vivo. Global co-localization of the nucleoid-associated protein H-NS with Rho-dependent terminators and genetic interactions between hns and rho suggest that H-NS aids Rho in suppression of antisense transcription. The combined actions of Rho, NusG, and H-NS appear to be analogous to the Sen1-Nrd1-Nab3 and nucleosome systems that suppress antisense transcription in eukaryotes. RNA-seq experiments were performed on ribosome-depleted RNA from cells treated with 20 ug/ml bicyclomycin or untreated cells. The series contains 4 datasets.

2012-11-01 | E-GEOD-41939 | biostudies-arrayexpress

Rho and NusG suppress pervasive antisense transcription in Escherichia coli [tiling array].

Project description:Despite the prevalence of antisense transcripts in bacterial transcriptomes, little is known about how their synthesis is controlled. We report that a major function of the Escherichia coli termination factor Rho and its co-factor NusG is suppression of ubiquitous antisense transcription genome-wide. Rho binds C-rich unstructured nascent RNA (high C/G ratio) prior to its ATP-dependent dissociation of transcription complexes. NusG is required for efficient termination at minority subsets (~20%) of both antisense and sense Rho-dependent terminators with lower C/G ratio sequences. In contrast, a widely studied nusA deletion proposed to compromise Rho-dependent termination had no effect on antisense or sense Rho-dependent terminators in vivo. Global co-localization of the nucleoid-associated protein H-NS with Rho-dependent terminators and genetic interactions between hns and rho suggest that H-NS aids Rho in suppression of antisense transcription. The combined actions of Rho, NusG, and H-NS appear to be analogous to the Sen1-Nrd1-Nab3 and nucleosome systems that suppress antisense transcription in eukaryotes. Tiling expression microarray experiments were performed in cells treated with 20 ug/ml bicyclomycin or cells deleted for nusG, or a partial deletion of nusA. Labeled cDNAs were hybridized to an E. coli K-12 MG1655 tiling array with overlapping probes at ~12bp spacing across the entire genome. The series contains 14 datasets.

2012-11-01 | E-GEOD-41938 | biostudies-arrayexpress

Rho and NusG suppress pervasive antisense transcription in Escherichia coli [tiling array].

2012-11-01 | GSE41938 | GEO

Rho and NusG suppress pervasive antisense transcription in Escherichia coli [ChIP-chip].

2012-11-01 | GSE41936 | GEO

Rho and NusG suppress pervasive antisense transcription in Escherichia coli [RNA-seq].

2012-11-01 | GSE41939 | GEO

Analysis of Staphylococcus aureus transcriptome changes in response to inhibition of Rho activity by bicyclomycin

Project description:The transcriptomes of Δrho mutant cells and of wild-type cells grown in RPMI medium with or without 40 or 80 µg/ml of BCM were analyzed using Agilent microarrays in order to analyze the effects of the Rho inhibitor bicyclomycin (BCM) treatment on antisense transcription and changes in the expression of protein coding genes. All antisense RNAs exhibiting higher levels in the rho mutant compared to the wild-type showed similar levels in BCM-treated wild-type cells (80 µg/ml BCM). In addition, microarray data revealed activation of the SaeR regulon in the presence of BCM as observed in the rho mutant.

2018-09-21 | GSE112980 | GEO

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