RNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA

ABSTRACT Ribonucleases facilitate rapid turnover of RNA, providing cells with another mechanism to adjust transcript and protein levels in response to environmental conditions. While many examples have been documented, a comprehensive list of RNase targets is not available. To address this knowledge...

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Autores principales:	Gina C. Gordon, Jeffrey C. Cameron, Brian F. Pfleger
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2017
Materias:	Escherichia coli RNA degradation RNA sequencing RNA stability RNase III Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/03b9810c445b4c19a2aa297414444f73
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spelling	oai:doaj.org-article:03b9810c445b4c19a2aa297414444f732021-11-15T15:51:00ZRNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA10.1128/mBio.00128-172150-7511https://doaj.org/article/03b9810c445b4c19a2aa297414444f732017-05-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00128-17https://doaj.org/toc/2150-7511ABSTRACT Ribonucleases facilitate rapid turnover of RNA, providing cells with another mechanism to adjust transcript and protein levels in response to environmental conditions. While many examples have been documented, a comprehensive list of RNase targets is not available. To address this knowledge gap, we compared levels of RNA sequencing coverage of Escherichia coli and a corresponding RNase III mutant to expand the list of known RNase III targets. RNase III is a widespread endoribonuclease that binds and cleaves double-stranded RNA in many critical transcripts. RNase III cleavage at novel sites found in aceEF, proP, tnaC, dctA, pheM, sdhC, yhhQ, glpT, aceK, and gluQ accelerated RNA decay, consistent with previously described targets wherein RNase III cleavage initiates rapid degradation of secondary messages by other RNases. In contrast, cleavage at three novel sites in the ahpF, pflB, and yajQ transcripts led to stabilized secondary transcripts. Two other novel sites in hisL and pheM overlapped with transcriptional attenuators that likely serve to ensure turnover of these highly structured RNAs. Many of the new RNase III target sites are located on transcripts encoding metabolic enzymes. For instance, two novel RNase III sites are located within transcripts encoding enzymes near a key metabolic node connecting glycolysis and the tricarboxylic acid (TCA) cycle. Pyruvate dehydrogenase activity was increased in an rnc deletion mutant compared to the wild-type (WT) strain in early stationary phase, confirming the novel link between RNA turnover and regulation of pathway activity. Identification of these novel sites suggests that mRNA turnover may be an underappreciated mode of regulating metabolism. IMPORTANCE The concerted action and overlapping functions of endoribonucleases, exoribonucleases, and RNA processing enzymes complicate the study of global RNA turnover and recycling of specific transcripts. More information about RNase specificity and activity is needed to make predictions of transcript half-life and to design synthetic transcripts with optimal stability. RNase III does not have a conserved target sequence but instead recognizes RNA secondary structure. Prior to this study, only a few RNase III target sites in E. coli were known, so we used RNA sequencing to provide a more comprehensive list of cleavage sites and to examine the impact of RNase III on transcript degradation. With this added information on how RNase III participates in transcript regulation and recycling, a more complete picture of RNA turnover can be developed for E. coli. Similar approaches could be used to augment our understanding of RNA turnover in other bacteria.Gina C. GordonJeffrey C. CameronBrian F. PflegerAmerican Society for MicrobiologyarticleEscherichia coliRNA degradationRNA sequencingRNA stabilityRNase IIIMicrobiologyQR1-502ENmBio, Vol 8, Iss 2 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Escherichia coli RNA degradation RNA sequencing RNA stability RNase III Microbiology QR1-502
spellingShingle	Escherichia coli RNA degradation RNA sequencing RNA stability RNase III Microbiology QR1-502 Gina C. Gordon Jeffrey C. Cameron Brian F. Pfleger RNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA
description	ABSTRACT Ribonucleases facilitate rapid turnover of RNA, providing cells with another mechanism to adjust transcript and protein levels in response to environmental conditions. While many examples have been documented, a comprehensive list of RNase targets is not available. To address this knowledge gap, we compared levels of RNA sequencing coverage of Escherichia coli and a corresponding RNase III mutant to expand the list of known RNase III targets. RNase III is a widespread endoribonuclease that binds and cleaves double-stranded RNA in many critical transcripts. RNase III cleavage at novel sites found in aceEF, proP, tnaC, dctA, pheM, sdhC, yhhQ, glpT, aceK, and gluQ accelerated RNA decay, consistent with previously described targets wherein RNase III cleavage initiates rapid degradation of secondary messages by other RNases. In contrast, cleavage at three novel sites in the ahpF, pflB, and yajQ transcripts led to stabilized secondary transcripts. Two other novel sites in hisL and pheM overlapped with transcriptional attenuators that likely serve to ensure turnover of these highly structured RNAs. Many of the new RNase III target sites are located on transcripts encoding metabolic enzymes. For instance, two novel RNase III sites are located within transcripts encoding enzymes near a key metabolic node connecting glycolysis and the tricarboxylic acid (TCA) cycle. Pyruvate dehydrogenase activity was increased in an rnc deletion mutant compared to the wild-type (WT) strain in early stationary phase, confirming the novel link between RNA turnover and regulation of pathway activity. Identification of these novel sites suggests that mRNA turnover may be an underappreciated mode of regulating metabolism. IMPORTANCE The concerted action and overlapping functions of endoribonucleases, exoribonucleases, and RNA processing enzymes complicate the study of global RNA turnover and recycling of specific transcripts. More information about RNase specificity and activity is needed to make predictions of transcript half-life and to design synthetic transcripts with optimal stability. RNase III does not have a conserved target sequence but instead recognizes RNA secondary structure. Prior to this study, only a few RNase III target sites in E. coli were known, so we used RNA sequencing to provide a more comprehensive list of cleavage sites and to examine the impact of RNase III on transcript degradation. With this added information on how RNase III participates in transcript regulation and recycling, a more complete picture of RNA turnover can be developed for E. coli. Similar approaches could be used to augment our understanding of RNA turnover in other bacteria.
format	article
author	Gina C. Gordon Jeffrey C. Cameron Brian F. Pfleger
author_facet	Gina C. Gordon Jeffrey C. Cameron Brian F. Pfleger
author_sort	Gina C. Gordon
title	RNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA
title_short	RNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA
title_full	RNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA
title_fullStr	RNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA
title_full_unstemmed	RNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA
title_sort	rna sequencing identifies new rnase iii cleavage sites in <italic toggle="yes">escherichia coli</italic> and reveals increased regulation of mrna
publisher	American Society for Microbiology
publishDate	2017
url	https://doaj.org/article/03b9810c445b4c19a2aa297414444f73
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RNA Sequencing Identifies New RNase III Cleavage Sites in <italic toggle="yes">Escherichia coli</italic> and Reveals Increased Regulation of mRNA

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