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PMID: 18333883 Published · ppublish English Journal Article Research Support, N.I.H., Extramural

Prevalence of RNA polymerase stalling at Escherichia coli promoters after open complex formation.

Molecular microbiology ·Vol. 68 ·No. 1 ·2008-04-00 ·Pages 17-28

Hatoum A, Roberts J

Abstract

RNA polymerase (RNAP) trapped in intermediate stages of promoter escape, as well as RNAP paused at promoter-proximal sigma(70)-dependent pause sites, gives rise to stable, transcriptionally engaged stalled complexes that can limit promoter function and present potential sites for transcription regulation. To investigate the prevalence of such intermediates, we screened 118 Escherichia coli candidate promoters for RNAP stalling at or near the promoter, using in vivo KMnO(4) mapping of RNAP on chromosomal DNA. Of 34 active promoters, the seven preceding lacZ, tnaA, cspA, cspD, rplK, rpsA and rpsU harboured stalled RNAP in vivo; this finding suggests that RNAP stalling after initiation is widespread in E. coli. Consistent with the characteristics of both abortive and promoter-proximal sigma(70)-dependent paused complexes, RNAP trapping at most of the newly identified stall sites was eliminated by the rpoDL402Fsigma(70) mutational alteration and by site mutations, and was enhanced by GreA deficiency. In addition to promoter-proximal RNAP trapping, we observed transcription-dependent DNA modifications spanning the tnaA and cspA leader regions up to 100 bp downstream of the transcription start site.

MeSH Terms
Base Sequence DNA Footprinting DNA, Bacterial/metabolism DNA-Directed RNA Polymerases/metabolism Escherichia coli/genetics,metabolism Escherichia coli Proteins/genetics,metabolism Mutation Promoter Regions, Genetic/genetics Protein Binding RNA, Messenger/genetics,metabolism Sigma Factor/genetics,metabolism Transcription Factors/genetics,metabolism Transcription, Genetic/genetics
Chemicals
DNA, Bacterial Escherichia coli Proteins GreA protein, E coli RNA, Messenger Sigma Factor Transcription Factors DNA-Directed RNA Polymerases
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Hatoum Asma
Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA.
Roberts Jeffrey
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Article Info
Journal
Molecular microbiology
Abbr.
Mol Microbiol
ISSN
1365-2958
DOI
10.1111/j.1365-2958.2008.06138.x
Published
2008-04-00
Pages
17-28
Language
English
Region
England
NLM ID
8712028
PMCID
PMC2819085
Subset
IM
Grants
NIGMS NIH HHS · R01 GM021941 · United States
NIGMS NIH HHS · R01 GM021941-35 · United States
NIGMS NIH HHS · R37 GM021941 · United States
NIGMS NIH HHS · GM21941 · United States
Corrections
CommentIn
PMID: 18312277
External Links
PubMed source DOI: 10.1111/j.1365-2958.2008.06138.x PMC Full Text View journal details
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