Skip to main content
bioRxiv
  • Home
  • About
  • Submit
  • ALERTS / RSS
Advanced Search
New Results

Mechanism of upstream promoter element stimulation of transcription at a ribosomal RNA promoter determined by single-molecule imaging

Jeffrey P. Mumm, View ORCID ProfileLarry J. Friedman, View ORCID ProfileJeff Gelles
doi: https://doi.org/10.1101/2020.02.17.953182
Jeffrey P. Mumm
aDepartment of Biochemistry, Brandeis University, Waltham, MA 02454-9110, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Larry J. Friedman
aDepartment of Biochemistry, Brandeis University, Waltham, MA 02454-9110, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Larry J. Friedman
Jeff Gelles
aDepartment of Biochemistry, Brandeis University, Waltham, MA 02454-9110, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Jeff Gelles
  • For correspondence: gelles@brandeis.edu
  • Abstract
  • Full Text
  • Info/History
  • Metrics
  • Supplementary material
  • Preview PDF
Loading

Abstract

DNA elements upstream of transcription promoters play a role in regulating transcription initiation in all organisms. In bacteria, upstream A-T rich sequences called UP elements can stimulate transcription through contact with the α subunit C-terminal domain (αCTD) of core RNA polymerase (RNAP), but the kinetic mechanisms by which they do so remain unclear. We investigated the role of the UP element in stimulating initiation from the strong E. coli 16s rRNA promoter using single-molecule fluorescence microscopy to visualize σ70RNAP holoenzyme binding and the formation nascent RNA by oligonucleotide probe hybridization on individual DNA molecules containing the rrnB P1 promoter. By directly detecting initial binding of σ70RNAP to promoter and monitoring the lifetimes of promoter-polymerase complexes, the experiments reveal the kinetic mechanism of polymerase recruitment to the promoter and the subsequent conformational change that stabilizes binding. The presence of UP stimulated the rate of initial binding of polymerase to promoter by at least six-fold, and this stimulation was fully sufficient to account for the increase in initiation rate by UP. Thus, UP likely functions at this strong promoter simply by acting as a binding target for the rapidly reorienting αCTD domain tethered to the core polymerase. In contrast, there were only minor effects of UP on the measured rates of the conformational change or the dissociation rates of the initial σ70RNAP promoter complexes. These studies define a paradigmatic kinetic mechanism for stimulation of transcription initiation by direct αCTD-DNA interactions. This mechanism can serve as a building block of more complex regulatory architectures in which αCTD promotes transcription through interactions with both DNA and protein activators.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
Back to top
PreviousNext
Posted February 18, 2020.
Download PDF

Supplementary Material

Email

Thank you for your interest in spreading the word about bioRxiv.

NOTE: Your email address is requested solely to identify you as the sender of this article.

Enter multiple addresses on separate lines or separate them with commas.
Mechanism of upstream promoter element stimulation of transcription at a ribosomal RNA promoter determined by single-molecule imaging
(Your Name) has forwarded a page to you from bioRxiv
(Your Name) thought you would like to see this page from the bioRxiv website.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
Mechanism of upstream promoter element stimulation of transcription at a ribosomal RNA promoter determined by single-molecule imaging
Jeffrey P. Mumm, Larry J. Friedman, Jeff Gelles
bioRxiv 2020.02.17.953182; doi: https://doi.org/10.1101/2020.02.17.953182
Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
Citation Tools
Mechanism of upstream promoter element stimulation of transcription at a ribosomal RNA promoter determined by single-molecule imaging
Jeffrey P. Mumm, Larry J. Friedman, Jeff Gelles
bioRxiv 2020.02.17.953182; doi: https://doi.org/10.1101/2020.02.17.953182

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
  • Tweet Widget
  • Facebook Like
  • Google Plus One

Subject Area

  • Biochemistry
Subject Areas
All Articles
  • Animal Behavior and Cognition (2504)
  • Biochemistry (4951)
  • Bioengineering (3452)
  • Bioinformatics (15126)
  • Biophysics (6854)
  • Cancer Biology (5356)
  • Cell Biology (7680)
  • Clinical Trials (138)
  • Developmental Biology (4502)
  • Ecology (7109)
  • Epidemiology (2059)
  • Evolutionary Biology (10178)
  • Genetics (7491)
  • Genomics (9750)
  • Immunology (4801)
  • Microbiology (13131)
  • Molecular Biology (5109)
  • Neuroscience (29284)
  • Paleontology (203)
  • Pathology (832)
  • Pharmacology and Toxicology (1454)
  • Physiology (2116)
  • Plant Biology (4714)
  • Scientific Communication and Education (1002)
  • Synthetic Biology (1332)
  • Systems Biology (3993)
  • Zoology (766)