RNA polymerase supply and flux through the lac operon in Escherichia coli.

Sendy, Bandar and Lee, David J and Busby, Stephen J W and Bryant, Jack A (2016) RNA polymerase supply and flux through the lac operon in Escherichia coli. Philosophical transactions of the Royal Society of London. Series B, Biological sciences, 371 (1707). ISSN 1471-2970

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Abstract

Chromatin immunoprecipitation, followed by quantification of immunoprecipitated DNA, can be used to measure RNA polymerase binding to any DNA segment in Escherichia coli By calibrating measurements against the signal from a single RNA polymerase bound at a single promoter, we can calculate both promoter occupancy levels and the flux of transcribing RNA polymerase through transcription units. Here, we have applied the methodology to the E. coli lactose operon promoter. We confirm that promoter occupancy is limited by recruitment and that the supply of RNA polymerase to the lactose operon promoter depends on its location in the E. coli chromosome. Measurements of RNA polymerase binding to DNA segments within the lactose operon show that flux of RNA polymerase through the operon is low, with, on average, over 18 s elapsing between the passage of transcribing polymerases. Similar low levels of flux were found when semi-synthetic promoters were used to drive transcript initiation, even when the promoter elements were changed to ensure full occupancy of the promoter by RNA polymerase.This article is part of the themed issue 'The new bacteriology'.

Item Type: Article
Identification Number: https://doi.org/10.1098/rstb.2016.0080
Date: 5 November 2016
Uncontrolled Keywords: polymerase per second, chromatin immunoprecipitation, promoter occupation, RNA polymerase, polymerase density, rifampicin
Subjects: C100 Biology
C400 Genetics
C500 Microbiology
Divisions: Faculty of Health, Education and Life Sciences > School of Health Sciences
Depositing User: David Lee
Date Deposited: 14 Jan 2020 08:47
Last Modified: 12 Feb 2020 13:19
URI: http://www.open-access.bcu.ac.uk/id/eprint/8710

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