Multiple Roles for the sRNA GcvB in the Regulation of Slp Levels inEscherichia coli

Lorraine T Stauffer,George V Stauffer

doi:10.1155/2013/918106

Abstract

TheEscherichia coli gcvBgene encodes a small RNA that regulates many genes involved in the transport of dipeptides, oligopeptides, and amino acids (oppA,dppA,cycA, andsstT). A microarray analysis of RNA isolated from anE. coliwild-type and aΔgcvBstrain grown to midlog phase in Luria-Bertani broth indicated that genes not involved in transport are also regulated by GcvB. One gene identified wasslpthat encodes an outer membrane lipoprotein of unknown function induced when cells enter stationary phase. The aim of this study was to verify thatslpis a new target for GcvB-mediated regulation. In this study we used RT-PCR to show that GcvB regulatesslpmRNA levels. GcvB negatively controlsslp::lacZin cells grown in Luria-Bertani broth by preventing an Hfq-mediated activation mechanism forslp::lacZexpression. In contrast, in glucose minimal medium supplemented with glycine, GcvB is required for inhibition ofslp::lacZexpression, and Hfq prevents GcvB-mediated repression. Thus, GcvB regulatesslpin both LB and in glucose minimal + glycine media and likely by mechanisms different than how it regulatessstT,dppA,cycA,andoppA. Repression ofslpby GcvB results in an increase in resistance to chloramphenicol, and overexpression ofslpin aΔgcvBstrain results in an increase in sensitivity to chloramphenicol.

Highlights

The E. coli chromosome encodes ∼100 small non-translated regulatory RNAs [1]
Microarray analysis of RNA isolated from an E. coli wild type (WT) strain and an otherwise isogenic ΔgcvB strain grown in Luria-Bertani broth (LB) suggested that Slp, an outer membrane lipoprotein of unknown function [25], is negatively regulated by GcvB [15]
We observed an increased level of DNA corresponding to the slp mRNA amplified from the ΔgcvB sample compared to the level of DNA amplified from either the WT or ΔgcvB complemented strain (Figure 2, compare lane 3 with lanes 2 and 4)

Summary

Introduction

The E. coli chromosome encodes ∼100 small non-translated regulatory RNAs (sRNAs) [1]. A number of these sRNAs have been shown to function as regulators of outer membrane proteins and play important roles in stress responses and virulence gene regulation [2,3,4]. Most of these sRNAs regulate expression of target genes posttranscriptionally by base pairing with the target mRNAs [5]. The Hfq protein is required for regulation by this class of sRNAs [6]. Base pairing results in negative regulation of translational activity and altered stability of the target mRNA [5]. DsrA and RprA bind to rpoS mRNA, likely preventing formation of an inhibitory secondary structure that sequesters the ribosome-binding site, resulting in increased translation [7,8,9]

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