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J Bacteriol. 2012 Jan;194(1):79-89. doi: 10.1128/JB.06209-11. Epub 2011 Oct 28.

Translational repression of NhaR, a novel pathway for multi-tier regulation of biofilm circuitry by CsrA.

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  • 1Department of Microbiology and Cell Science, University of Florida, Gainesville, Florida, USA.

Abstract

The RNA binding protein CsrA (RsmA) represses biofilm formation in several proteobacterial species. In Escherichia coli, it represses the production of the polysaccharide adhesin poly-β-1,6-N-acetyl-D-glucosamine (PGA) by binding to the pgaABCD mRNA leader, inhibiting pgaA translation, and destabilizing this transcript. In addition, CsrA represses genes responsible for the synthesis of cyclic di-GMP, an activator of PGA production. Here we determined that CsrA also represses NhaR, a LysR-type transcriptional regulator which responds to elevated [Na(+)] and alkaline pH and activates the transcription of the pgaABCD operon. Gel shift studies revealed that CsrA binds at two sites in the 5' untranslated segment of nhaR, one of which overlaps the Shine-Dalgarno sequence. An epitope-tagged NhaR protein, expressed from the nhaR chromosomal locus, and an nhaR posttranscriptional reporter fusion (PlacUV5-nhaR'-'lacZ) both showed robust repression by CsrA. Northern blotting revealed a complex transcription pattern for the nhaAR locus. Nevertheless, CsrA did not repress nhaR mRNA levels. Toeprinting assays showed that CsrA competes effectively with the ribosome for binding to the translation initiation region of nhaR. Together, these findings indicate that CsrA blocks nhaR translation. Epistasis studies with a pgaA-lacZ transcriptional fusion confirmed a model in which CsrA indirectly represses pgaABCD transcription via NhaR. We conclude that CsrA regulates the horizontally acquired pgaABCD operon and PGA biosynthesis at multiple levels. Furthermore, nhaR repression exemplifies an expanding role for CsrA as a global regulator of stress response systems.

PMID:
22037401
[PubMed - indexed for MEDLINE]
PMCID:
PMC3256615
Free PMC Article
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