PMID:22037401

Citation	Pannuri, A, Yakhnin, H, Vakulskas, CA, Edwards, AN, Babitzke, P and Romeo, T (2012) Translational repression of NhaR, a novel pathway for multi-tier regulation of biofilm circuitry by CsrA. J. Bacteriol. 194:79-89
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.
Links	PubMed PMC3256615 Online version:10.1128/JB.06209-11
Keywords	Biofilms/growth & development; Cyclic GMP/analogs & derivatives; Cyclic GMP/metabolism; DNA-Binding Proteins/genetics; DNA-Binding Proteins/metabolism; Down-Regulation; Escherichia coli/physiology; Escherichia coli Proteins/genetics; Escherichia coli Proteins/metabolism; Gene Expression Regulation, Bacterial/physiology; Hydrogen-Ion Concentration; Porins; Protein Biosynthesis/physiology; RNA, Messenger/genetics; RNA, Messenger/metabolism; RNA-Binding Proteins/genetics; RNA-Binding Proteins/metabolism; Repressor Proteins/genetics; Repressor Proteins/metabolism; Sodium; Transcription Factors/genetics; Transcription Factors/metabolism; beta-Glucans/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
ECOLI:CSRA	involved_in	GO:0045947: negative regulation of translational initiation	ECO:0000314: direct assay evidence used in manual assertion		P		Seeded From UniProt	complete
ECOLI:CSRA	enables	GO:0048027: mRNA 5'-UTR binding	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete

References

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