Phosphorylated CpxR Restricts Production of the RovA Global Regulator in Yersinia pseudotuberculosis

BACKGROUND: RovA is a global transcriptional regulator of gene expression in pathogenic Yersinia. RovA levels are kept in check by a sophisticated layering of distinct transcriptional and post-transcriptional regulatory mechanisms. In the enteropathogen Y. pseudotuberculosis, we have previously reported that the extracytoplasmic stress sensing CpxA-CpxR two-component regulatory system modulates rovA expression. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we characterized CpxR phosphorylation (CpxR∼P) in vitro, and determined that phosphorylation was necessary for CpxR to efficiently bind to the PCR-amplified upstream regulatory region of rovA. The precise CpxR∼P binding site was mapped by a nuclease protection assay and directed mutagenesis confirmed that in vivo binding to the rovA promoter inhibits transcription. Reduced RovA production was most pronounced following CpxR∼P accumulation in the Yersinia cytoplasm during chronic Cpx pathway activation and by the indiscriminate phosphodonor action of acetyl phosphate. CONCLUSIONS/SIGNIFICANCE: Cpx pathway activation restricts levels of the RovA global regulator. The regulatory influence of CpxR∼P must therefore extend well beyond periplasmic quality control in the Yersinia envelope, to include genes involved in environmental survival and pathogenicity.