Regulation of the Yersinia pseudotuberculosis Type III Secretion System by the CpxAR Two‐Component System

Host tissue cues including temperature, oxygen, and iron availability regulate expression of the Yersinia type III secretion system (T3SS) master regulator LcrF. We found that CpxR inhibits LcrF through an indirect, multi-factorial mechanism and identified the osmolarity-sensing OmpR/EnvZ system as an additional regulator of LcrF, highlighting integrated environmental control of virulence gene expression. ABSTRACT The type III secretion system (T3SS) is a cell envelope-spanning injectisome found in many Gram-negative pathogens. Expression of the T3SS is controlled by extracellular signals such as cell envelope stress. The CpxAR two-component system negatively regulates the Yersinia pseudotuberculosis Ysc T3SS, although the mechanism remains unclear. As expected, we found that mutants with constitutive CpxR activity (∆cpxA and cpxR D51E) led to decreased expression of the Ysc T3SS and its master regulator LcrF. CpxR did not bind to the regulatory regions of lcrF, suggesting an indirect mechanism of regulation. Transcriptome analysis showed that 101 genes were upregulated and 77 genes downregulated in both the ∆cpxA and cpxR D51E strains compared to wildtype, including seven genes known to modulate transcription. Individual deletion of these seven regulatory factors did not identify any single gene responsible for CpxR-dependent repression of LcrF, suggesting that CpxAR may act through multiple pathways to regulate the Ysc T3SS. However, this analysis led us to examine the role of the osmolarity sensing two-component regulatory system OmpR/EnvZ. Deletion of ompR led to an increase in LcrF and T3SS expression. These results suggest more complex regulation of the Ysc T3SS by two-component regulatory systems than previously appreciated.