OmpR positively regulates urease expression to enhance acid survival of Yersinia pseudotuberculosis

Abstract

Yersinia pseudotuberculosis is an enteric bacterium which must overcome the acidic stress in host organs for successful colonization, but how this bacterium survives in acidic conditions remains largely unknown. In the present study, the importance of OmpR in acid survival of Y. pseudotuberculosis YpIII was confirmed by the fact that mutation of ompR (strain ΔompR) greatly reduced cell survival at pH 4.5 or lower. To characterize the regulatory role of OmpR in this acid survival process, proteomic analysis was carried out to compare YpIII at pH 7.0 and pH 4.5 with ΔompR at pH 7.0, and urease components were revealed to be the main targets for OmpR regulation. Addition of urea to the culture medium also enhanced acid survival of YpIII but not ΔompR and urease activity was significantly induced by acid in YpIII but not in ΔompR. Each of the seven components of the YpIII urease gene cluster was fused to a lacZ reporter and their expression was dramatically decreased in a ΔompR background; this supports the notion that OmpR positively regulates urease expression. Furthermore, gel shift analysis revealed that OmpR binds to the deduced promoter regions of three polycistronic transcriptional units (ureABC, ureEF and ureGD) in the urease cluster, suggesting that the regulation of OmpR to urease components is direct. Taken together, these data strongly suggest that OmpR activates urease expression to enhance acid survival in Y. pseudotuberculosis.

LcrQ Blocks the Role of LcrF in Regulating the Ysc-Yop Type III Secretion Genes in Yersinia pseudotuberculosis

“…All these amplified non-coding fragments were cloned into the promoter cloning vector pZT100 [9]. These clones were respectively transformed into YPIII or its derivative strains and the β-galactosidase assay was performed at least in triplicate as previously described [26] at 26°C, or 37°C or 37°C with Ca 2+ depletion.…”

“…The gel-retardation was performed as described previously [26]. The LcrF protein was expressed in E. coli BL21(DE3) by pET28a plasmid carrying the lcrF gene from YPIII (named pET28a-LcrF), and purified by Ni-NTA resin.…”

“…To over-express LcrQ protein, lcrQ gene was amplified from YPIII and cloned into pOVR200 to obtain pOVR-LcrQ, which was transformed into YPIII. For co-expression of LcrF and LcrQ (or GST-LcrQ), the low copy plasmid named pKT100 [26], which contains the pOVR200 compatible P15A replicon and a promoter region of the chloramphenicol resistance gene, was used to express the target proteins together with the pOVR200 plasmid.…”

Pathogenesis ofY. enterocoliticaandY. pseudotuberculosisin Human Yersiniosis

“…A drop in pH induces the expression of the YPT aspertase ( aspA ) gene; the encoded gene product, AspA, subsequently produces ammonia, allowing the ingested organisms to survive the acidic gastrointestinal environment [125]. Other bacterial factors that promote survival under acidic conditions include urease [126], TatC [127], PhoP, OmpR, and PmrA [128, 129]. Acidic pH also induces a downregulation of the transcriptional regulator, Cra (for catabolite repressor/activator), which increases bacterial acid survival [130].…”

Transcriptomic and Phenotypic Analysis Reveals New Functions for the Tat Pathway in Yersinia pseudotuberculosis

“…In addition to its role in metabolism, the urease operon is known to be essential for virulence and acid resistance of enteric pathogens, including Y. enterocolitica (54). It has been suggested that the expression of urease genes is regulated by OmpR in Y. pseudotuberculosis (55), but there is no direct evidence for a role of urease in virulence (56). The adiAC genes, encoding arginine decarboxylase and the arginine-agmatine antiporter, respectively, were highly upregulated (fold change, 7.4 and 6.8, respectively; Table S2).…”