The prpZ gene cluster encoding eukaryotic-type Ser/Thr protein kinases and phosphatases is repressed by oxidative stress and involved in Salmonella enterica serovar Typhi survival in human macrophages

Abstract

The prpZ gene cluster consists of three ORFs coding for proteins with homology to eukaryotic-type Ser/Thr protein phosphatases 2C (prpZ) and Ser/Thr protein kinases (prkY and prkX). This cluster is present in the sequenced genomes of Salmonella enterica serovar Typhi (S. Typhi) strains Ty2 and CT18. This study investigated the genetic organization of this gene cluster, its regulation and its putative involvement in virulence. The three genes are transcribed as a polycistronic mRNA as demonstrated by reverse transcriptase (RT)-PCR. Analysis of a prpZ::lacZ transcriptional fusion showed that the prpZ locus is expressed throughout the growth phase. LacZ activity and real-time RT-PCR showed that transcription of the mRNA is negatively regulated upon exposure of cells to HOCl and, to a lesser extent, hydrogen peroxide. A deletion mutant of the prpZ gene cluster showed a significantly lower level of survival than the parental strain Ty2 in human macrophages at 48 h postinfection. Together these data suggest that prpZ, prkY and prkX are virulence genes that may be part of a signaling pathway controlling long-term survival of S. Typhi in host cells.

An Inducible and Secreted Eukaryote-Like Serine/Threonine Kinase of Salmonella enterica Serovar Typhi Promotes Intracellular Survival and Pathogenesis

“…Although T4519 was predicted previously to be a Ser/Thr kinase, this was never experimentally validated (17). We used in vitro kinase assays to investigate whether T4519 is a bona fide Ser/Thr kinase.…”

“…Hydrogen peroxide, but not hypochlorite, N minimal medium (low pH, low Mg 2ϩ ), or iron, was capable of inducing T4519, indicating that superoxide ions drive T4519 expression (Fig. 2G) (17).…”

“…However, these eSTKs were not found to play a significant role in pathogenesis (16). eSTK sequences were also identified in the S. Typhi genome (10,17). We show here that T4519 is an authentic eSTK of S. Typhi that is induced within macrophages in a reactive oxygen species (ROS)-dependent manner.…”

Eukaryote-Like Serine/Threonine Kinases and Phosphatases in Bacteria

“…Homologs of PrkC have also been implicated in S. pyogenes adherence and invasion and in E. faecalis persistence (72,82). The involvement of eSTKs in virulence has also been reported for several other pathogens, such as Streptococcus pneumoniae, E. faecalis, Pseudomonas aeruginosa, and Salmonella enterica serovar Typhi (33,43,46,(144)(145)(146)191).…”

Typhi genes expressed during infection or involved in pathogenesis

“…PagC is produced in typhoid patients [13]. On SPI-10, the prpZ locus encoding for proteins with homology to eukaryotic-type Ser/Thr protein phosphatase and kinases has been found to promote survival in macrophages [35] (Table 1).…”