TheSalmonellaSPI1 Type Three Secretion System Responds to Periplasmic Disulfide Bond Status via the Flagellar Apparatus and the RcsCDB System

Abstract

Upon contact with intestinal epithelial cells, Salmonella enterica serovar Typhimurium injects a set of effector proteins into the host cell cytoplasm via the Salmonella pathogenicity island 1 (SPI1) type III secretion system (T3SS) to induce inflammatory diarrhea and bacterial uptake. The master SPI1 regulatory gene hilA is controlled directly by three AraC-like regulators: HilD, HilC, and RtsA. Previous work suggested a role for DsbA, a periplasmic disulfide bond oxidase, in SPI1 T3SS function. RtsA directly activates dsbA, and deletion of dsbA leads to loss of SPI1-dependent secretion. We have studied the dsbA phenotypes by monitoring expression of SPI1 regulatory, structural, and effector genes. Here we present evidence that loss of DsbA independently affects SPI1 regulation and SPI1 function. The dsbA-mediated feedback inhibition of SPI1 transcription is not due to defects in the SPI1 T3SS apparatus. Rather, the transcriptional response is dependent on both the flagellar protein FliZ and the RcsCDB system, which also affects fliZ transcription. Thus, the status of disulfide bonds in the periplasm affects expression of the SPI1 system indirectly via the flagellar apparatus. RcsCDB can also affect SPI1 independently of FliZ. All regulation is through HilD, consistent with our current model for SPI1 regulation.

DsbL and DsbI contribute to periplasmic disulfide bond formation in Salmonella enterica serovar Typhimurium

“…The hilA gene, encoding the master regulator of the SPI1 T3SS (Bajaj et al, 1995), is transcriptionally controlled by a complex regulatory circuit (Ellermeier et al, 2005). Consistent with our previous data (Lin et al, 2008), deletion of dsbA or dsbB significantly decreased the expression of hilA (Fig. 2a).…”

“…of the SPI1 system (Lin et al, 2008). SPI1 transcriptionally responds to disulfide bond status through the flagellar apparatus and the RcsCDB system (Lin et al, 2008). Here we present evidence that DsbL and DsbI also contribute to disulfide bond formation in the Salmonella periplasm.…”

“…We previously showed that loss of DsbA, the primary disulfide bond oxidoreductase in the periplasm, or the cognate DsbB, decreases transcription of the SPI1 genes in serovar Typhimurium (Ellermeier & Slauch, 2004;Lin et al, 2008). During our studies, we recognized a pair of DsbA and DsbB paralogues encoded together in the Salmonella genome.…”

“…The lacZ fusion construct was subsequently introduced as described above. The hilA-lacZ fusion, integrated at the lambda attachment site, had been constructed previously (Lin et al, 2008). The following genes were amplified using primers carrying EcoRI or BamHI sites and then cloned into vector pWKS30 (Wang & Kushner, 1991) with base-pairs indicated relative to the start site of translation of the first gene in the construct: assT (214A+1808); assT dsbLI (214A+3194); dsbL (214A+687); dsbI (214A+693); dsbLI (214A+1379); dsbA (230A+645); dsbB (221A+547).…”

“…DsbL and DsbI homologues, not associated with ASST enzymes, are also apparent in a variety of Gram-negative organisms (Kimball et al, 2003;Raczko et al, 2005). of the SPI1 system (Lin et al, 2008). SPI1 transcriptionally responds to disulfide bond status through the flagellar apparatus and the RcsCDB system (Lin et al, 2008).…”