Abstract
Type III secretion systems (T3SSs) contribute to microbial pathogenesis of Vibrio species, but the regulatory mechanisms are complex. We determined if the classic ExsACDE protein-protein regulatory model from Pseudomonas aeruginosa applies to Vibrio alginolyticus. Deletion mutants in V. alginolyticus demonstrated that, as expected, the T3SS is positively regulated by ExsA and ExsC and negatively regulated by ExsD and ExsE. Interestingly, deletion of exsE enhanced the ability of V. alginolyticus to induce host-cell death while cytotoxicity was inhibited by in trans complementation of this gene in a wild-type strain, a result that differs from a similar experiment with Vibrio parahaemolyticus ExsE. We further showed that ExsE is a secreted protein that does not contribute to adhesion to Fathead minnow epithelial cells. An in vitro co-immunoprecipitation assay confirmed that ExsE binds to ExsC to exert negative regulatory effect on T3SS genes. T3SS in V. alginolyticus can be activated in the absence of physical contact with host cells and a separate regulatory pathway appears to contribute to the regulation of ExsA. Consequently, like ExsE from P. aeruginosa, ExsE is a negative regulator for T3SS gene expression in V. alginolyticus. Unlike the V. parahaemolyticus orthologue, however, deletion of exsE from V. alginolyticus enhanced in vitro cytotoxicity.
Highlights
The type III secretion system (T3SS) is an important virulence-associated surface structure of many Gram-negative pathogens, where it functions to translocate bacterial effector proteins across the bacterial and host membranes directly into the cytosol of host cells (Tseng et al, 2009)
The type III secretion system (T3SS) gene expression is induced by contact with eukaryotic cells or under specific environmental conditions
In P. aeruginosa T3SS genes are upregulated under low-calcium growth condition (Finck-Barbancon et al, 1997; Vallis et al, 1999) and ExsE serves as a secreted regulator that indirectly contributes to the transcriptional activator ExsA by releasing ExsC to bind to ExsD (Rietsch et al, 2005)
Summary
The type III secretion system (T3SS) is an important virulence-associated surface structure of many Gram-negative pathogens, where it functions to translocate bacterial effector proteins across the bacterial and host membranes directly into the cytosol of host cells (Tseng et al, 2009). T3SSs are composed of a secretion apparatus, translocation apparatus and effector proteins (Hueck, 1998). The regulatory pathway can be remarkably complex and usually involves a series of interacting proteins (Yahr and Wolfgang, 2006; Hauser, 2009). Transcription of T3SS genes in Pseudomonas aeruginosa is controlled by ExsA, which is a member of the AraC/XyIS family of transcriptional regulators (Yahr and Wolfgang, 2006; Hauser, 2009). The transcriptional activity of ExsA is regulated by three additional interacting proteins: ExsE in Vibrio alginolyticus
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