Abstract
Type three secretion systems (T3SSs) are virulence determinants employed by several pathogenic bacteria as molecular syringes to inject effector proteins into host cells. Diarrhea‐producing enteropathogenic Escherichia coli (EPEC) uses a T3SS to colonize the intestinal tract. T3S is a highly coordinated process that ensures hierarchical delivery of three classes of substrates: early (inner rod and needle subunits), middle (translocators), and late (effectors). Translocation of effectors is triggered upon host‐cell contact in response to different environmental cues, such as calcium levels. The T3S substrate specificity switch from middle to late substrates in EPEC is regulated by the SepL and SepD proteins, which interact with each other and form a trimeric complex with the chaperone CesL. In this study, we investigated the link between calcium concentration and secretion regulation by the gatekeeper SepL. We found that calcium depletion promotes late substrate secretion in a translocon‐independent manner. Furthermore, the stability, formation, and subcellular localization of the SepL/SepD/CesL regulatory complex were not affected by the absence of calcium. In addition, we demonstrate that SepL interacts in a calcium‐independent manner with the major export gate component EscV, which in turn interacts with both middle and late secretion substrates, providing a docking site for T3S. These results suggest that EscV serves as a binding platform for both the SepL regulatory protein and secreted substrates during the ordered assembly of the T3SS.
Highlights
Type III secretion systems (T3SSs) are essential virulence determinants of many animal and plant bacterial pathogens, as they serve to inject virulence proteins, or effectors, directly into the host cell cytoplasm in a process called translocation (Buttner, 2012; Gaytan, Martinez- Santos, Soto, & Gonzalez-Pedrajo, 2016)
Since the proteins forming the SsaL/SpiC/SsaM complex in Salmonella are homologous to the ones forming the SepL/SepD/CesL complex in enteropathogenic Escherichia coli (EPEC), and both protein complexes participate in the substrate specificity switch from translocators to effectors without secretion of the gatekeeper protein (Coombes et al, 2004; Younis et al, 2010; Yu et al, 2010), we argued that the mechanism by which these complexes respond to environmental cues that induce effector secretion could be similar
In EPEC, this secretion hierarchy is proposed to be controlled by the gatekeeper SepL (Deng et al, 2004, 2005; Wang et al, 2008), and to respond to environmental cues such as calcium concentration (Deng et al, 2005; Shaulov et al, 2017)
Summary
Type III secretion systems (T3SSs) are essential virulence determinants of many animal and plant bacterial pathogens, as they serve to inject virulence proteins, or effectors, directly into the host cell cytoplasm in a process called translocation (Buttner, 2012; Gaytan, Martinez- Santos, Soto, & Gonzalez-Pedrajo, 2016). SepL belongs to a family of proteins whose members include MxiC from Shigella (Botteaux, Sory, Biskri, Parsot, & Allaoui, 2009), InvE and SsaL from Salmonella pathogenicity islands 1 and 2, respectively (Coombes, Brown, Valdez, Brumell, & Finlay, 2004; Kubori & Galan, 2002), CopN from Chlamydia (Silva-Herzog et al, 2011), YopN/ TyeA from Yersinia (Forsberg, Viitanen, Skurnik, & Wolf-Watz, 1991; Iriarte et al, 1998), and PopN/Pcr from Pseudomonas (Yang et al, 2007) These proteins, known as gatekeepers, prevent premature effector secretion before host cell contact is established. We investigated the effect of calcium depletion in the gatekeeper-dependent triggering of effector secretion and uncovered the existence of novel protein interactions
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