ABSTRACTTheStaphylococcus aureustwo-component regulatory system, GraRS, is involved in resistance to killing by distinct host defense cationic antimicrobial peptides (HD-CAPs). It is believed to regulate downstream target genes such asmprFanddltABCDto modify theS. aureussurface charge. However, the detailed mechanism(s) by which the histidine kinase, GraS, senses specific HD-CAPs is not well defined. Here, we studied a well-characterized clinical methicillin-resistantS. aureus(MRSA) strain (MW2), its isogenicgraSdeletion mutant (ΔgraSstrain), a nonameric extracellular loop mutant (ΔEL strain), and four residue-specific ΔEL mutants (D37A, P39A, P39S, and D35G D37G D41G strains). The ΔgraSand ΔEL strains were unable to inducemprFanddltAexpression and, in turn, demonstrated significantly increased susceptibilities to daptomycin, polymyxin B, and two prototypical HD-CAPs (hNP-1 and RP-1). Further, P39A, P39S, and D35G-D37G-D41G ΔEL mutations correlated with moderate increases in HD-CAP susceptibility. Reductions ofmprFanddltAinduction by PMB were also found in the ΔEL mutants, suggesting these residues are pivotal to appropriate activation of the GraS sensor kinase. Importantly, a synthetic exogenous soluble EL mimic of GraS protected the parental MW2 strain against hNP-1- and RP-1-mediated killing, suggesting a direct interaction of the EL with HD-CAPs in GraS activation.In vivo, the ΔgraSand ΔEL strains displayed dramatic reductions in achieved target tissue MRSA counts in an endocarditis model. Taken together, our results provide new insights into potential roles of GraS inS. aureussensing of HD-CAPs to induce adaptive survival responses to these molecules.
Direct interaction between sensor kinase proteins mediates acute and chronic disease phenotypes in a bacterial pathogen