Timing Is Everything:Impact of Naturally Occurring Staphylococcus aureus AgrC Cytoplasmic Domain Adaptive Mutations on Autoinduction

Abstract

Mutations in the polymorphic Staphylococcus aureus agr locus responsible for quorum sensing (QS) dependent virulence gene regulation occur frequently during host adaptation. In two genomically closely related S. aureus clinical isolates exhibiting marked differences in Pantone-Valentine leukocidin production, a mutation conferring an N267I substitution was identified in the cytoplasmic domain of the QS sensor kinase, AgrC. This natural mutation delayed the onset and accumulation of auto-inducing peptide (AIP) and showed reduced responsiveness to exogenous AIPs. Other S. aureus strains harbouring naturally occurring AgrC cytoplasmic domain mutations were identified including T247I, I311T, A343T, L245S and F264C. These mutations were associated with reduced cytotoxicity, delayed/reduced AIP production and impaired sensitivity to exogenous AIP. Molecular dynamics simulations were used to model the AgrC cytoplasmic domain conformational changes arising. While mutations were localised in different parts of the C-terminal domain, their impact on molecular structure was manifested by twisting of the leading helical hairpin α1-α2, accompanied by repositioning of the H-box and G-box along with closure of the flexible loop connecting the two and occlusion of the ATP-binding site. Such conformational rearrangements of key functional subdomains in these mutants highlight the cooperative response of molecular structure involving dimerization, ATP binding and phosphorylation, as well as the binding site for the downstream response element AgrA. These appear to increase the threshold for agr activation via AIP-dependent autoinduction so reducing virulence and maintaining S. aureus in an agr-down-regulated ‘colonization’ mode