The goal of this project is to determine the mechanism of peptide-determined autoinduction of virulence gene expression in Staphylococcus aureus with the long-term objective of defining how this regulation contributes to pathogenesis of S. aureus diseases. The accessory gene regulator (agr) regulon is the central regulatory system that controls the expression of virulence genes in S. aureus. This agr regulon includes: 1) a two-component signal transduction system (AgrA/AgrC); 2) a modified peptide (autoinducing peptide or AIP) that is probably secreted via AgrB, processed from the AgrD propeptide, and used as the ligand for the AgrC sensor; and 3) an RNA molecule, RNAIII, that is the actual regulator of the agr response. We discovered that AIPs produced by some S. aureus strains inhibited the agr response in other strains, and we identified three groups of S. aureus strains that exhibited distinct activation-inhibition activities. Sequence analysis of the agr loci for selected members of each of these three groups revealed a striking pattern of group-specific sequence conservation and divergence. We propose in this application to delineate the steps required for AIP production. We hypothesize that the processing of AgrD propeptide and the secretion of the AIP are jointly catalyzed by AgrB. To test these proposals, we will pursue two specific aims designated to: (1) analyze the mechanisms of AgrD processing and AIP secretion, and (2) determine the function of AgrB in AgrD processing and AIP secretion and identify the catalytic domains and the critical amino acid residues involved in these processes. Genetic and biochemical methods will be used for both aims. The results obtained will facilitate our understanding of this complex agr system and may result in the development of new therapeutic antistaphylococcal drugs that target agr.
