Characterization of the sar locus and its interaction with agr in Staphylococcus aureus

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Abstract
The expression of cell wall and extracellular proteins in Staphylococcus aureus is controlled by global regulatory systems, including sar and agr. We have previously shown that a transposon insertion into the 372-bp sarA gene within the sar locus resulted in decreased expression of several extracellular and cell wall proteins (A. L. Cheung and S. J. Projan, J. Bacteriol. 176:4168-4172, 1994). In this study, Northern (RNA blot) analysis with a 732-bp sarA probe indicated that two major transcripts (0.56 and 1.2 kb) were absent in the sar mutant compared with the parental strain RN6390. Additional transcriptional studies revealed that the sarA gene is encoded within the 0.56-kg transcript. Notably, a plasmid carrying the sarA gene together with a 1.2-kb upstream fragment (1.7 kb total) was able to reestablish the 1.2-kb transcript in the mutant. Although reconstitution of the parental phenotype by the sarA gene was incomplete, the introduction of a plasmid carrying the 1.7-kb fragment to the mutant restored the parental phenotype. Transcription of RNAII and RNAIII, which encode the structural and regulatory genes of agr, respectively, was diminished in the mutant but restored to wild-type levels by complementation with the 1.7-kb fragment. In gel shift assays, cell extracts of this clone were able to retard the mobility of a labeled RNAII promoter probe but not an RNAIII promoter element. These data suggest that sarA and the adjacent upstream DNA are essential to the expression of a DNA-binding protein(s) with specificity for the RNAII promoter, thereby controlling agr-related transcription.