Characterization of a two-component system in Streptococcus pyogenes whichis involved in regulation of hyaluronic acid production

Hyaluronic acid production by group A streptococci is regulated by transcri ptional control. In this study, transposon mutagenesis of an unencapsulated strain yielded an encapsulated mutant. Two genes homologous to sensors and response regulators of bacterial two-component systems were identified dow nstream of the transposon insertion. Inactivation of the putative sensor ge ne, csrS, in three different unencapsulated strains yielded encapsulated mu tant strains. Electrophoretic mobility shift assays determined factor(s) in a cytoplasmic extract of an unencapsulated group A streptococcal strain wa s binding to a double-stranded DNA fragment derived from the has operon pro moter. In contrast, similarly prepared cytoplasmic extracts from a csrS del etion mutant did not shift the fragment. The putative response regulator, C srR, was partially purified and was shown to bind the has operon promoter f ragment, The affinity and specificity of CsrR for the fragment were increas ed significantly after incubation with acetyl phosphate. DNase I footprinti ng determined that the acetyl phosphate-treated CsrR was binding to key seq uences in the promoter and the coding region of hasA. Therefore, a two-comp onent system is repressing the production of hyaluronic acid in group A str eptococci using a phosphorylation-dependent binding interaction between the response regulator CsrR and the promoter region of the has operon.