Group A streptococcal growth phase-associated virulence factor regulation by a novel operon (Fas) with homologies to two-component-type regulators requires a small RNA molecule

A novel growth phase-associated two-component-type regulator, Fas ((f) unde r bar ibronectin/fibrinogen binding/haemolytic (a) under bar ctivity/(s) un der bar treptokinase regulator), of Streptococcus pyogenes was identified i n the M1 genome sequence, based on homologies to the histidine protein kina se (HPK) and response regulator (RR) part of the Staphylococcus aureus Agr and Streptococcus pneumoniae Com quorum-sensing systems. The fas operon, pr esent in all 12 tested M serotypes, was transcribed as polycystronic messag e (fasBCA) and contained genes encoding two potential HPKs (FasB and FasC) and one RR (FasA). Downstream of fasBCA, we identified a small 300 nucleoti de monocistronic transcript, designated fasX, that did not appear to encode true peptide sequences. Measurements of luciferase promoter fusions reveal ed a growth phase-associated transcription of fasBCA and fasX, with peak ac tivities during the late exponential phase. Insertional mutagenesis disrupt ing fasBCA and fasA led to a phenotype similar to agr-null mutations in S. aureus, with prolonged expression of extracellular matrix protein-binding a dhesins and reduced expression of secreted virulence factors such as strept okinase and streptolysin S. In addition, fasX transcription was dependent o n the RR FasA; however, deletion mutagenesis of fasX resulted in a similar phenotype to that of the fasBCA or fasA mutants. Complementation of the fas X deletion mutant, with the fasX gene expressed in trans from a plasmid, re stored the wild-type fasBCA regulation pattern. This strongly suggested tha t fasX, a putative non-translated RNA, is the main effector molecule of the fas regulon. However, using spent culture supernatants from wild-type and fas mutant strains, we were not able to show an influence on the logarithmi c growth phase expression of fas and dependent genes. Thus, despite structu ral and functional similarities between fas and agr, to date the fas operon appears not to be involved in group A streptococcal (GAS) quorum-sensing r egulation.