Constitutive expression of fibronectin binding in Streptococcus pyogenes as a result of anaerobic activation of rofA

Abstract

Protein F is a fibronectin-binding surface protein of Streptococcus pyogenes (group A streptococcus) that mediates adherence to host cells. A gene product encoded by rofA activates transcription of the gene that encodes protein F (prtF) and was identified in a strain of S. pyogenes that expressed high levels of protein F under all conditions tested. Insertional inactivation of rofA in this strain results in a phenotype similar to that of other strains where high-level transcription of prtF occurs only in response to increased oxygen tension. In this study, we have compared the regulation of prtF and rofA in O 2 -regulated and constitutive strains in order to gain further insight into the function of rofA. Comparison of the prtF and rofA transcripts by S1 nuclease and primer extension assays indicated that the same promoters for each transcript are used in both O 2 -regulated and constitutive strains. However, analyses of rofA-lacZ reporter alleles revealed that a key difference between strains involves regulation of rofA itself. In O 2 -regulated strains, expression of rofA was elevated following culture under conditions of reduced O 2 tension. However, a much more robust activation of rofA expression was observed when constitutive strains were grown under similar conditions. Exchange of reporter and rofA alleles between strains demonstrated that host genetic background, and not the sequence of the respective rofA allele or regulatory region, dictates the expression phenotype. Activation of rofA required RofA, and RofA was shown to bind specifically to DNA containing the promoters for rofA and prtF. Finally, overexpression of either allele of rofA caused constitutive expression of prtF regardless of host background. These data suggest a model where anaerobic expression of prtF in constitutive hosts is controlled at the level of transcription of rofA and implicate additional factors in this regulatory pathway.Studies of many different bacterial pathogens have revealed that complex regulatory networks which sense environmental signals and control expression of particular sets of virulenceassociated genes are essential for adaptation to different environments in the host during infection (reviewed in references 7, 24, and 26). Consequently, an understanding of how virulence factors are regulated can provide important insights into pathogenesis and the contribution of specific genes to different stages of the infection process.

Therapeutic Failures of Antibiotics Used To Treat Macrolide-Susceptible Streptococcus pyogenes Infections May Be Due to Biofilm Formation

“…It is reasonable that persistence of S. pyogenes on surfaces exposed to mechanical stress may be facilitated by biofilm formation. One possible explanation of the stimulating action of lower oxygen tensions on biofilm formation may reside in differential control by regulators at the transcriptional level, as already shown for other virulence traits of S. pyogenes (7,20). Moreover, S. pyogenes may start to form biofilm during the early colonization stages even under nonoptimal conditions; as biofilm forms, the oxygen tension in the lower layers decreases, which may further stimulate biofilm growth.…”

Identification of a Streptolysin S-Associated Gene Cluster and Its Role in the Pathogenesis ofStreptococcus iniaeDisease

“…These findings suggest that a regulatory mechanism, rather than structural differences in SagA, affects the oxygen-dependent expression of SLS in disease-associated strains. In GAS, the CovR-CovS two-component regulatory system and the negative regulators Nra and RofA negatively influence sagA transcription (2,14,37), and rofA is also affected by oxygen conditions (16,17). A small number of strains expressing hemolytic phenotypes opposite to those described for their genotypes (disease associated versus commensal) have been identified.…”

A Novel Sortase, SrtC2, from Streptococcus pyogenes Anchors a Surface Protein Containing a QVPTGV Motif to the Cell Wall

“…1) appear to be differentially regulated by RofA and Nra, respectively. RofA is a positive regulator of protein F under anaerobic conditions in the M6 strain JRS4 (16,17), and under aerobic conditions, expression of protein F is induced by a RofA-independent mechanism (17). In an M49 strain, Nra has been reported to be a negative regulator of Cpa expression (42).…”