Staphylococcus aureus regulates the expression and production of the staphylococcal superantigen‐like secreted proteins in a Rot‐dependent manner

Abstract

SUMMARY S. aureus over-produces a subset of immunomodulatory proteins known as the staphylococcal superantigen like proteins (Ssls) under conditions of pore-mediated membrane stress. In this study we demonstrate that overproduction of Ssls during membrane stress is due to the impaired activation of the two-component module of the quorum sensing Accessory gene regulator (Agr) system. Agr-dependent repression of ssl expression is indirect and mediated by the transcription factor Repressor of toxins (Rot). Surprisingly, we observed that Rot directly interacts with and activates the ssl promoters. The role of Agr and Rot as regulators of ssl expression was observed across several clinically relevant strains, suggesting that over-production of immunomodulatory proteins benefits agr-defective strains. In support of this notion, we demonstrate that Ssls contribute to the residual virulence of S. aureus lacking agr in a murine model of systemic infection. Altogether, these results suggest that S. aureus compensates for the inactivation of Agr by producing immunomodulatory exoproteins that could protect the bacterium from host-mediated clearance.

Rot and SaeRS Cooperate To Activate Expression of the Staphylococcal Superantigen-Like Exoproteins

“…The construction of ssl7, ssl9, and ssl11 reporter plasmids was described previously (7). For construction of a plasmid expressing the green fluorescent protein (GFP) gene under the control of the sarA promoter, a PCR amplicon was made using pCM11 as the template to amplify the sarA promoter and GFP gene using primers 347 and 348.…”

“…GFP reporter assays were performed as described previously (7). Briefly, overnight cultures of S. aureus grown in RPMI-CAS plus CM were diluted 1:100 into 5 ml of RPMI-CAS plus CM.…”

“…We have previously demonstrated that the regulation of ssl expression by Rot is due to the promoter activation by this transcription factor (7). To test whether Sae directly regulates the activation of the ssl promoters, the ssl7 and ssl9 promoters were fused to the GFP gene and the resulting transcriptional reporters were transformed into WT S. aureus strain Newman or the rot or sae-QRS isogenic mutants.…”

“…We recently demonstrated that inactivation of agr in strain Newman resulted in the production of higher levels of Ssls than in the wild-type (WT) strain and that the ability of Agr to repress ssls was dependent on the RNAIII-mediated regulation of Rot (7). Surprisingly, we found that Rot regulates ssl expression by directly binding to and activating the ssl promoters.…”

“…Among the immunomodulators, the staphylococcal superantigen-like proteins (Ssls) represent a family of 14 secreted proteins, each with a molecular mass of around 25 kDa, that have a variety of immunomodulatory properties, including inhibition of complement activation and neutrophil recruitment, as well as blocking of opsonization by IgG and IgA (17). Importantly, Ssl overproduction is associated with S. aureus pathogenesis in animal models of systemic infection (3,7,52).…”