SarA plays a predominant role in controlling the production of extracellular proteases in the diverse clinical isolates of Staphylococcus aureus LAC and UAMS-1

SUMMARY

Using DNA affinity chromatography we demonstrate that the S. aureus regulatory proteins MgrA, Rot, SarA, and SarS bind DNA baits derived from the promoter regions associated with the genes encoding aureolysin, ScpAB, SspABC, and SplA-F. Three of four baits also bound SarR and SarZ, the exception in both cases being the ScpAB-associated bait. Using the USA300, methicillin-resistant strain LAC and the USA200, methicillin-sensitive strain UAMS-1, we generated mutations in the genes encoding each of these proteins alone and in combination with sarA and examined the impact on protease production, the accumulation of high molecular weight proteins, and biofilm formation. These studies confirmed that multiple regulatory loci are involved in limiting protease production to a degree that impacts all of these phenotypes, but also demonstrate that sarA plays a predominant role in this regard. Using sarA mutants unable to produce individual proteases alone and in combination with each other, we also demonstrate that the increased production of aureolysin and ScpA is particularly important in defining the biofilm-deficient phenotype of LAC and UAMS-1 sarA mutants, while aureolysin alone plays a key role in defining the reduced accumulation of alpha toxin and overall cytotoxicity as assessed using both osteoblasts and osteoclasts.

KEYWORDS:

• Staphylococcus aureus
• extracellular protease
• regulation
• biofilm
• sarA
• rot
• mgrA
• sarS
• sarZ
• sarR

Acknowledgments

This work was supported by NIH grant R01-AI119380 to MSS. Additional support was provided by a generous gift from the Texas Hip and Knee Center and research core facilities supported by the Center for Microbial Pathogenesis and Host Inflammatory Responses (P20-GM103450), the Translational Research Institute (UL1TR000039), the United States Army Congressionally Directed Medical Research Programs (W81X1H-14-PRORP-EA), and NIH grant R01-AI124458 to LNS. AJT acknowledges support from the National Institutes of Health grants P20GM121293, UL1TR000039, S10OD018445, and P20GM103429.

Author contributions

Conceived and designed the experiments: AR, KB, MS. Performed the experiments: AR, KB, SB, BG. Analyzed the data: AR, KB, SB, AT, LS, MS. Wrote the paper: AR, KB, MS.

Disclosure statement

The authors report no conflict of interest.

Supplementary material

Supplemental data for this article can be accessed here.

Additional information

Funding

This work was supported by the National Institute of Allergy and Infectious Diseases [AI119380].