The alkaliphilic side ofStaphylococcus aureus

Abstract

AbstractThe genome ofStaphylococcus aureushas eight structurally distinctcation/protonantiporters (CPA) that play significant roles in maintaining cytoplasmic pH and ions in extreme conditions. These antiporters enableS. aureusto persist under conditions that are favorable to the bacterium but unfavorable to animal host including humans. In this study, we report physiological roles and catalytic properties of NhaC (NhaC1, NhaC2 and NhaC3), CPA1 (CPA1-1 and CPA1-2) and CPA2 family antiporters and how these antiporters crosstalk with Mnh1, a CPA3 family antiporter, recently shown to play important roles in virulence and pH tolerance. Catalytic properties of antiporters were determined by Na+/H+and K+/H+antiport assays using everted membrane vesicles of a CPA-deficientE. coliKNabC host. NhaC and CPA1 candidates exhibited Na+/H+and K+/H+antiporter activity in the pH range between pH 7 to 9.5 but did not show significant role in halotolerance and osmotolerance alone. Interestingly, NhaC3 exhibited significant antiporter activity at alkaline pH and play major roles in pH and salt tolerance. CPA2 neither exhibited Na+or K+/H+exchange nor showed any active role in pH and salt tolerance. Double deletion ofmnhA1withnhaC1, nhaC3, cpa1-1 or cpa1-2respectively, madeS. aureusseverely sensitive at pH 7.5 under stress conditions indicating synergistic relationship of Mnh1 with these antiporters. The functional loss study of these antiporters inin-vivomouse infection model,nhaC3deletion showed significant loss ofS. aureusvirulence. Altogether, the current study indicates NhaC3 as a potential target againstS. aureusvirulence under extreme pH and salt conditions.ImportanceIn this study, we established catalytic properties and physiological roles ofS. aureusNhaC, CPA1 and CPA2 family antiporters and their importance under salt and alkaline stress conditions. Except CPA2, all five antiporters of both families were active for Na+/H+and K+/H+exchange. CPA1-1 showed significant role in pH homeostasis at pH 7.5 whereas CPA1-2 and NhaCs were major contributors to halotolerance and osmotolerance at alkaline pH. The severity of growth deficit in double knockouts ofmnhA1with each ofnhaC1, nhaC2, nhaC3, cpa1-1orcpa1-2establishes their synergistic relationship in regulating pH and salt homeostasis. Deletion ofcpa1-1, cpa1-2andnhaC1, nhaC2, andnhaC3were assessed in mice model and NhaC3 was shown to play a major role inS. aureusvirulence.