Our previous studies revealed that a novel two-component signal transduction system, YhcSR, is essential for the survival of Staphylococcus aureus; however, the biological function of YhcSR remains unknown. In this study, we demonstrated that YhcSR plays an important role in the modulation of the nitrate respiratory pathway under anaerobic conditions. Specifically, we determined that nitrate induces yhcS transcription in the early log phase of growth under anaerobic conditions and that the downregulation of yhcSR expression eliminates the stimulatory effect of nitrate on bacterial growth. Using semiquantitative real-time reverse transcription-PCR (qPCR) and promoter-lux reporter fusions, we established that YhcSR positively modulates the transcription of the narG operon, which is involved in the nitrate respiratory pathway. Our gel shift assays revealed that YhcR binds to the promoter regions of narG and nreABC. Collectively, the above data indicate that the yhcSR system directly regulates the expression of both narG and nreABC operons, which in turn positively modulate the nitrate respiratory pathway of S. aureus under anaerobic conditions. These results provide a new insight into the biological functions of the essential two-component YhcSR system.