Staphylococcus aureus is a well-known pathogenic bacterium that can produce a variety of virulence factors that cause disease in humans. RpiRB is a regulator of metabolic response, whose regulatory role in the virulence of S. aureus is largely unknown. In this study, we demonstrated that the disruption of rpiRB led to down-regulation of the transcription levels of agr-related virulence factors, and reduced the hemolytic activity of methicillin-resistance S. aureus (MRSA). In addition, we also found that RpiRB was involved in regulating the inflammatory response of the host. A mouse subcutaneous abscess model showed that the pathogenicity of the strain was significantly reduced after the destruction of rpiRB. Interestingly, RpiRB enhanced the pathogenic capacity of S. aureus in an agr-dependent manner, while it was directly inhibited by SarA. This study aims to highlight the role of RpiRB in the regulation of the pathogenicity of S. aureus, so as to provide theoretical references for illustrating the infection mechanism and coping strategies of S. aureus.