The Staphylococcus aureus nitric oxide synthase (saNOS) modulates respiratory metabolism and cell physiology

Nitric oxide synthase (NOS) enzymes produce nitric oxide (NO), a highly reactive free radical capable of interacting with multiple cellular targets. Although saNOS contributes to Staphylococcus aureus virulence, as well as protection against exogenous oxidative stress and antimicrobials, the current mechanism behind these phenotypes is unknown. Here we report a previously-undescribed role for saNOS in modulating S. aureus physiology. When grown aerobically, endogenous reactive oxygen species (ROS) and superoxide levels were elevated in a S. aureus nos mutant. NO has been shown to slow respiration in other organisms, and likewise, comparison of respiratory dehydrogenase activity and membrane potential in wild-type, nos mutant, and complement strains suggested that saNOS-derived NO limits S. aureus aerobic respiration. Multiple transcriptional and metabolic changes were also observed in a S. aureus nos mutant, as assessed by RNAseq and targeted metabolomics analyses, respectively. Specifically, expression of genes associated with oxidative and nitrosative stress responses, anaerobic and lactate metabolism, and cytochrome biosynthesis and assembly were increased in the nos mutant relative to wild-type. Metabolites utilized to produce reducing equivalents by the right arm of the TCA cycle were depleted in a nos mutant (citrate and α-ketoglutarate), whereas fumarate and malate levels were increased relative to wild-type and complement strains. A significant reduction in lactate levels was also observed in the nos mutant. Collectively, these results support a model in which the absence of saNOS results in increased respiration and ROS accumulation, which may signal the cells to switch to an alternative lactate-based respiratory metabolism. Three RNAseq data sets in total. Each sample was generated by pooling three independent biological replicate RNA preps