The glutathione import system satisfies the Staphylococcus aureus nutrient sulfur requirement and promotes interspecies competition.

Sulfur is an indispensable element for bacterial proliferation. Prior studies demonstrated that the human pathogen Staphylococcus aureus utilizes glutathione (GSH) as a source of nutrient sulfur; however, mechanisms of GSH acquisition are not defined. Here, we identify a five-gene locus comprising a putative ABC-transporter and predicted γ-glutamyl transpeptidase (ggt) that promotes S. aureus proliferation in medium supplemented with either reduced or oxidized GSH (GSSG) as the sole source of nutrient sulfur. Based on these phenotypes, we name this transporter operon the glutathione import system (gisABCD). Ggt is encoded within the gisBCD operon, and we show that the enzyme is capable of liberating glutamate using either GSH or GSSG as substrates, demonstrating it is a bona fide γ-glutamyl transpeptidase. We also determine that Ggt is expressed in the cytoplasm, representing only the second example of cytoplasmic Ggt localization, the other being Neisseria meningitidis. Bioinformatic analyses revealed that Staphylococcus species closely related to S. aureus encode GisABCD-Ggt homologs. However, homologous systems were not detected in Staphylococcus epidermidis. Consequently, we establish that GisABCD-Ggt provides a competitive advantage for S. aureus over S. epidermidis in a GSH- and GSSG-dependent manner. Overall, this study describes the discovery of a nutrient sulfur acquisition system in S. aureus that targets GSSG in addition to GSH and promotes competition against other staphylococci commonly associated with the human microbiota.

The Staphylococcus aureus Cystine Transporters TcyABC and TcyP Facilitate Nutrient Sulfur Acquisition during Infection.

Staphylococcus aureus is a significant human pathogen due to its capacity to cause a multitude of diseases. As such, S. aureus efficiently pillages vital nutrients from the host; however, the molecular mechanisms that support sulfur acquisition during infection have not been established. One of the most abundant extracellular sulfur-containing metabolites within the host is cysteine, which acts as the major redox buffer in the blood by transitioning between reduced and oxidized (cystine) forms. We therefore hypothesized that S. aureus acquires host-derived cysteine and cystine as sources of nutrient sulfur during systemic infection. To test this hypothesis, we used the toxic cystine analogue selenocystine to initially characterize S. aureus homologues of the Bacillus subtilis cystine transporters TcyABC and TcyP. We found that genetic inactivation of both TcyA and TcyP induced selenocystine resistance. The double mutant also failed to proliferate in medium supplemented with cystine, cysteine, or N-acetyl cysteine as the sole sulfur source. However, only TcyABC was necessary for proliferation in defined medium containing homocystine as the sulfur source. Using a murine model of systemic infection, we observed tcyP-dependent competitive defects in the liver and heart, indicating that this sulfur acquisition strategy supports proliferation of S. aureus in these organs. Phylogenetic analyses identified TcyP homologues in many pathogenic species, implying that this sulfur procurement strategy is conserved. In total, this study is the first to experimentally validate sulfur acquisition systems in S. aureus and establish their importance during pathogenesis.