Role and Regulation of the Secreted Proteases of Staphylococcus aureus - Abstract S. aureus possesses 4 major secreted proteases carried on the core genome: a metalloprotease (aureolysin), a serine protease (V8), and two cysteine proteases (Staphopain A and Staphopain B). In addition, most strains encode up to 9 serine-protease-like enzymes (Spls) carried on the vSab pathogenicity island. Despite the first of these enzymes being identified >45 years ago, we have only recently begun to understand their importance to S. aureus infection. Using a 10-protease gene deletion (Protease Null (PN)) we revealed that secreted protease are key mediators of S. aureus disease. Importantly, their role is biphasic as: (i) Secreted protease deletion leads to markedly increased mortality of infected animals during sepsis; whilst (ii) the PN strain also displays impaired survival in human blood, decreased resistance to phagocytosis, and impaired ability for dissemination and survival during septic infection. The explanation for these seemingly contradictory findings stems from their differing roles, and substrates, during infection. The enhanced mortality of mice is driven by a role for these enzymes in controlling the stability of virulence factors (VF). Thus, in the absence of secreted proteases, VF exist unchecked, accumulating to high levels and causing aggressive and deadly infections. Conversely, virulence attenuation is mediated by a key role for these enzymes in attacking the host, cleaving proteins that facilitate nutrition, immune evasion, inflammation, inactivation of host defenses, tissue destruction, and dysregulation of host processes. However, despite this important knowledge, much remains unknown about how these enzymes are regulated, how they regulate the progression of infection, and how they interact with the host. As such, herein we will dissect the role and regulation of secreted proteases by exploring: Aim #1: The Regulation of S. aureus Secreted Proteases: Secreted proteases are produced by S. aureus alongside its many VF, to modulate their stability, and control the course of infection. During the previous period of support, we identified two non-canonical regulators that profoundly influence protease production. Thus, in this aim we will explore how each of these (a novel regulatory RNA and a unique two-component system) modulate protease activity. Aim #2: The Role of S. aureus Secreted Proteases: During septic infection, the PN strain demonstrates marked decreased in dissemination/survival within mouse organs. This attenuation is mediated by a key role for these enzymes in attacking the host. We know that not all proteases are required for this phenotype, thus we will unravel which proteases contribute to survival during a disseminated septic infection. We will next understand how the proteases mediate their role in vivo using cutting edge mass-spectrometry. Finally, we will conclude this aim determining the role of the Spls during pneumonia.
