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The regulation of apoptosis by toxic shock syndrome toxin-1 and associated mutants Hung, Ryan Wei Yan


Toxic shock syndrome toxin-1 (TSST-1), the primary virulence factor in toxic shock syndrome (TSS), is an exotoxin produced by Staphylococcus aureus and a member of the superantigen family of T cell mitogens. Activation-induced cell death (AICD) of superantigen-reactive T cells is thought to play a role in modulating the hyperactive immune response and promoting host survival. While this form of apoptotic cell death occurs with the related staphylococcal enterotoxins (SE) A and B, conflicting reports exist for TSST-1. Therefore, the aim of this project was to investigate the regulation of apoptosis by TSST-1 and site-directed mutants. TSST-1 was found to exert either pro- or anti-apoptotic effects depending on the dose, thus resolving some of the contradictory reports in the literature. At commonly used concentrations (<1 μM), TSST-1 was less potent at inducing apoptosis than SEB. Moreover, there was evidence of early induction of anti-apoptotic pathways. The glycine 31-to-arginine site-directed mutant of TSST-1 (G31R) was serendipitously found to be potently pro-apoptotic at doses as low as 10-nM. This apoptosis was very rapid and broadly directed, affecting T cells, monocytes and T lymphoma-derived cell lines. Surprisingly, high doses of TSST-1 (<1 μM) also exhibited similarly potent, non-specific apoptosis much like G31R. The mechanism behind this novel form of apoptosis was examined by dissecting the involvement of components of the extrinsic and intrinsic apoptotic pathways. When ligated, classical death receptors like Fas receptor utilize the extrinsic pathway. In contrast, conditions unfavourable to cell survival such as growth factor withdrawal, ionizing radiation, and other cell damage can converge at the level of the mitochondria to trigger death through the intrinsic pathway. G31R-induced apoptosis was dependent on the intrinsic, but not the extrinsic, pathway. Because TSST-1 is not known to possess direct enzymatic effects that could account for its cytotoxicity, the most likely site of action for the induction of apoptosis by G31R and high-dose TSST-1 was considered to be a non-classical death receptor coupled to the intrinsic pathway, a phenomenon previously described. However, the upstream pathways remain unclear, so that other possibilities may include direct cell membrane disruption and/or induction of reactive oxygen species. Either way, this previously unknown apoptotic activity of TSST-1 may be relevant in pathogenesis, acting as a local immunosuppressant at infection sites by deleting lymphocytes and monocytes that transit into the region. With identification of the apical mechanism for apoptosis in this system, possibilities exist for the therapeutic use of G31R or its derivatives in the treatment of tumours and autoimmune diseases.

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