Fas aggregation does not correlate with Fas-mediated apoptosis

Cross-linking of cell surface Fas molecules by Fas ligand or by agonistic a nti-Fas Abs induces cell death by apoptosis. We found that a serine proteas e inhibitor, N-tosyl-L-lysine chloromethyl ketone (TLCK), dramatically enha nces Fas-mediated apoptosis in the human T cell line Jurkat and in various B cell lines resistant to Fas-mediated apoptosis. The enhancing effect of T LCK is specific to Fas-induced cell death, with no effect seen on TNF-alpha or TNF-related apoptosis-inducing ligand-induced apoptosis. TLCK treatment had no effect on Fas expression levels on the cell surface, and neither pr omoted death-inducing signaling complex formation nor decreased expression levels of cellular inhibitors of apoptosis (FLICE inhibitory protein, X chr omosome-linked inhibitor of apoptosis, and Bcl-2). Activation of the Fas-me diated apoptotic pathway by anti-Fas Ab is accompanied by aggregation of Fa s molecules to form oligomers that are stable to boiling in SDS and beta -N M. Fas aggregation is often considered to be required for Fas-mediated apop tosis. However, sensitization of cells to Fas-mediated apoptosis by TLCK or other agents (cycloheximide, protein kinase C inhibitors) causes less Fas aggregation during the apoptotic process compared with that in nonsensitize d cells. These results show that Fas aggregation and Fas-mediated apoptosis are not directly correlated and may even be inversely correlated.