Protein kinase C regulates FADD recruitment and death-inducing signaling complex formation in Fas/CD95-induced apoptosis

Activation of protein kinase C (PKC) triggers cellular signals that inhibit Fas/CD95-induced cell death in Jurkat T-cells by poorly defined mechanisms . Previously, we have shown that one effect of PKC on Fas/CD95-dependent ce ll death occurs through inhibition of cell shrinkage and K+ efflux (Gomez-A ngelats, M., Bortner, C. D., and Cidlowski, J. A. (2000) J. Biol. Chem. 275 , 19609-19619). Here we report that PKC alters Fas/CD95 signaling from the plasma membrane to the activation of caspases by exerting a profound action on survival/cell death decisions. Specific activation of PKC with 12-O-tet radecanoylphorbol-13-acetate or bryostatin-1 induced translocation of PKC f rom the cytosol to the membrane and effectively inhibited cell shrinkage an d cell death triggered by anti-Fas antibody in Jurkat cells. In contrast, i nhibition of classical PKC isotypes with Go6976 exacerbated the effect of F as activation on both apoptotic volume decrease and cell death. PKC activat ion/inhibition did not affect anti-Fas antibody binding to the cell surface , intracellular levels of FADD ((F) under bar as-(a) under bar ssociated pr otein with (d) under bar eath (d) under bar omain), or c-FLIP (cellular (FL ) under bar ICE-like (i) under bar nhibitory (p) under bar rotein) expressi on. However, processing/activation of both caspase-8 and caspase-3 and BID cleavage were markedly blocked upon PKC activation and, conversely, were au gmented during PKC inhibition, suggesting a role for PKC upstream of caspas e-8 processing and activation. Analysis of death-inducing signaling complex (DISC) formation was carried out to examine the influence of PKC on recrui tment of both FADD and procaspase-8 to the Fas receptor. PKC activation blo cked FADD recruitment and caspase-8 activation and thus DISC formation in b oth type I and II cells. In contrast, inhibition of classical PKCs promoted the opposite effect on the Fas pathway by rapidly increasing FADD recruitm ent, caspase-8 activation, and DISC formation. Together, these data show th at PKC finely modulates Fas/CD95 signaling by altering the efficiency of DI SC formation.