Activation of atypical protein kinase C zeta by caspase processing and degradation by the ubiquitin-proteasome system

Atypical protein kinase C zeta (PKC zeta) is known to transduce signals tha t influence cell proliferation and survival. Here we show that recombinant human caspases can process PKC zeta at three sites in the hinge region betw een the regulatory and catalytic domains. Caspase-3, -6, -7, and -8 chiefly cleaved human PKC zeta at EETD down arrow G, and caspase-3 and -7 also cle aved PKC zeta at DGMD down arrow G and DSED down arrow L, respectively. Pro cessing of PKC zeta expressed in transfected cells occurred chiefly at EETD down arrow G and DGMD down arrow G and produced carboxyl-terminal polypept ides that contained the catalytic domain. Epitope-tagged PKC zeta that lack ed the regulatory domain was catalytically active following expression in H eLa cells. Induction of apoptosis in HeLa cells by tumor necrosis factor cu plus cycloheximide evoked the conversion of full-length epitope-tagged PKC zeta to two catalytic domain polypeptides and increased PKC zeta activity. A caspase inhibitor, zVAD-fmk, prevented epitope-tagged PKC zeta processin g and activation following the induction of apoptosis. Induction of apoptos is in rat parotid C5 cells produced catalytic domain polypeptides of endoge nous PKC zeta and increased PKC zeta activity. Caspase inhibitors prevented the increase in PKC zeta activity and production of the catalytic domain p olypeptides. Treatment with lactacystin, a selective inhibitor of the prote asome, caused polyubiquitin-PKC zeta conjugates to accumulate in cells tran sfected with the catalytic domain or full-length PKC alpha, or with a PKC z eta mutant that was resistant to caspase processing. We conclude that caspa ses process PKC zeta to carboxyl-terminal fragments that are catalytically active and that are degraded by the ubiquitin-proteasome pathway.