Protein kinase C V₃ domain mutants with differential sensitivities to m‐calpain are not resistant to phorbol‐ester‐induced down‐regulation

Distinct linker sequences were introduced into the protease-sensitive V3 domain of protein kinase C-alpha and the mutant proteins were expressed in COS-1 cells. Partially purified preparations of these mutants were functionally similar to wild-type protein kinase C-alpha, however their susceptibility to m-calpain was quite distinct, with one mutant being insensitive to cleavage. The three mutants, after expression in COS-1 cells, were found to behave in a manner indistinguishable from wild-type protein kinase C-alpha with respect to subcellular distribution, acute responses to 12-O-tetradecanoyl-phorbol 13-acetate and 12-O-tetradecanoyl-phorbol-13-acetate-induced down-regulation. The data imply that down-regulation of protein kinase C-alpha is likely to involve a general degradative process rather than cleavage by a site-specific protease.