Interaction of protein kinase C with filamentous actin: Isozyme specificity resulting from divergent phorbol ester and calcium dependencies

The mechanism of activation of protein kinase C isoforms by filamentous act in (F-actin) was investigated with respect to isozyme specificity and phorb ol ester and Ca2+ dependencies. It was found that the "conventional" (cPKC) , alpha, beta I, beta II, and gamma, "novel" (nPKC) delta and epsilon, and "atypical" (aPKC) zeta isoforms were each activated by F-actin with varying potencies. The level of activity along with the affinity for binding to F- actin was further potentiated by the phorbol ester 4 beta-12-O-tetradecanoy lphorbol 13-acetate (TPA), the potency of which again varied for each isofo rm. By contrast to the other cPKC isoforms, the level of cPKC-gamma activit y was unaffected by TPA, as was also the case for aPKC-zeta. It was found t hat whereas in the absence of F-actin the soluble form of cPKC-beta I conta ined two phorbol ester binding sites of low and high affinity, respectively , as previously reported for cPKC-alpha [Slater et al. (1998) J. Biol, Chem , 273, 23160-23168], the F-actin-bound form of the isozyme contained only a single site of relatively low affinity, The level of TPA required to induc e cPKC-alpha, -beta I, and -beta II activity and the binding of these isozy mes to F-actin was reduced in the presence of Ca2+. By contrast, the activi ty of cPKC-gamma was unaffected by Ca2+, as were the activities of nPKC-del ta and -epsilon and aPKC-zeta, as expected. Thus, the interaction with F-ac tin appears to be a general property of each of the seven PKC isozymes test ed. However, isoform specificity may, in part, be directed by differences i n the phorbol eater and Ca2+ dependences, which, with the notable exception of cPKC-gamma, appear to resemble those observed for the activation of eac h isoform by membrane association. The observation that cPKC isoforms may t ranslocate to F-actin as well as the membrane as a response to an elevation of Ca2+ levels may allow for the functional coupling of fluctuations of in tracellular Ca2+ levels through cPKC to F-actin cytoskeleton-mediated proce sses.