UNIQUE PHOSPHORYLATION OF PROTEIN-KINASE C-ALPHA IN PC12 CELLS INDUCES RESISTANCE TO TRANSLOCATION AND DOWN-REGULATION

Cell exposure to phorbol ester stimulates translocation and activation of protein kinase C (PKC), ultimately followed by its down-regulation Upon activation, PKC-alpha, the best studied isotype of the PRC famil y, undergoes changes in its phosphorylation state. With a two-dimensio nal immunoblot procedure we have previously shown the existence in PC1 2 cells of several multiply phosphorylated forms of PRC-alpha, whose n umber increases in response to phorbol esters (Gatti, A., Wang, X., an d Robinson, P. J. (1996) Biochim. Biophys. Acta 1313, 111-118), Using the same experimental system, here we report that besides the predomin ant pool of 80-kDa PKC-alpha forms that respond to phorbol ester by tr anslocating to the cell membranes and down-regulating, there is a smal l pool of cytosolic 82-kDa PKC-alpha forms that are characterized by a more acidic pi and by an unique resistance to phorbol ester-mediated translocation and down-regulation, The appearance of similarly slower migrating and more acidic PKC-alpha forms is reproduced upon in vitro autophosphorylation in the presence of phosphatidylserine and phorbol ester, but not in the presence of calcium, These results suggest that site-specific transphosphorylation or autophosphorylation of this kina se may regulate its subcellular localization and susceptibility to dow n-regulation.