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 PKC family, undergoes changes in its phosphorylation state. With a two-dimensional immunoblot procedure we have previously shown the existence in PC12 cells of several multiply phosphorylated forms of PKC-alpha, whose number increases in response to phorbol esters (Gatti, A., Wang, X., and Robinson, P. J. (1996) Biochim. Biophys. Acta 1313, 111-118). Using the same experimental system, here we report that besides the predominant pool of 80-kDa PKC-alpha forms that respond to phorbol ester by translocating to the cell membranes and down-regulating, there is a small 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 kinase may regulate its subcellular localization and susceptibility to down-regulation.
Gatti
A
A
Department of Experimental Oncology, European Institute of Oncology, 20141 Milan, Italy. agatti@ieo.cilea.it
Robinson
P J
PJ
D000818:Animals; D001921:Brain / Q000201:enzymology; D015536:Down-Regulation; D015180:Electrophoresis, Gel, Two-Dimensional; D007527:Isoenzymes / Q000378:metabolism*; D016716:PC12 Cells; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D051381:Rats; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014178:Translocation, Genetic
D007527:Isoenzymes; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate