Vinculin phosphorylation in response to calcium and phorbol esters in intact cells.
Vinculin phosphorylation in both chick embryo fibroblasts and Swiss 3T3 cells was increased by either calcium or biologically active phorbol esters. Increased phosphorylation of vinculin was noted as early as 10 min following phorbol 12-myristate 13-acetate treatment and was maximal at about 1 h. Maximal increases in phosphorylation were noted at approximately 100 nM phorbol 12-myristate 13-acetate. Phorbol 12,13-dibutyrate (80 nM), a less potent phorbol ester, resulted in smaller increases in vinculin phosphorylation than phorbol 12-myristate 13-acetate at equimolar concentrations. Phorbol, dibutyryl cAMP, and dibutyryl cGMP had no significant effect on phosphorylation. No correlation was found between vinculin phosphorylation and the morphological changes induced by phorbol esters. Tryptic peptide analysis of vinculin revealed multisite phosphorylation. Phosphorylation of only three of the peptides was significantly increased following phorbol 12-myristate 13-acetate treatment. Phosphoamino acid analysis revealed increases at both serine and threonine residues. The low level of phosphotyrosine present in control cells was not significantly increased by phorbol 12-myristate 13-acetate treatment. These findings combined with studies of vinculin phosphorylation by purified protein kinase C (Werth, D. K., Niedel, J. E., and Pastan I. (1983) J. Biol. Chem. 258, 11423-11426) suggest the hypothesis that protein kinase C may be involved in regulation of phosphorylation of vinculin, a cytoskeletal protein.
D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology*; D002478:Cells, Cultured; D002642:Chick Embryo; D003995:Dibutyryl Cyclic GMP / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D007700:Kinetics; D051379:Mice; D009124:Muscle Proteins / Q000378:metabolism*; D010446:Peptide Fragments / Q000032:analysis; D015240:Phorbol 12,13-Dibutyrate; D010703:Phorbol Esters / Q000494:pharmacology*; D010704:Phorbols / Q000494:pharmacology*; D010748:Phosphopeptides / Q000032:analysis; D010766:Phosphorylation; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014357:Trypsin; D016596:Vinculin
D009124:Muscle Proteins; D010446:Peptide Fragments; D010703:Phorbol Esters; D010704:Phorbols; D010748:Phosphopeptides; D016596:Vinculin; D003995:Dibutyryl Cyclic GMP; D015240:Phorbol 12,13-Dibutyrate; D003994:Bucladesine; D014357:Trypsin; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium