INTRACELLULAR CALCIUM-DEPENDENT ACTIVATION OF P72(SYK) IN PLATELETS

We previously demonstrated that thrombin-induced activation of p72(syk ) was independent of intracellular Ca2+ elevation in platelets. Howeve r, our previous studies also demonstrated that activation of platelets by ionophore A23187 results in a dramatic increase in tyrosine phosph orylation of several cellular proteins. In the present study, we inves tigated the effect of Ca2+ elevation on the activity of p72(syk). When washed porcine platelets were stimulated with ionophore A23187 and th e activity of p72(syk) was assessed by means of an immunoprecipitation kinase assay, A23187 caused a time- and dose-dependent increase in th e specific activity of p72(syk). In addition, pretreatment of platelet s with both aspirin and ADP scavengers or chelation of extracellular C a2+ by EGTA had no effect on the A23187-induced activation of p72(syk) . These results indicate that A23187-induced activation of p72(syk) is independent of the formation of endoperoxide/thromboxage A(2), releas ed ADP and extracellular Ca2+, suggesting the existence of a novel pat hway for activation of p72(syk). Furthermore, evidence is presented wh ich indicates a synergistic effect of A23187 and thrombin on the activ ation of p72(syk), and an inhibitory effect of pretreatment with phorb ol 12-myristate 13-acetate, a protein kinase C activator, on the activ ation of p72(syk) induced by either A23187 or thrombin.