Protein kinase C-mediated formation of phosphatidylinositol 3,4-bisphosphate in human platelets

The effect of phorbol 12,13-dibutyrate on the formation of phosphatidylinositol 3,4-bisphosphate in washed human platelets was studied. Platelets labelled with [32P]Pi were stimulated with phorbol 12,13-dibutyrate or thrombin in the presence or absence of staurosporine. Lipids were extracted, and deacylated, and the glycerophosphoinositol derivatives were analyzed by high performance liquid chromatography. Phorbol 12,13-dibutyrate increased formation of phosphatidylinositol 4-monophosphate and phosphatidylinositol 3,4-bisphosphate in a dose- and time-dependent manner. Thrombin also increased formation of phosphatidylinositol 3,4-bisphosphate. Staurosporine completely inhibited phorbol 12,13-dibutyrate or thrombin-stimulated production of phosphatidylinositol 3,4-bisphosphate. These data indicate that production of phosphatidylinositol 3,4-bisphosphate and phosphatidylinositol 4-monophosphate is mediated by protein kinase C. It is widely recognized that production of phosphatidylinositol 3,4-bisphosphate is caused by the tyrosine kinase-mediated activation of phosphatidylinositol 3-kinase. However, in platelets, production of phosphatidylinositol 3,4-bisphosphate might be related to stimulation of phosphatidylinositol 4-kinase, which is activated by protein kinase C.