Biphasic Activation of PKBα/Akt In Platelets (original) (raw)

1998, Journal of Biological Chemistry

Stimulation of platelet thrombin receptors or protein kinase C causes fibrinogen-dependent aggregation that is a function of integrin ␣ IIb ␤ 3 activation. Such platelets rapidly and transiently form phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P 3) and a small amount of phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P 2). After aggregation, a larger amount of PtdIns(3,4)P 2 is generated. We report that this latter PtdIns(3,4)P 2 arises largely through wortmannin-inhibitable generation of PtdIns3P and then phosphorylation by PtdIns3P 4-kinase (PtdIns3P 4-K), a novel pathway apparently contingent upon the activation of the Ca 2؉-dependent protease calpain. Elevation of cytosolic Ca 2؉ by ionophore, without integrin/ligand binding, is insufficient to activate the pathway. PtdIns3P 4-K is not the recently described "PIP5KII␣." Cytoskeletal activities of phosphatidylinositol 3-kinase and PtdIns3P 4-K increase after aggregation. Prior to aggregation, PtdIns3P 4-K can be regulated negatively by the ␤␥ subunit of heterotrimeric GTP-binding protein. After aggregation, PtdIns3P 4-K calpain-dependently loses its susceptibility to G␤␥ and is, in addition, activated. Both PtdIns(3,4,5)P 3 and PtdIns(3,4)P 2 have been shown to stimulate PKB␣/Akt phosphorylation and activation by phosphoinositide-dependent kinase 1. We find that activation of PKB␣/Akt in platelets is phosphorylation-dependent and biphasic; the initial phase is PtdIns(3,4,5)P 3-dependent and more efficient, whereas the second phase depends upon PtdIns(3,4)P 2 generated after aggregation. There is thus potential for both pre-and postaggregation-dependent signaling by PKB␣/Akt.