title: Biphasic activation of PKBalpha/Akt in platelets. Evidence for stimulation both by phosphatidylinositol 3,4-bisphosphate, produced via a novel pathway, and by phosphatidylinositol 3,4,5-trisphosphate.
abstract: Stimulation of platelet thrombin receptors or protein kinase C causes fibrinogen-dependent aggregation that is a function of integrin alphaIIb beta3 activation. Such platelets rapidly and transiently form phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) and a small amount of phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P2). After aggregation, a larger amount of PtdIns(3,4)P2 is generated. We report that this latter PtdIns(3,4)P2 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 Ca2+-dependent protease calpain. Elevation of cytosolic Ca2+ by ionophore, without integrin/ligand binding, is insufficient to activate the pathway. PtdIns3P 4-K is not the recently described "PIP5KIIalpha." 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 beta gamma subunit of heterotrimeric GTP-binding protein. After aggregation, PtdIns3P 4-K calpain-dependently loses its susceptibility to Gbeta gamma and is, in addition, activated. Both PtdIns(3,4,5)P3 and PtdIns(3,4)P2 have been shown to stimulate PKBalpha/Akt phosphorylation and activation by phosphoinositide-dependent kinase 1. We find that activation of PKBalpha/Akt in platelets is phosphorylation-dependent and biphasic; the initial phase is PtdIns(3,4,5)P3-dependent and more efficient, whereas the second phase depends upon PtdIns(3,4)P2 generated after aggregation. There is thus potential for both pre- and post-aggregation-dependent signaling by PKBalpha/Akt.
authors.affiliation: Kimmel Cancer Institute and Cardeza Foundation for Hematologic Research, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
authors.forename: H
authors.initials: H
authors.lastname: Banfić
authors.forename: C P
authors.initials: CP
authors.lastname: Downes
authors.forename: S E
authors.initials: SE
authors.lastname: Rittenhouse
mesh_terms: D001792:Blood Platelets / Q000201:enzymology*; D002118:Calcium / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D004151:Dipeptides / Q000494:pharmacology; D004789:Enzyme Activation; D005340:Fibrinogen / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism*; D015539:Platelet Activation; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D051552:beta-Adrenergic Receptor Kinases
chemical_list: D004151:Dipeptides; D018129:Phosphatidylinositol Phosphates; D011518:Proto-Oncogene Proteins; C060974:phosphatidylinositol 3,4,5-triphosphate; C060980:phosphatidylinositol 3,4-diphosphate; C071482:calpeptin; D005340:Fibrinogen; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D019204:GTP-Binding Proteins; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium