The P2Y(12) receptor induces platelet aggregation through weak activation of the alpha(IIb)beta(3) integrin - a phosphoinositide 3-kinase-dependent mechanism

High concentrations of adenosine-5'-diphosphate ADP are able to induce part ial aggregation without shape change of P2Y(1) receptor-deficient mouse pla telets through activation of the P2Y(12) receptor. In the present work we s tudied the transduction pathways selectively involved in this phenomenon. F low cytometric analyses using R-phycoerythrin-conjugated JON/A antibody (JO N/A-PE), an antibody which recognizes activated Mouse alpha (IIb)beta (3) i ntegrin, revealed a low level activation Of alpha (IIb)beta (3) in P2Y(1) r eceptor-deficient platelets in response to 100 muM ADP or 1 muM 2MeS-ADP. A drenaline induced no such activation but strongly potentiated the effect of ADP in a dose-dependent manner. Global phosphorylation of P-32-labeled pla telets showed that P2Y(12)-mediated aggregation was not accompanied by an i ncrease in the phosphorylation of myosin light chain (P-20) or pleckstrin ( P-47) and was not affected by the protein kinase C (PKC) inhibitor staurosp orine. On the other hand, two unrelated phosphoinositide 3-kinase inhibitor s, wortmannin and LY294002, inhibited this aggregation. Our results indicat e that (i) the P2Y(12) receptor is able to trigger a P2Y(1) receptor-indepe ndent inside-out signal leading to alpha (IIb)beta (3) integrin activation and platelet aggregation, (ii) ADP and adrenaline use different signaling p athways which synergize to activate the alpha (IIb)beta (3) integrin, and ( iii) the transduction pathway triggered by the P2Y12 receptor is independen t of PKC but dependent on phosphoinositide 3-kinase. (C) 2001 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rig hts reserved.