Identification of a unique co-operative phosphoinositide 3-kinase signaling mechanism regulating integrin alpha IIb beta 3 adhesive function in platelets.
Phosphoinositide (PI) 3-kinases play an important role in regulating the adhesive function of a variety of cell types through affinity modulation of integrins. Two type I PI 3-kinase isoforms (p110 beta and p110 gamma) have been implicated in G(i)-dependent integrin alpha(IIb)beta(3) regulation in platelets, however, the mechanisms by which they coordinate their signaling function remains unknown. By employing isoform-selective PI 3-kinase inhibitors and knock-out mouse models we have identified a unique mechanism of PI 3-kinase signaling co-operativity in platelets. We demonstrate that p110 beta is primarily responsible for G(i)-dependent phosphatidylinositol 3,4-bisphosphate (PI(3,4)P(2)) production in ADP-stimulated platelets and is linked to the activation of Rap1b and AKT. In contrast, defective integrin alpha(IIb)beta(3) activation in p110 gamma(-/-) platelets was not associated with alterations in the levels of PI(3,4)P(2) or active Rap1b/AKT. Analysis of the effects of active site pharmacological inhibitors confirmed that p110 gamma principally regulated integrin alpha(IIb)beta(3) activation through a non-catalytic signaling mechanism. Inhibition of the kinase function of PI 3-kinases, combined with deletion of p110 gamma, led to a major reduction in integrin alpha(IIb)beta(3) activation, resulting in a profound defect in platelet aggregation, hemostatic plug formation, and arterial thrombosis. These studies demonstrate a kinase-independent signaling function for p110 gamma in platelets. Moreover, they demonstrate that the combined catalytic and non-catalytic signaling function of p110 beta and p110 gamma is critical for P2Y(12)/G(i)-dependent integrin alpha(IIb)beta(3) regulation. These findings have potentially important implications for the rationale design of novel antiplatelet therapies targeting PI 3-kinase signaling pathways.
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Simone M
SM
Schoenwaelder
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Akiko
A
Ono
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Sharelle
S
Sturgeon
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Siew Mei
SM
Chan
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Pierre
P
Mangin
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Mhairi J
MJ
Maxwell
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Shannon
S
Turnbull
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Megha
M
Mulchandani
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Karen
K
Anderson
Auckland Cancer Society Research Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland 1020, New Zealand.
Gilles
G
Kauffenstein
Auckland Cancer Society Research Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland 1020, New Zealand.
Gordon W
GW
Rewcastle
Auckland Cancer Society Research Centre, Faculty of Medical and Health Sciences, University of Auckland, Auckland 1020, New Zealand.
Jackie
J
Kendall
INSERM U.311, EFS-Alsace, Strasbourg, France.
Christian
C
Gachet
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004.
Hatem H
HH
Salem
Australian Centre for Blood Diseases, Monash University, Alfred Medical Research and Education Precinct (AMREP), 89 Commercial Road, Melbourne, Victoria, Australia 3004. Electronic address: Shaun.Jackson@med.monash.edu.au.
Shaun P
SP
Jackson
D000244:Adenosine Diphosphate / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001792:Blood Platelets / Q000166:cytology / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000819:agonists / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010973:Platelet Adhesiveness / Q000187:drug effects / Q000502:physiology*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D058910:Purinergic P2 Receptor Agonists; D018048:Receptors, Purinergic P2 / Q000378:metabolism; D058925:Receptors, Purinergic P2Y12; D015290:Second Messenger Systems / Q000187:drug effects / Q000502:physiology*; D020668:rap GTP-Binding Proteins / Q000378:metabolism
D000244:Adenosine Diphosphate; D004791:Enzyme Inhibitors; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D007527:Isoenzymes; D008565:Membrane Proteins; D019869:Phosphatidylinositol 3-Kinases; D018129:Phosphatidylinositol Phosphates; D000081082:Phosphoinositide-3 Kinase Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D051057:Proto-Oncogene Proteins c-akt; D058910:Purinergic P2 Receptor Agonists; D018048:Receptors, Purinergic P2; D058925:Receptors, Purinergic P2Y12; D020668:rap GTP-Binding Proteins; C548577:P2ry12 protein, mouse; C060980:phosphatidylinositol 3,4-diphosphate; C492136:Rap1b protein, mouse