REVIEW ARTICLES: Platelet G protein-coupled receptors in hemostasis and thrombosis

Abstract

The same processes that govern hemostasis to stem blood loss after injury also contribute to thrombosis, particularly at sites of arterial atherosclerosis [1Ruggeri Z.M. Platelets in atherothrombosis.Nat Med. 2002; 8: 1227-34Crossref PubMed Scopus (1351) Google Scholar]. Platelet activation is initiated at sites of injury or atherosclerotic lesions as platelets encounter the combination of matrix proteins revealed beneath a damaged endothelial lining and the shear stress of non-laminar blood flow [2Jackson S.P. Nesbitt W.S. Kulkarni S. Signaling events underlying thrombus formation.J Thromb Haemost. 2003; 1: 1602-12Abstract Full Text Full Text PDF PubMed Scopus (337) Google Scholar, 3Savage B. Almus-Jacobs F. Ruggeri Z.M. Specific synergy of multiple substrate-receptor interactions in platelet thrombus formation under flow.Cell. 1998; 94: 657-66Abstract Full Text Full Text PDF PubMed Scopus (678) Google Scholar, 4Badimon L. Badimon J.J. Turitto V.T. Vallabhajosula S. Fuster V. Platelet thrombus formation on collagen type I. A model of deep vessel injury. Influence of blood rheology, von Willebrand factor, and blood coagulation.Circulation. 1988; 78: 1431-42Crossref PubMed Google Scholar]. Thus, a greater understanding of the mechanisms regulating hemostasis also have a broader application as we seek to develop novel targeted therapies to control thrombosis. G protein-coupled receptors (GPCRs) or the agonists that activate them have already proven to be key targets for platelet-directed therapy in reducing thrombotic complications in patients at risk for recurrent heart attack and stroke [5Micieli G. Cavallini A. New therapeutic strategies with antiplatelet agents.Neurol Sci. 2004; 25: S13-5Crossref PubMed Scopus (10) Google Scholar]. The most recent examples of platelet-directed therapeutics that target GPCRs are clopidigrel and ticlopidine, which target the P2Y12 receptor for adenosine diphosphate (ADP) [6Dorsam R.T. Kunapuli S.P. Central role of the P2Y12 receptor in platelet activation.J Clin Invest. 2004; 113: 340-5Crossref PubMed Google Scholar]. However, inhibitors of cyclooxygenase, such as aspirin, reduce production of the platelet agonist thromboxane A2, which also stimulates a GPCR. This review will detail the role of individual GPCRs in hemostasis and thrombosis, the functions of the G proteins activated by these agonists, and the known effectors of G protein-mediated signaling in platelets. All of the known soluble platelet agonists bind to GPCRs on the platelet surface, although these generally do not provide the initial stimulus for hemostatic plug formation in vivo. Platelets first become activated at sites of vessel injury by a shear-dependent interaction with von Willebrand factor (vWF) and collagen exposed beneath the damaged endothelial lining [2Jackson S.P. Nesbitt W.S. Kulkarni S. Signaling events underlying thrombus formation.J Thromb Haemost. 2003; 1: 1602-12Abstract Full Text Full Text PDF PubMed Scopus (337) Google Scholar, 3Savage B. Almus-Jacobs F. Ruggeri Z.M. Specific synergy of multiple substrate-receptor interactions in platelet thrombus formation under flow.Cell. 1998; 94: 657-66Abstract Full Text Full Text PDF PubMed Scopus (678) Google Scholar, 4Badimon L. Badimon J.J. Turitto V.T. Vallabhajosula S. Fuster V. Platelet thrombus formation on collagen type I. A model of deep vessel injury. Influence of blood rheology, von Willebrand factor, and blood coagulation.Circulation. 1988; 78: 1431-42Crossref PubMed Google Scholar]. vWF and collagen induce platelet signaling and adhesion to the subendothelial matrix, which is immediately followed by the generation of thrombin on the platelet surface [7Dorsam R.T. Tuluc M. Kunapuli S.P. Role of protease-activated and ADP receptor subtypes in thrombin generation on human platelets.J Thromb Haemost. 2004; 2: 804-12Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar], the generation of thromboxane A2 from arachidonic acid [8FitzGerald G.A. Mechanisms of platelet activation: thromboxane A2 as an amplifying signal for other agonists.Am J Cardiol. 1991; 68: 11B-15BAbstract Full Text PDF PubMed Google Scholar, 9Nakamura T. Jamieson G.A. Okuma M. Kambayashi J. Tandon N.N. Platelet adhesion to native type I collagen fibrils. Role of GPVI in divalent cation-dependent and -independent adhesion and thromboxane A2 generation.J Biol Chem. 1998; 273: 4338-44Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 10Hanasaki K. Nakano T. Arita H. Two phasic generation of thromboxane A2 by the action of collagen on rat platelets.Thromb Res. 1987; 46: 425-36Abstract Full Text PDF PubMed Scopus (0) Google Scholar], and the release of ADP from platelet dense granules [11Reed G.L. Platelet secretory mechanisms.Semin Thromb Hemost. 2004; 30: 441-50Crossref PubMed Scopus (0) Google Scholar, 12Legrand C. Nurden A.T. Studies on platelets of patients with inherited platelet disorders suggest that collagen-induced fibrinogen binding to membrane receptors requires secreted ADP but not released alpha-granule proteins.Thromb Haemost. 1985; 54: 603-6Crossref PubMed Scopus (0) Google Scholar]. Each of these soluble agonists binds to GPCRs [13Woulfe D. Yang J. Prevost N. O'Brien P. Fortna R. Tognolini M. Jiang H. Wu J. Brass L.F. Signaling receptors on platelets and megakaryocytes.Methods Mol Biol. 2004; 273: 3-32PubMed Google Scholar, 14Vu T.K. Hung D.T. Wheaton V.I. Coughlin S.R. Molecular cloning of a functional thrombin receptor reveals a novel proteolytic mechanism of receptor activation.Cell. 1991; 64: 1057-68Abstract Full Text PDF PubMed Scopus (2654) Google Scholar, 15Hirata M. Hayashi Y. Ushikubi F. Yokota Y. Kageyama R. Nakanishi S. Narumiya S. Cloning and expression of cDNA for a human thromboxane A2 receptor.Nature. 1991; 349: 617-20Crossref PubMed Google Scholar, 16Hollopeter G. Jantzen H.M. Vincent D. Li G. England L. Ramakrishnan V. Yang R.B. Nurden P. Nurden A. Julius D. Conley P.B. Identification of the platelet ADP receptor targeted by antithrombotic drugs.Nature. 2001; 409: 202-7Crossref PubMed Scopus (1280) Google Scholar, 17Webb T.E. Simon J. Krishek B.J. Bateson A.N. Smart T.G. King B.F. Burnstock G. Barnard E.A. Cloning and functional expression of a brain G-protein-coupled ATP receptor.FEBS Lett. 1993; 324: 219-25Crossref PubMed Scopus (464) Google Scholar]. The activation of GPCRs by platelet agonists in turn initiates multiple signaling cascades that enhance secretion of platelet granule contents and trigger fibrinogen binding to integrin αIIbβ3 [18Shattil S.J. Newman P.J. Integrins: dynamic scaffolds for adhesion and signaling in platelets.Blood. 2004; 104: 1606-15Crossref PubMed Scopus (436) Google Scholar, 19De Cristofaro R. Landolfi R. De Candia E. Castagnola M. Di Cera E. Wyman J. Allosteric equilibria in the binding of fibrinogen to platelets.Proc Natl Acad Sci U S A. 1988; 85: 8473-6Crossref PubMed Google Scholar, 20Bennett J.S. Vilaire G. Exposure of platelet fibrinogen receptors by ADP and epinephrine.J Clin Invest. 1979; 64: 1393-401Crossref PubMed Google Scholar, 21Shattil S.J. Brass L.F. Induction of the fibrinogen receptor on human platelets by intracellular mediators.J Biol Chem. 1987; 262: 992-1000Abstract Full Text PDF PubMed Google Scholar]. Soluble agonists, such as ADP released from platelet dense granules, extend platelet plug formation by recruiting circulating platelets to the growing platelet plug. The platelet plug is then stabilized by additional forces, such as the crosslinking of fibrin by factor XIII and αIIbβ3-dependent ‘outside-in’ signaling [22Law D.A. DeGuzman F.R. Heiser P. Ministri-Madrid K. Killeen N. Phillips D.R. Integrin cytoplasmic tyrosine motif is required for outside-in alphaIIbbeta3 signalling and platelet function.Nature. 1999; 401: 808-11Crossref PubMed Scopus (0) Google Scholar, 23Francis C.W. Marder V.J. Rapid formation of large molecular weight alpha-polymers in cross-linked fibrin induced by high factor XIII concentrations. Role of platelet factor XIII.J Clin Invest. 1987; 80: 1459-65Crossref PubMed Google Scholar]. The recruitment of platelets by soluble agonists to enhance growth of the platelet plug is generally simulated in vitro by observing platelet aggregation in solution. Platelet aggregation can be stimulated by thrombin, thromboxane A2, epinephrine, or ADP. Binding of any of these agonists to the platelet surface results in platelet shape change within seconds, followed by aggregation over the course of several minutes. Reorganization of the actin cytoskeleton is required for platelet shape change and appears to be primarily regulated in a G protein-dependent fashion by Rho activation and phosphorylation of myosin light chain kinase (MLCK) [24Klages B. Brandt U. Simon M.I. Schultz G. Offermanns S. Activation of G12/G13 results in shape change and Rho/Rho-kinase-mediated myosin light chain phosphorylation in mouse platelets.J Cell Biol. 1999; 144: 745-54Crossref PubMed Scopus (305) Google Scholar]. Multiple signals have been shown to be required for aggregation: two of the critical changes in second messengers observed are an increase in the intracellular calcium concentration and a decrease in cytosolic cyclic adenosine monophosphate (cAMP) [25Jin J. Kunapuli S.P. Coactivation of two different G protein-coupled receptors is essential for ADP-induced platelet aggregation.Proc Natl Acad Sci U S A. 1998; 95: 8070-4Crossref PubMed Scopus (470) Google Scholar, 26Pulcinelli F.M. Ciampa M.T. Favilla M. Pignatelli P. Riondino S. Gazzaniga P.P. Concomitant activation of Gi protein-coupled receptor and protein kinase C or phospholipase C is required for platelet aggregation.FEBS Lett. 1999; 460: 37-40Crossref PubMed Scopus (0) Google Scholar]. Both of these changes are necessary for inducing a conformational change in the platelet-specific integrin, αIIbβ3, which consequently becomes competent to bind fibrinogen [18Shattil S.J. Newman P.J. Integrins: dynamic scaffolds for adhesion and signaling in platelets.Blood. 2004; 104: 1606-15Crossref PubMed Scopus (436) Google Scholar, 27Brass L.F. Manning D.R. Cichowski K. Abrams C.S. Signaling through G proteins in platelets: to the integrins and beyond.Thromb Haemost. 1997; 78: 581-9Crossref PubMed Scopus (165) Google Scholar]. The change in the integrin that allows fibrinogen binding, termed ‘inside-out signaling’ is generally accepted to be the central event in platelet aggregation, but the detailed mechanisms which lead from G protein activation to inside-out signaling are still not completely understood. Nevertheless, much progress has been made, particularly in regard to defining the receptor-dependent activation of specific G proteins in platelets, the signaling pathways stimulated by these G proteins, and whether these contribute to fibrinogen binding or reorganization of the platelet cytoskeleton [18Shattil S.J. Newman P.J. Integrins: dynamic scaffolds for adhesion and signaling in platelets.Blood. 2004; 104: 1606-15Crossref PubMed Scopus (436) Google Scholar]. All GPCRs share a common serpentine structure of seven transmembrane-spanning domains, with an extracellular N-terminus and an intracellular C-terminus [28Kroeze W.K. Sheffler D.J. Roth B.L. G-protein-coupled receptors at a glance.J Cell Sci. 2003; 116: 4867-9Crossref PubMed Scopus (254) Google Scholar, 29Dohlman H.G. Bouvier M. Benovic J.L. Caron M.G. Lefkowitz R.J. The multiple membrane spanning topography of the beta 2-adrenergic receptor. Localization of the sites of binding, glycosylation, and regulatory phosphorylation by limited proteolysis.J Biol Chem. 1987; 262: 14282-8Abstract Full Text PDF PubMed Google Scholar, 30Nathans J. Hogness D.S. Isolation, sequence analysis, and intron-exon arrangement of the gene encoding bovine rhodopsin.Cell. 1983; 34: 807-14Abstract Full Text PDF PubMed Scopus (0) Google Scholar, 31Peralta E.G. Winslow J.W. Peterson G.L. Smith D.H. Ashkenazi A. Ramachandran J. Schimerlik M.I. Capon D.J. Primary structure and biochemical properties of an M2 muscarinic receptor.Science. 1987; 236: 600-5Crossref PubMed Scopus (333) Google Scholar]. GPCRs are so-named because they are physically associated with heterotrimeric G proteins, which typically dock in the region of the third intracellular loop on the internal membrane surface [32Wong S.K. Parker E.M. Ross E.M. Chimeric muscarinic cholinergic: beta-adrenergic receptors that activate Gs in response to muscarinic agonists.J Biol Chem. 1990; 265: 6219-24Abstract Full Text PDF PubMed Google Scholar, 33Wess J. Brann M.R. Bonner T.I. Identification of a small intracellular region of the muscarinic m3 receptor as a determinant of selective coupling to PI turnover.FEBS Lett. 1989; 258: 133-6Crossref PubMed Scopus (76) Google Scholar, 34Kobilka B.K. Kobilka T.S. Daniel K. Regan J.W. Caron M.G. Lefkowitz R.J. Chimeric alpha 2-,beta 2-adrenergic receptors: delineation of domains involved in effector coupling and ligand binding specificity.Science. 1988; 240: 1310-6Crossref PubMed Google Scholar]. Each heterotrimeric G protein is composed of an α, β, and γ subunit, the β and γ subunits of which remain tightly associated. Agonist binding to the receptor initiates dissociation of the α from the βγ subunits, which allows exposure of surfaces on both the α and βγ subunits for interaction with effector proteins. G proteins are generally classified into four families based on the identity and function of the α subunits: Gs, Gi, Gq, and G12/13 [35Wettschureck N. Moers A. Offermanns S. Mouse models to study G-protein-mediated signaling.Pharmacol Ther. 2004; 101: 75-89Crossref PubMed Scopus (0) Google Scholar]. Whether specific members of the βγ family interact only with specific α subunits or with particular receptors is still not well understood and has not been explored at all in platelets. The term G protein generally refers to the heterotrimer, and for the purposes of this review, a particular heterotrimer will be represented as, for example, Gs. The α subunit in particular will be represented as Gαs: these terms, however, are often used interchangeably. Representatives of each of the four G protein families are present in platelets and are listed together with the receptors to which they couple in Table 1 [36Williams A.G. Woolkalis M.J. Poncz M. Manning D.R. Gewirtz A.M. Brass L.F. Identification of the pertussis toxin-sensitive G proteins in platelets, megakaryocytes, and human erythroleukemia cells.Blood. 1990; 76: 721-30Crossref PubMed Google Scholar, 37Gagnon A.W. Manning D.R. Catani L. Gewirtz A. Poncz M. Brass L.F. Identification of Gz alpha as a pertussis toxin-insensitive G protein in human platelets and megakaryocytes.Blood. 1991; 78: 1247-53Crossref PubMed Google Scholar, 38Shenker A. Goldsmith P. Unson C.G. Spiegel AM. The G. protein coupled to the thromboxane A2 receptor in human platelets is a member of the novel Gq family.J Biol Chem. 1991; 266: 9309-13Abstract Full Text PDF PubMed Google Scholar, 39van Willigen G. Donath J. Lapetina E.G. Akkerman J.W. Identification of alpha-subunits of trimeric GTP-binding proteins in human platelets by RT-PCR.Biochem Biophys Res Commun. 1995; 214: 254-62Crossref PubMed Scopus (0) Google Scholar]. Agonists for receptors coupled to Gs stimulate adenylyl cyclase, increasing cAMP production. A high concentration of cytosolic cAMP suppresses platelet function [40Keularts I.M. van Gorp R.M. Feijge M.A. Vuist W.M. Heemskerk J.W. Alpha(2A)-adrenergic receptor stimulation potentiates calcium release in platelets by modulating cAMP levels.J Biol Chem. 2000; 275: 1763-72Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 41Yada Y. Nagao S. Okano Y. Nozawa Y. Inhibition by cyclic AMP of guanine nucleotide-induced activation of phosphoinositide-specific phospholipase C in human platelets.FEBS Lett. 1989; 242: 368-72Crossref PubMed Scopus (43) Google Scholar], as do agonists that stimulate Gs-coupled receptors on platelets [42Yang J. Wu J. Jiang H. Mortensen R. Austin S. Manning D.R. Woulfe D. Brass L.F. Signaling through Gi family members in platelets. Redundancy and specificity in the regulation of adenylyl cyclase and other effectors.J Biol Chem. 2002; 277: 46035-42Abstract Full Text Full Text PDF PubMed Scopus (128) Google Scholar]. Activation of Gi family members inhibits adenylyl cyclase, inhibiting the accumulation of cAMP. Activation of Gi-coupled receptors relieves the tonic suppression of function mediated by high cytosolic concentrations of cAMP [40Keularts I.M. van Gorp R.M. Feijge M.A. Vuist W.M. Heemskerk J.W. Alpha(2A)-adrenergic receptor stimulation potentiates calcium release in platelets by modulating cAMP levels.J Biol Chem. 2000; 275: 1763-72Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 42Yang J. Wu J. Jiang H. Mortensen R. Austin S. Manning D.R. Woulfe D. Brass L.F. Signaling through Gi family members in platelets. Redundancy and specificity in the regulation of adenylyl cyclase and other effectors.J Biol Chem. 2002; 277: 46035-42Abstract Full Text Full Text PDF PubMed Scopus (128) Google Scholar] and activates additional signaling pathways that are still being elucidated [40Keularts I.M. van Gorp R.M. Feijge M.A. Vuist W.M. Heemskerk J.W. Alpha(2A)-adrenergic receptor stimulation potentiates calcium release in platelets by modulating cAMP levels.J Biol Chem. 2000; 275: 1763-72Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 43Lova P. Paganini S. Hirsch E. Barberis L. Wymann M. Sinigaglia F. Balduini C. Torti M. A selective role for phosphatidylinositol 3,4, 5-trisphosphate in the Gi-dependent activation of platelet Rap1B.J Biol Chem. 2003; 278: 131-8Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 44Kauffenstein G. Bergmeier W. Eckly A. Ohlmann P. Leon C. Cazenave J.P. Nieswandt B. Gachet C. The P2Y(12) receptor induces platelet aggregation through weak activation of the alpha(IIb) beta(3) integrin – a phosphoinositide 3-kinase-dependent mechanism.FEBS Lett. 2001; 505: 281-90Crossref PubMed Scopus (0) Google Scholar]. Gq activation stimulates the activity of phospholipase Cβ, which catalyzes the release of diacylglycerol (DAG) and inositol tris-phosphate (IP3) from phosphatidylinositol-(4,5)-bisphosphate (PIP2). DAG and IP3 in turn activate protein kinase C (PKC) and stimulate the release of calcium from the dense tubular system [45Offermanns S. Toombs C.F. Hu Y.H. Simon M.I. Defective platelet activation in G alpha(q)-deficient mice.Nature. 1997; 389: 183-6Crossref PubMed Scopus (494) Google Scholar]. Activation of the G12/13 family in platelets stimulates activation of the small GTPase RhoA [24Klages B. Brandt U. Simon M.I. Schultz G. Offermanns S. Activation of G12/G13 results in shape change and Rho/Rho-kinase-mediated myosin light chain phosphorylation in mouse platelets.J Cell Biol. 1999; 144: 745-54Crossref PubMed Scopus (305) Google Scholar]. RhoA and Rho kinase (RhoK) activation have in turn been shown to stimulate phosphorylation of MLCK, which is proposed to regulate the reorganization of the actin cytoskeleton associated with platelet shape change [24Klages B. Brandt U. Simon M.I. Schultz G. Offermanns S. Activation of G12/G13 results in shape change and Rho/Rho-kinase-mediated myosin light chain phosphorylation in mouse platelets.J Cell Biol. 1999; 144: 745-54Crossref PubMed Scopus (305) Google Scholar].Table 1G protein-coupled receptors on mouse or human plateletsReceptorAgonistG proteinEffector/signalingReferencesPAR1 (human only)ThrombinGqPhospholipase C/Ca++ release, PKC activation36Williams A.G. Woolkalis M.J. Poncz M. Manning D.R. Gewirtz A.M. Brass L.F. Identification of the pertussis toxin-sensitive G proteins in platelets, megakaryocytes, and human erythroleukemia cells.Blood. 1990; 76: 721-30Crossref PubMed Google Scholar, 37Gagnon A.W. Manning D.R. Catani L. Gewirtz A. Poncz M. Brass L.F. Identification of Gz alpha as a pertussis toxin-insensitive G protein in human platelets and megakaryocytes.Blood. 1991; 78: 1247-53Crossref PubMed Google Scholar, 38Shenker A. Goldsmith P. Unson C.G. Spiegel AM. The G. protein coupled to the thromboxane A2 receptor in human platelets is a member of the novel Gq family.J Biol Chem. 1991; 266: 9309-13Abstract Full Text PDF PubMed Google ScholarG13Rho activation, actin remodelingPAR4ThrombinGqPhospholipase C/Ca++release, PKC activation38Shenker A. Goldsmith P. Unson C.G. Spiegel AM. The G. protein coupled to the thromboxane A2 receptor in human platelets is a member of the novel Gq family.J Biol Chem. 1991; 266: 9309-13Abstract Full Text PDF PubMed Google Scholar, 39van Willigen G. Donath J. Lapetina E.G. Akkerman J.W. Identification of alpha-subunits of trimeric GTP-binding proteins in human platelets by RT-PCR.Biochem Biophys Res Commun. 1995; 214: 254-62Crossref PubMed Scopus (0) Google Scholar, 40Keularts I.M. van Gorp R.M. Feijge M.A. Vuist W.M. Heemskerk J.W. Alpha(2A)-adrenergic receptor stimulation potentiates calcium release in platelets by modulating cAMP levels.J Biol Chem. 2000; 275: 1763-72Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 41Yada Y. Nagao S. Okano Y. Nozawa Y. Inhibition by cyclic AMP of guanine nucleotide-induced activation of phosphoinositide-specific phospholipase C in human platelets.FEBS Lett. 1989; 242: 368-72Crossref PubMed Scopus (43) Google ScholarG13Rho activation, actin remodelingPAR3 (mouse only)ThrombinNoneNone39van Willigen G. Donath J. Lapetina E.G. Akkerman J.W. Identification of alpha-subunits of trimeric GTP-binding proteins in human platelets by RT-PCR.Biochem Biophys Res Commun. 1995; 214: 254-62Crossref PubMed Scopus (0) Google Scholar, 40Keularts I.M. van Gorp R.M. Feijge M.A. Vuist W.M. Heemskerk J.W. Alpha(2A)-adrenergic receptor stimulation potentiates calcium release in platelets by modulating cAMP levels.J Biol Chem. 2000; 275: 1763-72Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 41Yada Y. Nagao S. Okano Y. Nozawa Y. Inhibition by cyclic AMP of guanine nucleotide-induced activation of phosphoinositide-specific phospholipase C in human platelets.FEBS Lett. 1989; 242: 368-72Crossref PubMed Scopus (43) Google ScholarP2Y1ADPGqPhospholipase C Ca++ release, PKC activation50–53P2Y12ADPGi2↓ cAMP, PI3Kγ activation27Brass L.F. Manning D.R. Cichowski K. Abrams C.S. Signaling through G proteins in platelets: to the integrins and beyond.Thromb Haemost. 1997; 78: 581-9Crossref PubMed Scopus (165) Google Scholar, 54Yamanishi J. Kawahara Y. Fukuzaki H. Effect of cyclic AMP on cytoplasmic free calcium in human platelets stimulated by thrombin: direct measurement with quin2.Thromb Res. 1983; 32: 183-8Abstract Full Text PDF PubMed Scopus (0) Google Scholar, 58Nanevicz T. 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Molecular basis for ADP-induced platelet activation. II. The P2Y1 receptor mediates ADP-induced intracellular calcium mobilization and shape change in platelets.J Biol Chem. 1998; 273: 2030-4Abstract Full Text Full Text PDF PubMed Scopus (412) Google ScholarIPPGI2Gs↑ cAMP25Jin J. Kunapuli S.P. Coactivation of two different G protein-coupled receptors is essential for ADP-induced platelet aggregation.Proc Natl Acad Sci U S A. 1998; 95: 8070-4Crossref PubMed Scopus (470) Google Scholar, 29Dohlman H.G. Bouvier M. Benovic J.L. Caron M.G. Lefkowitz R.J. The multiple membrane spanning topography of the beta 2-adrenergic receptor. Localization of the sites of binding, glycosylation, and regulatory phosphorylation by limited proteolysis.J Biol Chem. 1987; 262: 14282-8Abstract Full Text PDF PubMed Google Scholar, 30Nathans J. Hogness D.S. Isolation, sequence analysis, and intron-exon arrangement of the gene encoding bovine rhodopsin.Cell. 1983; 34: 807-14Abstract Full Text PDF PubMed Scopus (0) Google Scholar, 34Kobilka B.K. Kobilka T.S. Daniel K. Regan J.W. Caron M.G. Lefkowitz R.J. Chimeric alpha 2-,beta 2-adrenergic receptors: delineation of domains involved in effector coupling and ligand binding specificity.Science. 1988; 240: 1310-6Crossref PubMed Google ScholarTPThromboxane A2GqPhospholipase C, Ca++ release, PKC activation69–80G13Rho activation, actin remodelingα2A adrenergicEpinephrineGz↓ cAMP23Francis C.W. Marder V.J. Rapid formation of large molecular weight alpha-polymers in cross-linked fibrin induced by high factor XIII concentrations. Role of platelet factor XIII.J Clin Invest. 1987; 80: 1459-65Crossref PubMed Google Scholar, 81Bertoni A. Tadokoro S. Eto K. Pampori N. Parise L.V. White G.C. Shattil S.J. 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Akt activation in platelets depends on Gi signaling pathways.J Biol Chem. 2004; 279: 4186-95Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar Open table in a new tab To date, all the activity of GPCRs in platelets has been attributed to G protein-dependent pathways, as the knock-out of individual G proteins has been sufficient to disrupt platelet responses to receptor agonists [45Offermanns S. Toombs C.F. Hu Y.H. Simon M.I. Defective platelet activation in G alpha(q)-deficient mice.Nature. 1997; 389: 183-6Crossref PubMed Scopus (494) Google Scholar, 46Moers A. Nieswandt B. Massberg S. Wettschureck N. Gruner S. Konrad I. Schulte V. Aktas B. Gratacap M.P. Simon M.I. Gawaz M. Offermanns S. G13 is an essential mediator of platelet activation in hemostasis and thrombosis.Nat Med. 2003; 9: 1418-22Crossref PubMed Scopus (212) Google Scholar, 47Jantzen H.M. Milstone D.S. Gousset L. Conley P.B. Mortensen R.M. Impaired activation of murine platelets lacking G alpha(i2).J Clin Invest. 2001; 108: 477-83Crossref PubMed Google Scholar]. Most platelet agonists bind to GPCRs that activate or enhance activation of platelets (see specific examples in the following sections). An exception to this is the prostacyclin PGI2, which binds to the IP receptor and dampens platelet responses to other agonists by increasing cytosolic cAMP concentrations [48Eggerman T.L. Hartzell C.J. Selfe S. Andersen N.H. The single prostacyclin receptor of gel-filtered platelets provides a correlation with antiaggregatory potency of PGI2 mimics.Thromb Res. 1987; 45: 645-59Abstract Full Text PDF PubMed Google Scholar]. Platelet agonists may bind to a single GPCR or to multiple receptors, and the receptors in turn may couple to one or more G proteins (see below). Our understanding of the impact of individual GPCRs to platelet function has developed as the receptors have been cloned and as subtype-selective agonists and antagonists have been developed, and has accelerated with the study of receptor knock-outs in mice. In several cases, the impact of a particular receptor to human health has also become apparent, as patients with bleeding defects have been identified and the defect attributed to functional deficiencies in a particular receptor or pathway [16Hollopeter G. Jantzen H.M. Vincent D. Li G. England L. Ramakrishnan V. Yang R.B. Nurden P. Nurden A. Julius D. Conley P.B. Identification of the platelet ADP receptor targeted by antithrombotic drugs.Nature. 2001; 409: 202-7Crossref PubMed Scopus (1280) Google Scholar, 49Cattaneo M. Zighetti M.L. Lombardi R. Martinez C. Lecchi A. Conley P.B. Ware J. Ruggeri Z.M. Molecular bases of defective signal transduction in the platelet P2Y12 receptor of a patient with congenital bleeding.Proc Natl Acad Sci U S A. 2003; 100: 1978-83Crossref PubMed Scopus (0) Google Scholar, 50Mitsui T. Yokoyama S. Shimizu Y. Katsuura M. Akiba K. Hayasaka K. Defective signal transduction through the thromboxane A2 receptor in a patient with a mild bleeding disorder: deficiency of the inositol 1,4,5-triphosphate formation despite normal G-protein activation.Thromb Haemost. 1997; 77: 991-5Crossref PubMed Google Scholar, 51Fuse I. Higuchi W. Aizawa Y. Pathogenesis of a bleeding disorder characterized by platelet unresponsiveness to thromboxane A2.Semin Thromb Hemost. 2000; 26: 43-5Crossref PubMed Google Scholar]. In the following sections, I will describe the role of individual receptors in the signal transduction pathways initiated by the more common platelet agonists (and one platelet inactivator). A schematic representation of platelet signaling stimulated by agonist binding to GPCRs is provided in Fig. 1. Platelets circulating through the bloodstream are normally expo