title: Arrestin-2 differentially regulates PAR4 and ADP receptor signaling in platelets.
abstract: Arrestins can facilitate desensitization or signaling by G protein-coupled receptors (GPCR) in many cells, but their roles in platelets remain uncharacterized. Because of recent reports that arrestins can serve as scaffolds to recruit phosphatidylinositol-3 kinases (PI3K)s to GPCRs, we sought to determine whether arrestins regulate PI3K-dependent Akt signaling in platelets, with consequences for thrombosis. Co-immunoprecipitation experiments demonstrate that arrestin-2 associates with p85 PI3Kα/β subunits in thrombin-stimulated platelets, but not resting cells. The association is inhibited by inhibitors of P2Y12 and Src family kinases (SFKs). The function of arrestin-2 in platelets is agonist-specific, as PAR4-dependent Akt phosphorylation and fibrinogen binding were reduced in arrestin-2 knock-out platelets compared with WT controls, but ADP-stimulated signaling to Akt and fibrinogen binding were unaffected. ADP receptors regulate arrestin recruitment to PAR4, because co-immunoprecipitates of arrestin-2 with PAR4 are disrupted by inhibitors of P2Y1 or P2Y12. P2Y1 may regulate arrestin-2 recruitment to PAR4 through protein kinase C (PKC) activation, whereas P2Y12 directly interacts with PAR4 and therefore, may help to recruit arrestin-2 to PAR4. Finally, arrestin2(-/-) mice are less sensitive to ferric chloride-induced thrombosis than WT mice, suggesting that arrestin-2 can regulate thrombus formation in vivo. In conclusion, arrestin-2 regulates PAR4-dependent signaling pathways, but not responses to ADP alone, and contributes to thrombus formation in vivo.
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, Delaware 19716, USA.
authors.forename: Dongjun
authors.initials: D
authors.lastname: Li
authors.forename: Lauren
authors.initials: L
authors.lastname: D'Angelo
authors.forename: Massiel
authors.initials: M
authors.lastname: Chavez
authors.forename: Donna S
authors.initials: DS
authors.lastname: Woulfe
mesh_terms: D000818:Animals; D019390:Arrestins / Q000502:physiology*; D001792:Blood Platelets / Q000378:metabolism*; D058543:Class Ia Phosphatidylinositol 3-Kinase / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D021381:Protein Transport; D044462:Receptors, Proteinase-Activated / Q000378:metabolism*; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D015398:Signal Transduction; D013927:Thrombosis / Q000209:etiology; D000071557:beta-Arrestins
chemical_list: D019390:Arrestins; D044462:Receptors, Proteinase-Activated; D018048:Receptors, Purinergic P2; D000071557:beta-Arrestins; C481942:protease-activated receptor 4, mouse; D058543:Class Ia Phosphatidylinositol 3-Kinase