The G-protein βγ subunits regulate platelet function
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AIMS: G-protein coupled receptors (GPCRs) tightly regulate platelet function by interacting with various physiological agonists. An essential mediator of GPCR signaling is the G protein αβγ heterotrimers, in which the βγ subunits are central players in downstream signaling. Herein, we investigated the role of Gβγ subunits in platelet function, hemostasis and thrombogenesis. METHODS: To achieve this goal, platelets from both mice and humans were employed in the context of a small molecule inhibitor of Gβγ, namely gallein. We used an aggregometer to examine aggregation and dense granules secretion. We also used flow cytometry for P-selectin and PAC1 to determine the impact of inhibiting Gβγ on α -granule secretion and αIIbβ3 activation. Clot retraction and the platelet spreading assay were used to examine Gβγ role in outside-in platelet signaling, whereas Western blot was employed to examine its role in Akt activation. Finally, we used the bleeding time assay and the FeCl3-induced carotid-artery injury thrombosis model to determine Gβγ contribution to in vivo platelet function. RESULTS: We observed that gallein inhibits platelet aggregation and secretion in response to agonist stimulation, in both mouse and human platelets. Furthermore, gallein also exerted inhibitory effects on integrin αIIbβ3 activation, clot retraction, platelet spreading and Akt activation/phosphorylation. Finally, gallein's inhibitory effects manifested in vivo, as documented by its ability to modulate physiological hemostasis and delay thrombus formation. CONCLUSION: Our findings demonstrate, for the first time, that Gβγ subunits directly regulate GPCR-dependent platelet function, in vitro and in vivo. Moreover, these data highlight Gβγ as a novel therapeutic target for managing thrombotic disorders.
author list (cited authors)
Alarabi, A. B., Karim, Z. A., Hinojos, V., Lozano, P. A., Hernandez, K. R., Montes Ramirez, J. E., ... Alshbool, F. Z.