Integrin β3E726 Regulates the Switch Between Platelet Spreading and Clot Retraction by Interfering Gα13/RhoA Pathway

Platelet spreading and clot retraction, albeit both mediated by integrin outside-in signaling, lead to platelet shape changes in two opposite directions. The mechanisms by which these processes are regulated are not fully understood. Our previous study found that E726Q mutation in β3 integrin caused impaired spreading in Chinese hamster ovary (CHO) cells on immobilized fibrinogen.The current study further utilized knock-in mice bearing the β3^E726Q mutation to explore the underlying mechanisms whereby the E⁷²⁶ residue differentially influences platelet spreading and clot retraction.Compared to wild type (WT) platelets, β3^E726Q platelets displayed similar level of β3 expression but partially impaired fibrinogen binding associated with attenuated responses in platelet aggregation and P-selectin exposure. Notably, β3^E726Q mutation resulted in defective platelet spreading but accelerated clot retraction concomitant with increased clot density. Functionally, β3^E726Q mice displayed prolonged bleeding time and defective thrombogenesis in vitro and in vivo. Further mechanistic study showed that in β3^E726Q platelets the activities of RhoA and Rac1 were significantly enhanced following thrombin stimulation, possibly due to reduced binding of Gα13 to the β3 cytoplasmic tail.Taken together, the β3E⁷²⁶ is a potential novel regulatory site that influences the direct interaction of β3 cytoplasmic tail with Gα13 and therefore the activity of downstream RhoA, a molecular switch that shifts platelet spreading into clot retraction.