Leukemia-associated Rho guanine nucleotide exchange factor (LARG) plays an agonist specific role in platelet function through RhoA activation

We studied the of LARG in murine and human megakaryocytes and platelets with Larg knockout, shRNA-mediated knockdown and small molecule-mediated inhibition. We found that LARG is important for human, but not murine, megakaryocyte maturation. Larg KO mice exhibit macrothrombocytopenia, internal bleeding in the ovaries and prolonged bleeding times. KO platelets have impaired aggregation, α -granule release and integrin α 2b β 3 activation in response to thrombin and thromboxane, but not to ADP. The same agonist-specific reductions in platelet aggregation occur in human platelets treated with a LARG inhibitor. Larg KO platelets have reduced RhoA activation and myosin light chain phosphorylation, suggesting that Larg plays an agonist-specific role in platelet signal transduction. Using 2 different in vivo assays, Larg KO mice are protected from in vivo thrombus formation. Together, these results establish that LARG regulates human megakaryocyte maturation, and is critical for platelet function in both humans and mice. that LARG deficiency or inhibition would affect thrombin-induced platelet activation by decreasing thrombin-induced RhoA activation. We assessed the role of LARG in platelet function by characterizing the in vitro and in vivo megakaryocyte and platelet phenotypes of Larg KO mice, and by assessing human megakaryocyte (MK) and platelet function in the presence of a LARG inhibitor. Murine MK were assessed in Larg KO mice and human megakaryopoiesis was assessed in vitro using shRNA and inhibitor treatments. We found that LARG is important for human MK maturation, but Larg KO mice had no detectable differences in megakaryopoiesis. However, LARG appears to play a role in activation of both murine and human platelets specifically in response to thrombin and thromboxane via activation of RhoA signaling. Moreover, LARG was shown to play an important role in two in vivo murine thrombosis models, carotid artery ligation and FeCl 3 induced carotid damage.