Platelet-Derived ERp57 Mediates Platelet Incorporation Into a Growing Thrombus By Regulation Of The αIIbβ3 Integrin

ERp57 is a member of protein disulfide isomerase (PDI) family. Although previous studies using antibodies have indicated ERp57 mediates platelet aggregation, the specific role of platelet-derived ERp57 in hemostasis and thrombosis is unknown. In this study we addressed this issue using a megakaryocyte/platelet specific ERp57-knockout mouse model (PF4-Cre/ERp57^fl/fl). Although PF4-Cre/ERp57^fl/fl mice had normal platelet counts and platelet glycoproteins expression, they had significantly prolonged tail-bleeding times, and thrombus occlusion times with FeCl3-induced carotid artery-injury. Using a mesenteric artery thrombosis model we found decreased incorporation of ERp57-deficient platelets into a growing thrombus. Upon stimulation with convulxin and thrombin, platelets lacking ERp57 had defective activation of the αIIbβ3 integrin and platelet aggregation. Addition of exogenous wild type ERp57 corrected the defect in aggregation implicating surface ERp57 in platelet aggregation. Moreover, using recombinant ERp57 mutants we found that the second active site of ERp57 targets a platelet surface substrate to potentiate platelet aggregation. Binding of Alexa 488-labeled ERp57 to thrombin-activated and Mn²⁺-treated platelets lacking β3 was substantially decreased, suggesting a direct interaction of ERp57 with αIIbβ3. Surface ERp57 activity in human platelets increased in response to stimulation with thrombin, concomitant with protein expression occurring in a physiologically relevant time frame. In conclusion, platelet-derived ERp57 directly interacts with αIIbβ3 during activation of this receptor and is required for incorporation of platelets into a growing thrombus.