Extracellular signal-regulated kinase 5 associates with casein kinase II to regulate GPIb-IX-mediated platelet activation via the PTEN/PI3K/Akt pathway

摘要

Background: The platelet glycoprotein (GP) Ib-IX complex plays essential roles in thrombosis and hemostasis. The mitogen-activated protein kinases (MAPKs) ERK1/2 and p38 have been shown to be important in the GPIb-IX-mediated signaling leading to integrin activation. However, the roles of the MAPK extracellular signal-regulated kinase 5 (ERK5) in GPIb-IX-mediated platelet activation are unknown. Objective: To reveal the function and mechanisms of ERK5 in GPIb-IX-mediated platelet activation. Methods: The functions of ERK5 in GPIb-IX-mediated human platelet activation were assessed using botrocetin/VWF, ristocetin/VWF, or platelet adhesion to von Willebrand factor (VWF) under shear stress in the presence of a specific inhibitor of ERK5. ERK5-associated proteins were pulled down from Chinese hamster ovary (CHO) cells transfected with HA-tagged-ERK5, identified by mass spectrometry, and confirmed in human platelets. Roles of ERK5-associated proteins in GPIb-IX-mediated platelet activation were clarified using specific inhibitors. Results: The phosphorylation levels of ERK5 were significantly enhanced in human platelets stimulated with botrocetin/VWF or ristocetin/VWF. The ERK5 inhibitor XMD8-92 suppressed the second wave of human platelet aggregation induced by botrocetin/VWF or ristocetin/VWF and inhibited human platelet adhesion on immobilized VWF under shear stress. Casein kinase II (CKII) was identified as an ERK5-associated protein in human platelets. The CKII inhibitor TBB, similar to the ERK5 inhibitor XMD8-92, specifically restrained PTEN phosphorylation, therefore suppressing Akt phosphorylation in human platelets treated with botrocetin/VWF. Conclusion: ERK5 associates with CKII to play essential roles in GPIb-IX-mediated platelet activation via the PTEN/PI3K/Akt pathway.