title: P2X1-mediated ERK2 activation amplifies the collagen-induced platelet secretion by enhancing myosin light chain kinase activation.
abstract: The ATP-gated P2X1 ion channel is the only P2X subtype expressed in human platelets. Via transmission electron microscopy, we found that P2X1 mediates fast, reversible platelet shape change, secretory granule centralization, and pseudopodia formation. In washed human platelets, the stable P2X1 agonist alpha,beta-methylene ATP (alpha,beta-meATP) causes rapid, transient (2-5 s), and dose-dependent myosin light chain (MLC) phosphorylation, requiring extracellular Ca2+. Phosphorylation was inhibited by the calmodulin (CaM) inhibitor W-7, but not by the Rho kinase inhibitor HA-1077, i.e. it is exclusively regulated by Ca2+/CaM-dependent MLC kinase. Correspondingly, the P2X1-induced platelet shape change was inhibited by W-7 and by the MLC kinase inhibitor ML-7 but not by HA-1077. W-7, ML-7, the protein kinase C inhibitor GF109203-X, and the Src family kinase inhibitor PP1 inhibited the collagen and convulxin-induced early platelet degranulation, shape change, and subsequent aggregation, indicating a role for Ca2+/CaM and MLC kinase in these glycoprotein VI-related platelet responses. The secreted ATP-mediated P2X1-dependent ERK2 activation induced by low collagen concentrations contributes to MLC kinase activation since P2X1 desensitization or blockade of ERK2 phosphorylation by U0126 strongly attenuated MLC phosphorylation, degranulation, and aggregation. We therefore conclude that at low doses of collagen, glycoprotein VI activation leads to early protein kinase C- and MLC kinase-dependent degranulation. Rapidly released ATP triggers P2X1 -mediated Ca2+ influx, activating ERK2, in turn amplifying platelet secretion by reinforcing the early MLC kinase phosphorylation. Hence, the P2X1-ERK2-MLC axis contributes to collagen-induced platelet activation by enhancing platelet degranulation.
mesh_terms: D000255:Adenosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D001792:Blood Platelets / Q000166:cytology / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D015550:Cell Degranulation; D048429:Cell Size; D003094:Collagen / Q000494:pharmacology; D004789:Enzyme Activation; D006801:Humans; D008854:Microscopy, Electron; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D058910:Purinergic P2 Receptor Agonists; D018048:Receptors, Purinergic P2 / Q000502:physiology*; D058469:Receptors, Purinergic P2X
chemical_list: D002147:Calmodulin; D058910:Purinergic P2 Receptor Agonists; D018048:Receptors, Purinergic P2; D058469:Receptors, Purinergic P2X; D000255:Adenosine Triphosphate; D003094:Collagen; D009219:Myosin-Light-Chain Kinase; D019950:Mitogen-Activated Protein Kinase 1; C002630:alpha,beta-methyleneadenosine 5'-triphosphate; D002118:Calcium