Polgár J, Clemetson J M, Kehrel B E, Wiedemann M, Magnenat E M, Wells T N, Clemetson K J
Theodor Kocher Institute, University of Berne, Freiestrasse 1, CH-3012 Berne, Switzerland.
J Biol Chem. 1997 May 23;272(21):13576-83. doi: 10.1074/jbc.272.21.13576.
Convulxin, a powerful platelet activator, was isolated from Crotalus durissus terrificus venom, and 20 amino acid N-terminal sequences of both subunits were determined. These indicated that convulxin belongs to the heterodimeric C-type lectin family. Neither antibodies against GPIb nor echicetin had any effect on convulxin-induced platelet aggregation showing that, in contrast to other venom C-type lectins acting on platelets, GPIb is not involved in convulxin-induced platelet activation. In addition, partially reduced/denatured convulxin only affects collagen-induced platelet aggregation. The mechanism of convulxin-induced platelet activation was examined by platelet aggregation, detection of time-dependent tyrosine phosphorylation of platelet proteins, and binding studies with 125I-convulxin. Convulxin induces signal transduction in part like collagen, involving the time-dependent tyrosine phosphorylation of Fc receptor gamma chain, phospholipase Cgamma2, p72(SYK), c-Cbl, and p36-38. However, unlike collagen, pp125(FAK) and some other bands are not tyrosine-phosphorylated. Convulxin binds to a glycosylated 62-kDa membrane component in platelet lysate and to p62/GPVI immunoprecipitated by human anti-p62/GPVI antibodies. Convulxin subunits inhibit both aggregation and tyrosine phosphorylation in response to collagen. Piceatannol, a tyrosine kinase inhibitor with some specificity for p72(SYK), showed differential effects on collagen and convulxin-stimulated signaling. These results suggest that convulxin uses the p62/GPVI but not the alpha2beta1 part of the collagen signaling pathways to activate platelets. Occupation and clustering of p62/GPVI may activate Src family kinases phosphorylating Fc receptor gamma chain and, by a mechanism previously described in T- and B-cells, activate p72(SYK) that is critical for downstream activation of platelets.
convulxin是一种强效血小板激活剂，从巴西矛头蝮蛇毒液中分离得到，并测定了两个亚基的20个氨基酸N端序列。这些结果表明convulxin属于异源二聚体C型凝集素家族。抗糖蛋白Ib（GPIb）抗体和echicetin对convulxin诱导的血小板聚集均无任何影响，这表明与其他作用于血小板的蛇毒C型凝集素相反，GPIb不参与convulxin诱导的血小板激活。此外，部分还原/变性的convulxin仅影响胶原诱导的血小板聚集。通过血小板聚集、检测血小板蛋白的时间依赖性酪氨酸磷酸化以及与125I-convulxin的结合研究，对convulxin诱导血小板激活的机制进行了研究。convulxin部分地像胶原一样诱导信号转导，涉及Fc受体γ链、磷脂酶Cγ2、p72（SYK）、c-Cbl和p36 - 38的时间依赖性酪氨酸磷酸化。然而，与胶原不同的是，pp125（黏着斑激酶）和其他一些条带未发生酪氨酸磷酸化。convulxin与血小板裂解物中一种糖基化的62 kDa膜成分以及人抗p62/GPVI抗体免疫沉淀的p62/GPVI结合。convulxin亚基抑制胶原诱导的聚集和酪氨酸磷酸化。白皮杉醇是一种对p72（SYK）具有一定特异性的酪氨酸激酶抑制剂，对胶原和convulxin刺激的信号传导表现出不同的作用。这些结果表明，convulxin利用胶原信号通路的p62/GPVI而非α2β1部分来激活血小板。p62/GPVI的占据和聚集可能激活Src家族激酶，使其磷酸化Fc受体γ链，并通过先前在T细胞和B细胞中描述的机制激活对血小板下游激活至关重要的p72（SYK）。
