Interaction between von Willebrand factor and glycoprotein Ib activates Src kinase in human platelets: role of phosphoinositide 3-kinase
Interaction between von Willebrand factor and glycoprotein Ib activates Src kinase in human platelets: role of phosphoinositide 3-kinase
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DOI：
10.1182/blood-2002-03-0806
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发表时间：
2003-05
期刊：
影响因子：
20.3
通讯作者：
Yi Wu;N. Asazuma;K. Satoh;Y. Yatomi;T. Takafuta;M. Berndt;Y. Ozaki
中科院分区：
文献类型：
--
作者：
Yi Wu;N. Asazuma;K. Satoh;Y. Yatomi;T. Takafuta;M. Berndt;Y. Ozaki
血管性血友病因子（VWF）与糖蛋白（GP）Ib-IX-V的结合刺激导致血小板粘附和聚集的跨膜信号传导事件。最近的研究表明，Src家族激酶的激活参与了GPIb介导的血小板活化，尽管相关的信号转导途径仍然不清楚。这项研究提供了Src和GPIb协会的重要作用的证据。在含有Complete（一种广谱蛋白酶抑制剂混合物）的血小板裂解液中，Src和林恩在VWF-肉毒素刺激下与GPIb动态相关。细胞松弛素D，抑制Src激酶易位到细胞骨架，进一步增加Src和GPIb协会。用肉毒素和单体A1结构域而不是完整的VWF获得了类似的结果，诱导Src活化和GPIb与Src之间的缔合。这些发现表明，GPIb的配体结合，没有受体聚集，足以激活Src。免疫沉淀研究表明，Src，磷脂酰肌醇3-激酶（PI 3-激酶），和GPIb在GPIb刺激的血小板形成复合物。当PI 3-激酶的p85亚基被免疫耗竭时，Src与GPIb的结合被废除。然而，wortmannin，一个特定的PI 3-激酶抑制剂，未能阻止Src和GPIb之间的复合物的形成。Src-SH 3结构域作为谷胱甘肽S-转移酶（GST）-融合蛋白共沉淀PI 3-激酶的p85亚基和GPIb。这些发现一起表明，PI 3-激酶的p85亚基介导GPIb相关的激活信号和激活Src独立的PI 3-激酶的酶活性。
The binding of von Willebrand factor (VWF) to glycoprotein (GP) Ib-IX-V stimulates transmembrane signaling events that lead to platelet adhesion and aggregation. Recent studies have implied that activation of Src family kinases is involved in GPIb-mediated platelet activation, although the related signal transduction pathway remains poorly defined. This study presents evidence for an important role of Src and GPIb association. In platelet lysates containing Complete, a broad-spectrum protease inhibitor mixture, Src and Lyn dynamically associated with GPIb on VWF-botrocetin stimulation. Cytochalasin D, which inhibits translocation of Src kinases to the cytoskeleton, further increased Src and GPIb association. Similar results were obtained with botrocetin and monomeric A1 domain, instead of intact VWF, with induction of both Src activation and association between GPIb and Src. These findings suggest that ligand binding of GPIb, without receptor clustering, is sufficient to activate Src. Immunoprecipitation studies demonstrated that Src, phosphoinositide 3– kinase (PI 3–kinase), and GPIb form a complex in GPIb-stimulated platelets. When the p85 subunit of PI 3–kinase was immunodepleted, association of Src with GPIb was abrogated. However, wortmannin, a specific PI 3–kinase inhibitor, failed to block complex formation between Src and GPIb. The Src-SH3 domain as a glutathione S-transferase (GST)–fusion protein coprecipitated the p85 subunit of PI 3–kinase and GPIb. These findings taken together suggest that the p85 subunit of PI 3–kinase mediates GPIb-related activation signals and activates Src independently of the enzymatic activity of PI 3– kinase.