TNF receptor-associated factor 4 (TRAF4) is a novel binding partner of glycoprotein Ib and glycoprotein VI in human platelets
TNF receptor-associated factor 4 (TRAF4) is a novel binding partner of glycoprotein Ib and glycoprotein VI in human platelets
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DOI：
10.1111/j.1538-7836.2010.04091.x
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发表时间：
2011-01-01
影响因子：
10.4
通讯作者：
Berndt, M. C.
中科院分区：
文献类型：
--
作者：
Arthur, J. F.;Shen, Y.;Berndt, M. C.
背景资料：活性氧物质的产生是血小板糖蛋白（GP）受体GPIb-IX-V和GPVI的配体接合的结果之一，其结合VWF/胶原蛋白并在动脉剪切下引发血栓形成;然而，将氧化还原途径活化与这些受体的接合偶联的精确分子机制尚不清楚。目的：本研究的目的是鉴定GPIb-IX-V和GPVI的新结合伴侣，其可以提供氧化还原途径和早期血小板信号传导事件之间的潜在联系。方法和结果：使用蛋白质阵列分析和亲和力结合试验，我们证明了孤儿TNF受体相关因子（TRAF）家族成员TRAF 4选择性结合GPIb β和GPVI的胞质序列。TRAF 4、p47 phox [NADPH氧化酶（Nox 2）酶复合物的]和其它氧化还原相关信号蛋白如Hic-5与来自人血小板裂解物的GPIb/GPVI共免疫沉淀，而MBP-TRAF 4或MBP-p47 phox融合蛋白特异性地下拉GPIb/GPVI。GPIb或GPVI选择性激动剂诱导TRAF 4相关蛋白Hic-5和Pyk 2的磷酸化，磷酸化通过Nox 2抑制而减弱。结论：这些结果描述了TRAF 4与受体的第一次直接关联，并确定了GPIb-IX-V和GPVI的新结合伴侣，提供了这些血小板受体和下游TRAF 4/Nox 2依赖性氧化还原途径之间的潜在联系。
Background: Reactive oxygen species generation is one consequence of ligand engagement of platelet glycoprotein (GP) receptors GPIb-IX-V and GPVI, which bind VWF/collagen and initiate thrombosis at arterial shear; however, the precise molecular mechanism coupling redox pathway activation to engagement of these receptors is unknown. Objective: The objective of this study was to identify novel binding partners for GPIb-IX-V and GPVI that could provide a potential link between redox pathways and early platelet signaling events. Methods and Results: Using protein array analysis and affinity-binding assays, we demonstrated that the orphan TNF receptor-associated factor (TRAF) family member, TRAF4, selectively binds cytoplasmic sequences of GPIb beta and GPVI. TRAF4, p47phox [of the NADPH oxidase (Nox2) enzyme complex] and other redox relevant signaling proteins such as Hic-5, co-immunoprecipitate with GPIb/GPVI from human platelet lysates whilst MBP-TRAF4 or MBP-p47phox fusion proteins specifically pull-down GPIb/GPVI. GPIb- or GPVI-selective agonists induce phosphorylation of the TRAF4-associated proteins, Hic-5 and Pyk2, with phosphorylation attenuated by Nox2 inhibition. Conclusion: These results describe the first direct association of TRAF4 with a receptor, and identify a novel binding partner for GPIb-IX-V and GPVI, providing a potential link between these platelet receptors and downstream TRAF4/Nox2-dependent redox pathways.