CrkL is an adapter for Wiskott-Aldrich syndrome protein and Syk
CrkL is an adapter for Wiskott-Aldrich syndrome protein and Syk
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DOI：
10.1182/blood.v97.9.2633
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发表时间：
2001-05-01
期刊：
影响因子：
20.3
通讯作者：
Ikeda, H
中科院分区：
文献类型：
--
作者：
Oda, A;Ochs, HD;Ikeda, H
Wiskott-Aldrich 综合征 (WAS) 和 X 连锁血小板减少症是由 WAS 蛋白 (WASP) 基因突变引起的。 WASP 可能参与足体的调节，足体是一种由各种类型的细胞形成的富含肌动蛋白的动态细胞粘附结构。 WASP 与足体或其他细胞粘附结构之间的分子联系尚不清楚。血小板表达 SH2-SH3 衔接分子 CrkL，它可以直接与位于足体中的桩蛋白结合。因此，测试了 CrkL 与 WASP 结合的假设。使用抗 CrkL 和 GST 融合蛋白的共沉淀实验结果表明，CrkL 通过其 SH3 结构域与 WASP 结合，并且该结合不受 WASP 酪氨酸磷酸化的影响。 PSTPIP1 的 GST 融合 SH3 结构域的体外结合也不受 WASP 酪氨酸磷酸化的影响，表明 SH3 结构域与 WASP 的结合不受 WASP 酪氨酸磷酸化的抑制。抗 CrkL 还可共沉淀一种 72 kd 的蛋白质，该蛋白质被鉴定为 syk 酪氨酸激酶，对于胶原蛋白诱导的血小板激活至关重要。体外激酶测定证明，CrkL 免疫沉淀物含有激酶活性 syk。使用 GST 融合 CrkL 蛋白的共沉淀实验表明，CrkL 的 SH2 和 SH3 结构域均参与 CrkL 与 syk、WASP、CrkL、syk 和血小板聚集后掺入血小板细胞骨架的桩蛋白样 Hic-5 的结合。因此，CrkL 是 WASP 和 syk 的新型分子接头，并可能通过与细胞骨架蛋白（如 Hic-5）结合将这些分子转移到细胞骨架。 (Blood, 2001; 97:2633-2639) (C) 2001 年，美国血液学会。
Wiskott-Aldrich syndrome (WAS) and X-linked thrombocytopenia are caused by mutations of the WAS protein (WASP) gene. WASP may be involved in the regulation of podosome, an actin-rich dynamic cell adhesion structure formed by various types of cells. The molecular links between WASP and podosomes or other cell adhesion structures are unknown. Platelets express an SH2-SH3 adapter molecule, CrkL, that can directly associate with paxillin, which is localized in podosomes, The hypothesis that CrkL binds to WASP was, therefore, tested. Results from coprecipitation experiments using anti-CrkL and GST-fusion proteins suggest that CrkL binds to WASP through its SH3 domain and that the binding was not affected by WASP tyrosine phosphorylation. The binding of GST-fusion SH3 domain of PSTPIP1 in vitro was also not affected by WASP tyrosine phosphorylation, suggesting that the binding of the SH3 domains to WASP is not inhibited by tyrosine phosphorylation of WASP. Anti-CrkL also coprecipitates a 72-kd protein, which was identified as syk tyrosine kinase, critical for collagen induced-platelet activation. CrkL immunoprecipitates contain kinase-active syk, as evidenced by an in vitro kinase assay. Coprecipitation experiments using GST-fusion CrkL proteins suggest that both SH2 and SH3 domains of CrkL are involved in the binding of CrkL to syk, WASP, CrkL, syk, and paxillin-like Hic-5 incorporated to platelet cytoskeleton after platelet aggregation. Thus, CrkL is a novel molecular adapter for WASP and syk and may potentially transfer these molecules to the cytoskeleton through association with cytoskeletal proteins such as Hic-5. (Blood, 2001; 97:2633-2639) (C) 2001 by The American Society of Hematology.