Identification of a novel proline-arginine motif involved in CIN85-dependent clustering of Cbl and down-regulation of epidermal growth factor receptors
Identification of a novel proline-arginine motif involved in CIN85-dependent clustering of Cbl and down-regulation of epidermal growth factor receptors
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DOI：
10.1074/jbc.m304541200
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发表时间：
2003-10-10
影响因子：
4.8
通讯作者：
Dikic, I
中科院分区：
文献类型：
--
作者：
Kowanetz, K;Szymkiewicz, I;Dikic, I
CIN85 是一种多结构域衔接蛋白，参与 Cbl 介导的受体酪氨酸激酶下调。 CIN85 与 Cbl 的结合在生长因子刺激后增加，对于将受体酪氨酸激酶靶向网格蛋白介导的内吞作用至关重要。在此，我们报告了一种新型聚脯氨酸-精氨酸基序 (PXXXPR) 的鉴定，该基序由 CIN85 及其同源物 CMS/CD2AP 的 SH3 结构域特异性识别。该基序对于 CIN85 与 Cbl/Cbl-b、其他 CIN85 SH3 结构域效应子的结合以及介导 SH3-A 结构域和 CIN85 富含脯氨酸区域之间的分子内相互作用是必不可少的。 CIN85 的各个 SH3 结构域以微摩尔亲和力与 Cbl/Cbl-b 的 PXXXPR 肽结合，而两个或三个 SH3 结构域的扩展结构以更高的化学计量和增加的对相同肽的亲和力结合。这使得全尺寸 CIN85 能够同时与多个 Cbl 分子相互作用，促进它们在哺乳动物细胞中聚集。 CIN85 聚集 Cbl 的能力对于配体诱导的 CIN85.Cbl.表皮生长因子受体复合物的稳定以及溶酶体中表皮生长因子受体的降解很重要。因此，CIN85 SH3 结构域与 Cbl 中 PXXXPR 基序的特异性相互作用在受体酪氨酸激酶的下调中发挥多种作用。
CIN85 is a multidomain adaptor protein implicated in Cbl-mediated down-regulation of receptor tyrosine kinases. CIN85 binding to Cbl is increased after growth factor stimulation and is critical for targeting receptor tyrosine kinases to clathrin-mediated endocytosis. Here we report the identification of a novel polyproline-arginine motif (PXXXPR), specifically recognized by the SH3 domains of CIN85 and its homologue CMS/CD2AP. This motif was indispensable for CIN85 binding to Cbl/Cbl-b, to other CIN85 SH3 domains' effectors, and for mediating an intramolecular interaction between the SH3-A domain and the proline-rich region of CIN85. Individual SH3 domains of CIN85 bound to PXXXPR peptides of Cbl/Cbl-b with micromolar affinities, whereas an extended structure of two or three SH3 domains bound with higher stoichiometry and increased affinity to the same peptides. This enabled full size CIN85 to simultaneously interact with multiple Cbl molecules, promoting their clustering in mammalian cells. The ability of CIN85 to cluster Cbl was important for ligand-induced stabilization of CIN85.Cbl.epidermal growth factor receptor complexes, as well as for epidermal growth factor receptor degradation in the lysosome. Thus, specific interactions of CIN85 SH3 domains with the PXXXPR motif in Cbl play multiple roles in down-regulation of receptor tyrosine kinases.