The UIM domain of Hrs couples receptor sorting to vesicle formation
The UIM domain of Hrs couples receptor sorting to vesicle formation
复制标题
DOI：
10.1242/jcs.00723
复制
发表时间：
2003-10-15
影响因子：
4
通讯作者：
Clague, MJ
中科院分区：
文献类型：
--
作者：
Urbé, S;Sachse, M;Clague, MJ
肝细胞生长因子调节的酪氨酸激酶底物(HRS)最初被认为是激活的酪氨酸激酶受体的底物，它是内体分类的双层网状蛋白涂层的主要成分。我们已经分析了HRs对表皮生长因子(EGF)刺激的反应，并表明进化保守的酪氨酸Y329和Y334提供了主要的磷酸化位点。HRS被认为是通过与其泛素相互作用基序(UIM)结构域的直接相互作用，将泛素化的受体集中在被笼蛋白覆盖的区域。我们发现，相同的UIM结构域是EGF刺激的HRs酪氨酸磷酸化所必需的。野生型HRs的过表达或EGF依赖的磷酸化缺陷的双突变体Y329/334F都通过抑制内部囊泡的形成而显著抑制EGF受体(EGFR)的降解，从而阻止EGFR整合到多囊泡体的腔小泡中。相反，HRS-UIM结构域的突变或缺失强烈地抑制了这一效应。此外，在内膜上也观察到UIM缺失和点突变，这表明在将受体复合体掺入腔小泡之前，未能从内体膜解离。我们的数据表明，作为腔小泡形成的前奏，HRs的UIM结构域在主动保留内体限制膜上的EGFR方面发挥了作用。
Hepatocyte growth factor regulated tyrosine kinase substrate (Hrs), a main component of the 'bilayered' clathrin coat on sorting endosomes, was originally identified as a substrate of activated tyrosine kinase receptors. We have analysed Hrs phosphorylation in response to epidermal growth factor (EGF) stimulation and show that the evolutionary conserved tyrosines Y329 and Y334 provide the principal phosphorylation sites. Hrs is proposed to concentrate ubiquitinated receptors within clathrin-coated regions via direct interaction with its UIM (ubiquitin interaction motif) domain. We show that the same UIM domain is necessary for EGF-stimulated tyrosine phosphorylation of Hrs. Over-expression of wildtype Hrs or a double mutant, Y329/334F, defective in EGF-dependent phosphorylation, both substantially retard EGF receptor (EGFR) degradation by inhibiting internal vesicle formation and thereby preventing EGFR incorporation into lumenal vesicles of the multivesicular bodies. In contrast, mutation or deletion of the Hrs-UIM domain strongly suppresses this effect. In addition the UIM-deletion and point mutants are also observed on internal membranes, indicating a failure to dissociate from the endosomal membrane prior to incorporation of the receptor complex into lumenal vesicles. Our data suggest a role for the UIM-domain of Hrs in actively retaining EGFR at the limiting membrane of endosomes as a prelude to lumenal vesicle formation.