Characterization of ubiquilin 1, an mTOR-interacting protein
Characterization of ubiquilin 1, an mTOR-interacting protein
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DOI：
10.1016/s0167-4889(01)00164-1
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发表时间：
2002-01-30
影响因子：
5.1
通讯作者：
Avruch, J
中科院分区：
文献类型：
--
作者：
Wu, SL;Mikhailov, A;Avruch, J
已知 mTOR 蛋白激酶通过调节翻译、转录、膜运输和蛋白质降解来控制细胞周期进程和细胞生长。 mTOR 的已知相互作用并不能解释该蛋白质的多种功能。使用 mTOR (1-670) 的非催化片段作为酵母双杂交筛选相互作用蛋白的诱饵，鉴定出泛素 1 (NM013438)。泛素 1 是功能未知的系统发育保守基因家族的成员，其特征在于 N 端泛素样 (Ubq) 结构域、C 端泛素相关 (Uba) 结构域和包含大量 NPXphi 基序（X，任意；phi 疏水性氨基酸）的中心区域。 GST-ubiquilin 1 与哺乳动物细胞中的 FLAG-mTOR（残基 1-670）特异性结合； mTOR 的残基 570-670 和泛素 1 的残基 226-323 是这种相互作用所必需的。 mTOR 和泛素免疫反应性均以细小斑点的形式遍布细胞质；未观察到与细胞骨架元件、早期内体或蛋白酶体的显着共定位。通过细胞分级分离评估，mTOR 主要与低密度膜相关，同时还有 10% 的泛素 1。泛素 1 是一种雷帕霉素不敏感的磷蛋白。在雷帕霉素存在或不存在的情况下，泛素 1 的过表达不会改变共转染 mTOR 的激酶活性或 mTOR 靶标 p70 S6 激酶的磷酸化。我们的数据表明我们已经鉴定出一种新的 mTOR 相互作用因子，即泛素 1。这种可能是基于膜的相互作用的生物学意义需要进一步研究。 (C) 2002 Elsevier Science B.V. 保留所有权利。
The mTOR protein kinase is known to control cell cycle progression and cell growth through regulation of translation, transcription, membrane traffic and protein degradation. Known interactions of mTOR do not account for the multiple functions of this protein. Using a non-catalytic segment of mTOR (1-670) as bait in a yeast two-hybrid screen for interacting proteins, ubiquilin 1 (NM013438) was identified. Ubiquilin 1 is a member of a phylogenetically conserved gene family of unknown function, characterized by an N-terminal ubiquitin-like (Ubq) domain, a C-terminal ubiquitin associated (Uba) domain and a central region containing numerous NPXphi motifs (X, any; phi hydrophobic amino acid). GST-ubiquilin 1 binds specifically to FLAG-mTOR (residues 1-670) in mammalian cells; residues 570-670 of mTOR and 226-323 of ubiquilin 1 are required for this interaction. Both mTOR and ubiquilin immunoreactivity appear as fine speckles throughout the cytoplasm; significant colocalization with cytoskeletal elements, early endosomes or proteasomes is not observed. As assessed by cell fractionation, mTOR is predominantly associated with low density membranes, along with 10% of ubiquilin 1. Ubiquilin 1 is a rapamycin-insensitive phosphoprotein. Overexpression of ubiquilin 1 does not alter the kinase activity of cotransfected mTOR or the phosphorylation of the mTOR target, p70 S6 kinase, in the presence or absence of rapamycin. Our data suggest that we have identified a novel mTOR interactor, ubiquilin 1. The biological significance of this, presumably membrane based, interaction, requires further study. (C) 2002 Elsevier Science B.V. All rights reserved.