The ubiquitin-protein ligase Nedd4-2 differentially interacts with and regulates members of the Tweety family of chloride ion channels
The ubiquitin-protein ligase Nedd4-2 differentially interacts with and regulates members of the Tweety family of chloride ion channels
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DOI：
10.1074/jbc.m803361200
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发表时间：
2008-08-29
影响因子：
4.8
通讯作者：
Hooper, John D.
中科院分区：
文献类型：
--
作者：
He, Yaowu;Hryciw, Deanne H.;Hooper, John D.
Twety蛋白包括一个氯离子通道家族，有三个成员在人类(TTYH1-3)以及苍蝇和小鼠物种中发现的同源物。在人类中，TTYH2表达增加与癌症进展有关，而Fly Twety与发育过程有关。在结构上，Twety蛋白由5个膜扫描结构域和N-糖链修饰组成，这些结构域对转运到质膜具有重要作用，这些蛋白的氨基末端位于细胞外，而羧基末端位于细胞质内。除了N-糖基化外，由Hect类型E3泛素连接酶Nedd4-2介导的泛素化也是一种翻译后修饰，对调节膜蛋白起重要作用。在本研究中，我们对每个TTYH1-3与Nedd4-2相互作用以及被该连接酶泛素化和调节的能力进行了全面的分析。我们的数据表明，Nedd4-2与两个家族成员TTYH2和TTYH3结合，TTYH2和TTYH3含有Hect型E3泛素连接酶的共识PY((L/P)PXY)结合部位，但不与TTYH1结合，TTYH1没有这个基序。一致地，Nedd4-2泛素化TTYH2和TTYH3。重要的是，我们已经证明了内源TTYH2和Nedd4-2是结合伙伴，并证明了TTYH2 PY基序在这些相互作用中是必不可少的。我们还表明，Nedd4-2介导的TTYH2泛素化是该蛋白细胞表面和细胞总水平的关键调节因子。这些数据表明，Nedd4-2与TTYH1-3有不同的相互作用和调节作用，这将对理解Twety蛋白在生理和疾病中的调控机制具有重要意义。
The Tweety proteins comprise a family of chloride ion channels with three members identified in humans (TTYH1-3) and orthologues in fly and murine species. In humans, increased TTYH2 expression is associated with cancer progression, whereas fly Tweety is associated with developmental processes. Structurally, Tweety proteins are characterized by five membranespanning domains and N-glycan modifications important for trafficking to the plasma membrane, where these proteins are oriented with the amino terminus located extracellularly and the carboxyl terminus cytoplasmically. In addition to N-glycosylation, ubiquitination mediated by the HECT type E3 ubiquitin ligase Nedd4-2 is a post-translation modification important in regulating membrane proteins. In the present study, we performed a comprehensive analysis of the ability of each of TTYH1-3 to interact with Nedd4-2 and to be ubiquitinated and regulated by this ligase. Our data indicate that Nedd4-2 binds to two family members, TTYH2 and TTYH3, which contain consensus PY ((L/P)PXY) binding sites for HECT type E3 ubiquitin ligases, but not to TTYH1, which lacks this motif. Consistently, Nedd4-2 ubiquitinates both TTYH2 and TTYH3. Importantly, we have shown that endogenous TTYH2 and Nedd4-2 are binding partners and demonstrated that the TTYH2PY motif is essential for these interactions. We have also shown that Nedd4-2-mediated ubiquitination of TTYH2 is a critical regulator of cell surface and total cellular levels of this protein. These data, indicating that Nedd4-2 differentially interacts with and regulates TTYH1-3, will be important for understanding mechanisms controlling Tweety proteins in physiology and disease.