泛素E3连接酶Nedd4L介导Pendrin降解的作用机制研究  

Nedd4L-mediated Ubiquitination of a Congenital Deafness-related Gene Pendrin Induces Pendrin Degradation

作　　者：方桂村 张碧君[1] 赵彦艳[1] FANG Guicun;ZHANG Bijun;ZHAO Yanyan(Department of Clinical Genetics,Shengjing Hospital,China Medical University)

机构地区：[1]中国医科大学,沈阳110122

出　　处：《中华耳科学杂志》2020年第3期570-575,共6页Chinese Journal of Otology

基　　金：国家重点研发计划(2016YFC1000702);国家自然科学基金面上项目(No.31571198)。

摘　　要：目的探讨泛素E3连接酶Nedd4L在Pendrin泛素化修饰中的作用及对Pendrin表达的影响。方法培养293T细胞,Co-IP实验验证Nedd4L与Pendrin存在体内结合;构建Nedd4L真核细胞表达载体;转染293T细胞后Western Blot检测Pendrin的表达量以及泛素化程度。结果Co-IP实验证实Nedd4L与Pendrin在293T细胞中存在体内结合;真核表达载体构建,能够表达完整Nedd4L蛋白;转染72h后,Pendrin在293T细胞中的表达量降低,泛素化修饰增加,在细胞膜表面表达降低。结论Nedd4L能够结合Pendrin并通过促进其泛素化修饰以及降低细胞膜表面表达量。其机制可能是通过促进Pendrin泛素化修饰诱导Pendrin内化并进入泛素蛋白酶体途径降解。Objective To investigate the role of ubiquitin E3 ligase Nedd4L in the ubiquitination modification of Pendrin.Methods 293T cells were cultured,and Nedd4L binding with Pendrin was verified by Co-IP assay.A Nedd4L eukaryotic expression vector was constructed.Pendrin expression and ubiquitination were detected by Western Blot after 293T cells were transfected.Results Co-IP assay confirmed that Nedd4L and Pendrin bound in 293T cells in vitro.The eukaryotic expression vector was successfully constructed to express the complete Nedd4L protein.After 72 hours of transfection,Pendrin expression in 293T cells decreased,ubiquitination modification increased,and expression on cell membrane surface decreased.Conclusion Nedd4L can bind Pendrin and induce its ubiquitination by promoting its ubiquitination modification,as well as decreasing its expression on cell membrane surface.The mechanism to induce Pendrin internalization and degradation may be through promoting Pendrin ubiquitination modification.

关 键 词：泛素化 Nedd4L 先天耳聋 PDS 

分 类 号：R764[医药卫生—耳鼻咽喉科]