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Regulatory mechanisms of CIC chloride channels by protein-protein interaction

蛋白质-蛋白质相互作用对 CIC 氯离子通道的调节机制

基本信息

批准号：
13670035
负责人：
FURUKAWA Tetsushi
金额：
$ 2.24万
依托单位：
Akita University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13670035/
关键词：
chloride channel protein-protein interaction yeast two-hybrid system cell cycle cyclin-dependent kinase ubiquitin CFTR PDZ domain 蛋白相互作用ユビキチン化

项目摘要

(1) The C-terminus of ClC-2 channel is directly phosphorylated by M phase-specific cyclin-dependent kinase, p34^<cdc2>/cyclin B, and de-phosphorylated by protein phosphatase 1, for which protein-protein interaction plays a crucial role. ClC-2 channel expressed in Xenopus oocytes is inhibited by phosphorylation and activated by dephosphorylation.(2) Western blot analysis and immunocytochemistry revealed that ClC-2 channel protein is expressed in dividing cells of the M phase of cell cycle, and promptly disappears after cell division. The PEST sequence is a signature of short-lived proteins via ubiquitination, and multiple PEST sequences are present in the C-terminus of ClC-2. Phosphorylation of ^<632>Ser of ClC-2 by p34^<cdc2>/cyclin B triggers ClC-2 channel ubiquitination, which underlines M phase-specific ubiquitination and degradation of ClC-2 channel protein.(3) ClC-3B is a chloride channel expressed predominantly in epithelial cells and is localized in its microvilli. ClC-3B interacts With EBP50, an epithelia-specific PDZ-containing protein, and thereby interacts with cystic fibrosis transmembrane conductance regulator (CFTR), a product of cystic fibrosis gene. ClC-3B chloride channel is activated by protein kinase A in the presence of CFTR, and is important in ionic transport across the epithelia.

(1)ClC-2通道的C-末端被M期特异性细胞周期蛋白依赖性激酶p34^ /cyclin B直接磷酸化<cdc2>，并被蛋白磷酸酶1去磷酸化，蛋白-蛋白相互作用在其中起关键作用。在非洲爪蟾卵母细胞中表达的ClC-2通道被磷酸化抑制，被去磷酸化激活。(2)Western blot分析和免疫细胞化学显示，ClC-2通道蛋白在细胞周期M期的分裂细胞中表达，并在细胞分裂后迅速消失。PEST序列是通过泛素化的短寿命蛋白质的特征，并且多个PEST序列存在于ClC-2的C-末端。p34 <632>^ /cyclin B对ClC-2的^ Ser<cdc2>的磷酸化触发ClC-2通道的泛素化，这强调了ClC-2通道蛋白的M期特异性泛素化和降解。(3)ClC-3B是主要在上皮细胞中表达的氯离子通道，并且定位于其微绒毛中。ClC-3B与EBP 50（一种上皮特异性含PDZ的蛋白）相互作用，从而与囊性纤维化跨膜传导调节因子（CFTR）（囊性纤维化基因的产物）相互作用。ClC-3B氯离子通道在CFTR存在下被蛋白激酶A激活，并且在跨上皮的离子转运中是重要的。