Syntaxin 16 Binds to Cystic Fibrosis Transmembrane Conductance Regulator and Regulates Its Membrane Trafficking in Epithelial Cells
Syntaxin 16 Binds to Cystic Fibrosis Transmembrane Conductance Regulator and Regulates Its Membrane Trafficking in Epithelial Cells
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DOI：
10.1074/jbc.m110.162438
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发表时间：
2010-11-12
影响因子：
4.8
通讯作者：
Lee, Min Goo
中科院分区：
文献类型：
--
作者：
Gee, Heon Yung;Tang, Bor Luen;Lee, Min Goo
囊性纤维化跨膜电导调节器（CFTR）是调节上皮通透性的上皮离子转运蛋白复杂网络中的关键膜蛋白。突触融合蛋白是细胞内运输和分泌蛋白膜靶向的主要决定因素之一。在本研究中，我们证明了 CFTR 和突触蛋白 16 (STX16) 之间的生化和功能关联，后者介导早期/晚期内体和跨高尔基体网络内的囊泡运输。大鼠结肠和 T84 人结肠上皮细胞的免疫沉淀实验表明 STX16 与 CFTR 相关。使用域特异性下拉分析的进一步分析表明，STX16 的螺旋结构域直接与 CFTR 的 N 末端区域相互作用。大鼠结肠和 T84 细胞的免疫染色显示 CFTR 和 STX16 在上皮细胞的顶端和顶端下区域高度共定位。有趣的是，通过敲低 T84 细胞中 STX16 的表达，CFTR 相关的氯电流减少。表面生物素化和回收测定表明 CFTR 氯电流的减少是由于质膜上 CFTR 表达的减少所致。这些结果表明STX16介导CFTR的再循环并构成肠上皮细胞中CFTR运输机制的重要组成部分。
The cystic fibrosis transmembrane conductance regulator (CFTR) is a key membrane protein in the complex network of epithelial ion transporters regulating epithelial permeability. Syntaxins are one of the major determinants in the intracellular trafficking and membrane targeting of secretory proteins. In the present study we demonstrate the biochemical and functional association between CFTR and syntaxin 16 (STX16) that mediates vesicle transport within the early/late endosomes and trans-Golgi network. Immunoprecipitation experiments in rat colon and T84 human colonic epithelial cells indicate that STX16 associates with CFTR. Further analyses using the domain-specific pulldown assay reveal that the helix domain of STX16 directly interacts with the N-terminal region of CFTR. Immunostainings in rat colon and T84 cells show that CFTR and STX16 highly co-localize at the apical and subapical regions of epithelial cells. Interestingly, CFTR-associated chloride current was reduced by the knockdown of STX16 expression in T84 cells. Surface biotinylation and recycling assays indicate that the reduction in CFTR chloride current is due to decreased CFTR expression on the plasma membrane. These results suggest that STX16 mediates recycling of CFTR and constitutes an important component of CFTR trafficking machinery in intestinal epithelial cells.