Sodium 4-phenylbutyrate downregulates Hsc70:: implications for intracellular trafficking of ΔF508-CFTR
Sodium 4-phenylbutyrate downregulates Hsc70:: implications for intracellular trafficking of ΔF508-CFTR
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DOI：
10.1152/ajpcell.2000.278.2.c259
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发表时间：
2000-02-01
影响因子：
5.5
通讯作者：
Zeitlin, PL
中科院分区：
文献类型：
--
作者：
Rubenstein, RC;Zeitlin, PL
囊性纤维化跨膜传导调节因子(CFTR)最常见的突变是Delta F508，它是一种运输突变，与分子伴侣有很长时间的联系，并至少部分地被泛素-蛋白酶体系统降解。4-苯丁酸钠(4PBA)在体外和体内均能改善Delta F508-CFTR在囊性纤维化上皮细胞中的转运和功能。为了进一步了解4PBA的作用机制，我们验证了4PBA通过减少囊性纤维化上皮细胞Hsc70的表达来调节Delta F508-CFTR泛素化和降解的靶向的假设。经0.05-5 mM 4PBA处理2天的IB3-1细胞(DELTA F508/W1282X)，Hsc70蛋白免疫反应性和mRNA水平呈剂量依赖性降低。用Hsc70特异性抗血清进行免疫沉淀表明，Hsc70和CFTR在对照条件下是结合的，而4PBA处理减少了这些复合体。4PBA处理不影响Hsp40、Hdj2、Hsp70、Hsp90和calnexin的表达水平。这些数据表明，4PBA可能通过允许更大比例的突变CFTR逃脱与Hsc70的关联，从而改善Delta F508-CFTR的运输。
The most common mutation of the cystic fibrosis transmembrane conductance regulator (CFTR), Delta F508, is a trafficking mutant that has prolonged associations with molecular chaperones and is rapidly degraded, at least in part by the ubiquitin-proteasome system. Sodium 4-phenylbutyrate (4PBA) improves Delta F508-CFTR trafficking and function in vitro in cystic fibrosis epithelial cells and in vivo. To further understand the mechanism of action of 4PBA, we tested the hypothesis that 4PBA modulates the targeting of Delta F508-CFTR for ubiquitination and degradation by reducing the expression of Hsc70 in cystic fibrosis epithelial cells. IB3-1 cells (genotype Delta F508/W1282X) that were treated with 0.05-5 mM 4PBA for 2 days in culture demonstrated a dose-dependent reduction in Hsc70 protein immunoreactivity and mRNA levels. Immunoprecipitation with Hsc70-specific antiserum demonstrated that Hsc70 and CFTR associated under control conditions and that treatment with 4PBA reduced these complexes. Levels of immunoreactive Hsp40, Hdj2, Hsp70, Hsp90, and calnexin were unaffected by 4PBA treatment. These data suggest that 4PBA may improve Delta F508-CFTR trafficking hy allowing a greater proportion of mutant CFTR to escape association with Hsc70.