Yang Y, Janich S, Cohn J A, Wilson J M
Department of Internal Medicine, University of Michigan Medical School, Ann Arbor 48109.
Proc Natl Acad Sci U S A. 1993 Oct 15;90(20):9480-4. doi: 10.1073/pnas.90.20.9480.
The most common cause of cystic fibrosis is deletion of Phe-508 (delta F508) from the cystic fibrosis transmembrane conductance regulator (CFTR). Previous studies have suggested that delta F508 CFTR is an unstable protein that retains a pattern of glycosylation specific to the endoplasmic reticulum. This report examines the mechanism responsible for the mislocalization of delta F508 CFTR in a human cystic fibrosis epithelial cell line overexpressing recombinant CFTR by virtue of adenovirus-mediated gene transfer. Immunoelectron microscopy confirmed that wild-type CFTR is delivered to the plasma membrane of these cells and that delta F508 CFTR is retained in the endoplasmic reticulum. Pulse-chase studies showed that newly synthesized CFTR complexes with the chaperone hsp70. The wild-type protein dissociates from hsp70 before its transport to the Golgi, and the protein is subsequently degraded in lysosomes. By contrast, the complex formed between delta F508 CFTR and hsp70 is retained in the endoplasmic reticulum and delta F508 CFTR is rapidly degraded in a pre-Golgi nonlysosomal compartment. Thus, hsp70 discriminates between the normal form of CFTR and the form of the protein that most commonly causes cystic fibrosis (delta F508). These findings clarify the mechanism by which mutation causing delta F508 affects the intracellular trafficking of CFTR and suggest another function for hsp70 in ensuring quality control during the biosynthesis of plasma-membrane proteins.
囊性纤维化最常见的病因是囊性纤维化跨膜传导调节因子(CFTR)中苯丙氨酸508(ΔF508)缺失。以往研究表明,ΔF508 CFTR是一种不稳定蛋白,保留着内质网特有的糖基化模式。本报告借助腺病毒介导的基因转移,研究了在过表达重组CFTR的人囊性纤维化上皮细胞系中,ΔF508 CFTR定位错误的机制。免疫电子显微镜证实,野生型CFTR被转运至这些细胞的质膜,而ΔF508 CFTR则保留在内质网中。脉冲追踪研究表明,新合成的CFTR与伴侣蛋白hsp70形成复合物。野生型蛋白在转运至高尔基体之前与hsp70解离,随后在溶酶体中降解。相比之下,ΔF508 CFTR与hsp70形成的复合物保留在内质网中,ΔF508 CFTR在高尔基体前非溶酶体区室中迅速降解。因此,hsp70能够区分正常形式的CFTR和最常导致囊性纤维化的蛋白形式(ΔF508)。这些发现阐明了导致ΔF508的突变影响CFTR细胞内运输的机制,并提示了hsp70在确保质膜蛋白生物合成过程中的质量控制方面的另一个功能。