The most common cystic fibrosis transmembrane conductance regulator (CFTR) mutant in cystic fibrosis patients, ⌬F508 CFTR, is retained in the endoplasmic reticulum (ER) and is consequently degraded by the ubiquitin-proteasome pathway known as ER-associated degradation (ERAD). Because the prolonged interaction of ⌬F508 CFTR with calnexin, an ER chaperone, results in the ERAD of ⌬F508 CFTR, calnexin seems to lead it to the ERAD pathway. However, the role of calnexin in the ERAD is controversial. In this study, we found that calnexin overexpression partially attenuated the ERAD of ⌬F508 CFTR. We observed the formation of concentric membranous bodies in the ER upon calnexin overexpression and that the ⌬F508 CFTR but not the wild-type CFTR was retained in the concentric membranous bodies. Furthermore, we observed that calnexin overexpression moderately inhibited the formation of aggresomes accumulating the ubiquitinated ⌬F508 CFTR. These findings suggest that the overexpression of calnexin may be able to create a pool of ⌬F508 CFTR in the ER. INTRODUCTIONCystic fibrosis transmembrane conductance regulator (CFTR) is a polytopic integral membrane protein that functions as a cAMP-dependent Cl Ϫ channel in the apical membrane of epithelial cells (Anderson et al., 1991;Drumm et al., 1991;Tabcharani et al., 1991;Bear et al., 1992). Mutations in the CFTR gene lead to the absence or malfunction of a regulated Cl Ϫ channel in the apical membrane of secretory epithelia, resulting in the clinical symptoms of cystic fibrosis (CF) (Kerem et al., 1989;Riordan et al., 1989;Rommens et al., 1989;Cheng et al., 1990). Therefore, potential CF therapies are aimed at overcoming the functional impairment of various mutant CFTRs, particularly ⌬F508 CFTR, in which a phenylalanine at position 508 is deleted from the first nucleotide-binding fold. This mutation is found in ϳ70% of CF chromosomes and results in a severe form of the disease; Ͼ90% of CF patients have at least one ⌬F508 allele (Tsui, 1992;Sferra and Collins, 1993). Although ⌬F508 CFTR is functionally competent as a cAMP-dependent Cl Ϫ channel in model situations where it reaches the plasma membrane (Drumm et al., 1991;Denning et al., 1992;Li et al., 1993), in mammalian cells ⌬F508 CFTR fails to reach the plasma membrane.CFTR biogenesis is inefficient. During insertion into the ER membrane, CFTR interacts with the cytosolic chaperones Hsc70/Hdj-2 and Hsp90, as well as the ER chaperone calnexin (CNX) (Yang et al., 1993;Pind et al., 1994;Loo et al., 1998;Meacham et al., 1999). After ATP-dependent conformational maturation, assisted by cytosolic and ER chaperones, the chaperones release CFTR, and only ϳ30% of the immature wild-type (wt) CFTR transits to the late secretory pathway, ultimately reaching the plasma membrane (Lukacs et al., 1994). However, most of the immature wt CFTR (ϳ70%), and nearly all of immature ⌬F508 CFTR, fail to mature and do not transit to the late secretory pathway (Lukacs et al., 1994). They are trapped in the ER as core-glycosylated products (ϳ140 kDa) and ar...

No abstract