Figure 4

p.Ile1234_Arg1239del-CFTR is misprocessed, retained in the endoplasmic reticulum, and partially rescued by Lumacaftor (currently in clinical trials for p.Phe508del patients). (a) Immunoblots show steady-state levels of wild-type (WT)-CFTR, p.Phe508del-CFTR, and p.Ile1234_Arg1239del-CFTR in baby hamster kidney (BHK) cells after 24 h at 37 °C. WT-CFTR was expressed as both the Golgi-modified, complex glycosylated (mature) band C form (broad 170-kDa band) and the endoplasmic reticulum–modified, core glycosylated (immature), band B form (sharp 150-kDa band). p.Phe508del-CFTR and p.Ile1234_Arg1239del-CFTR were expressed primarily as the immature, band B form. Maturation (expressed as percentage of band C/(band B + band C)) of WT-CFTR, p.Phe508del-CFTR, and p.Ile1234_Arg1239del-CFTR was quantified for three independent trials, and a significant difference was found between the mutant proteins (P < 0.01). (b) To evaluate glycosylation status, immunoblots show the sensitivity of WT-CFTR, p.Phe508del-CFTR, and p.Ile1234_Arg1239del-CFTR to endoglycosidase H and peptide-N-glycosidase F. White arrowhead, complex glycosylated; black arrowhead, core glycosylated; gray arrowhead, deglycosylated. (c) Immunoblots show the processing of WT-CFTR, p.Phe508del-CFTR, and p.Ile1234_Arg1239del-CFTR in BHK cells in the absence (dimethyl sulfoxide) and presence of 3 µmol/l Lumacaftor (or VX-809) after 24 h at 37 °C. Equivalent sample loading was confirmed by immunoblots of calnexin protein expression. Maturation (expressed as percentage of band C/(band B + band C)) of p.Phe508del-CFTR and p.Ile1234_Arg1239del-CFTR was quantified for three independent trials, and there was a significant difference between untreated (dimethyl sulfoxide) and Lumacaftor (VX-809)-treated cultures, as indicated by asterisks (**P < 0.01). CFTR, cystic fibrosis transmembrane conductance regulator.