Figure 6.

BAG-2 inhibits the chaperone-assisted degradation of CFTR and is essential for CFTR maturation. (A) HEK293 cells were transiently transfected with CFTR-, BAG-2-, and CHIP-encoding plasmids in the indicated combinations. Amounts of pcDNA3.1-bag-2 were 1- and 2-fold of the amount of pcDNA3.1-chip when indicated. The ER-localized, core-glycosylated B-form (B) and the completely glycosylated, plasma membrane-localized C-form of CFTR (C) were detected by immunoblotting. Likewise, BAG-2 and CHIP were detected with specific antibodies. Hsc70 and Hsp70 served as loading control. Each lane represents 60 μg of cellular extract. CFTR levels were quantified from four independent experiments and mean values are presented as fold change compared with the control reaction, which was set to 1. (B) HEK293 cells were transiently transfected with CFTR-, BAG-2 shRNA-, and BAG-2-expressing plasmids as indicated. Cell extracts, 60 μg, were separated by SDS-PAGE. Protein levels were analyzed by immunoblotting using specific antibodies. Hsc70/Hsp70 served as loading control. Samples that did not express BAG-2 shRNA received the pSUPER plasmid without an insert.