| Replacement or circumvention of mutant forms of the cystic fibrosis transmembrane conductance regulator (CFTR) with wild-type CFTR is one of the goals of CF Research. Such approaches may be complicated further by possible CFTR oligomerization or multimerization. Hints from the CF literature suggest a mild CF disease-like pheno-type in CF heterozygous carriers that is most likely attributed to the DeltaF508-CFTR mutation. As such, we coexpressed wild-type CFTR with increasing amounts of DeltaF508-CFTR in IB3-1 CF epithelial cells and HEK-293 human embryonic kidney cells null for CFTR protein. Increasing amounts of DeltaF508-CFTR inhibited wild-type CFTR processing and cyclic AMP-activated Cl- channel function in IB3-1 CF cells but not in the HEK293T cells, which suggests background specificity. In addition, CALU-3 non-CF epithelial cells, which endogenously express wild-type CFTR, stably expressing DeltaF508-CFTR were used to perform short-circuit current (ISC) recordings via Ussing chamber and in parallel protein biochemistry. These transfectants showed a reduction in cAMP stimulated short-circuit current and wild-type CFTR processing. An ultimate test of this dominant negative-like effect of DeltaF508-CFTR on wild-type CFTR was the parallel study of CFTR function and processing in two CF mouse models, the DeltaF508-CFTR mouse and CFTR lung and airway knock-out mouse. Three genotypes (wild-type, heterozygous, and homozygous) were assessed in each model. WT/DeltaF508 heterozygotes had an intermediate phenotype with regard to CFTR agonist responses in in vivo nasal potential difference (NPD) recordings and in vitro short-circuit current recordings on mouse tracheal epithelial cells monolayer isolated from each mouse genotype In contrast, +/- heterozygotes had no difference in their responses versus +/+ wild-type mice. Taken together, these data suggest that DeltaF508-CFTR inhibits the processing and function of wild-type CFTR. |
