Mutations in CFTR associated with mild-disease-form CI^- channels with altered pore properties

Abstract

THE cystic fibrosis transmembrane conductance regulator (CFTR) is a phosphorylation-regulated Cl^- channel located in the apical membrane of epithelia^1–10. Although cystic fibrosis (CF) is caused by mutations in a single gene encoding CFTR^11,12, the disease has a variable clinical phenotype^13,14. The most common mutation associated with cystic fibrosis, deletion of a phenylalanine at position 508 (frequency, 67%), is associated with severe disease^15–17. But some missense mutations, for example ones in which arginine is replaced by histidine at residue at 117 (R117H; 0.8%), tryptophan at 334 (0.4%), or proline at 347 (0.5%), are associated with milder disease^15,17,18. These missense mutations affect basic residues located at the external end of the second (M2) and in the sixth (M6) putative membrane-spanning sequences. Here we report that, when expressed in heterologous epithelial cells, all three mutants were correctly processed and generated cyclic AMP-regulated apical Cl^- currents. Although the macroscopic current properties were normal, the amount of current was reduced. Patch-clamp analysis revealed that all three mutants had reduced single-channel conductances. In addition, R117H showed altered sensitivity to external pH and had altered single-channel kinetics. These results explain the quantitative decrease in macroscopic Cl^- current, and suggest that R117, R334 and R347 contribute to the pore of the CFTR Cl^- channel. Our results also suggest why R117H, R334W and R347P produce less severe clinical disease and have implications for our understanding of cystic fibrosis.