The ubiquitous ClC-2 Cl⁻channel is thought to contribute to epithelial Cl⁻secretion, but the distribution of the ClC-2 protein in human epithelia has not been investigated. We have studied the distribution of ClC-2 in adult human and rat intestine and airways by immunoblotting and confocal microscopy. In the rat, ClC-2 was present in the lateral membranes of villus enterocytes and was predominant at the basolateral membranes of luminal colon enterocytes. The expression pattern of ClC-2 in the human intestine differed significantly, because ClC-2 was mainly detected in a supranuclear compartment of colon cells. We found significant expression of ClC-2 at the apex of ciliated cells in both rat and human airways. These results show that the distribution of ClC-2 in airways is consistent with participation of ClC-2 channels in Cl⁻ secretion and indicate that extrapolation of results from studies of ClC-2 function in rat intestine to human intestine is not straightforward.

CFTR (cystic fibrosis transmembrane conductance regulator) is an epithelial Cl− channel inhibited with high affinity and selectivity by the thiazolidinone compound CFTRinh-172. In the present study, we provide evidence that CFTRinh-172 acts directly on the CFTR. We introduced mutations in amino acid residues of the sixth transmembrane helix of the CFTR protein, a domain that has an important role in the formation of the channel pore. Basic and hydrophilic amino acids at positions 334–352 were replaced with alanine residues and the sensitivity to CFTRinh-172 was assessed using functional assays. We found that an arginine-to-alanine change at position 347 reduced the inhibitory potency of CFTRinh-172 by 20–30-fold. Mutagenesis of Arg347 to other amino acids also decreased the inhibitory potency, with aspartate producing near total loss of CFTRinh-172 activity. The results of the present study provide evidence that CFTRinh-172 interacts directly with CFTR, and that Arg347 is important for the interaction.