Molecular cloning and transmembrane structure of hCLCA2 from human lung, trachea, and mammary gland

The CLCA family of Ca2+-activated Cl- channels has recently been discovered , with an increasing number of closely related members isolated from differ ent species. Here we report the cloning of the second human homolog, hCLCA2 , from a human lung cDNA library. Northern blot and RT-PCR analyses reveale d additional expression in trachea and mammary gland. A primary translation product of 120 kDa was cleaved into two cell surface-associated glycoprote ins of 86 and 34 kDa in transfected HEK-293 cells. hCLCA2 is the first CLCA homolog for which the transmembrane structure has been systematically stud ied. Glycosylation site scanning and protease protection assays revealed fi ve transmembrane domains with a large, cysteine-rich, amino-terminal extrac ellular domain. Whole cell patch-clamp recordings of hCLCA2-transfected HEK -293 cells detected a slightly outwardly rectifying anion conductance that was increased in the presence of the Ca2+ ionophore ionomycin and inhibited by DIDS, dithiothreitol, niflumic acid, and tamoxifen. Expression in human trachea and lung suggests that hCLCA2 may play a role in the complex patho genesis of cystic fibrosis.