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

The CLCA family ofCa²⁺-activatedCl channels has recentlybeen discovered, with an increasing number of closely related members isolated from different species. Here we report the cloning of thesecond human homolog, hCLCA2, from a human lung cDNA library. Northernblot and RT-PCR analyses revealed additional expression in trachea andmammary gland. A primary translation product of 120 kDa was cleavedinto two cell surface-associated glycoproteins of 86 and 34 kDa intransfected HEK-293 cells. hCLCA2 is the first CLCA homolog for whichthe transmembrane structure has been systematically studied.Glycosylation site scanning and protease protection assays revealedfive transmembrane domains with a large, cysteine-rich, amino-terminalextracellular domain. Whole cell patch-clamp recordings ofhCLCA2-transfected HEK-293 cells detected a slightly outwardly rectifying anion conductance that was increased in the presence of theCa²⁺ ionophore ionomycin andinhibited by DIDS, dithiothreitol, niflumic acid, and tamoxifen.Expression in human trachea and lung suggests that hCLCA2 may play arole in the complex pathogenesis of cystic fibrosis.