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 different species. Here we report the cloning of the second human homolog, hCLCA2, from a human lung cDNA library. Northern blot and RT-PCR analyses revealed additional expression in trachea and mammary gland. A primary translation product of 120 kDa was cleaved into two cell surface-associated glycoproteins of 86 and 34 kDa in transfected HEK-293 cells. hCLCA2 is the first CLCA homolog for which the transmembrane structure has been systematically studied. Glycosylation site scanning and protease protection assays revealed five transmembrane domains with a large, cysteine-rich, amino-terminal extracellular 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 pathogenesis of cystic fibrosis.
Gruber
A D
AD
Cancer Biology Laboratories, Department of Molecular Medicine, Cornell University College of Veterinary Medicine, Ithaca, New York 14853, USA.
Schreur
K D
KD
Ji
H L
HL
Fuller
C M
CM
Pauli
B U
BU
D000595:Amino Acid Sequence / Q000235:genetics; D001483:Base Sequence / Q000235:genetics; D001940:Breast / Q000378:metabolism*; D002460:Cell Line; D018118:Chloride Channels / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004594:Electrophysiology; D005260:Female; D006801:Humans; D008168:Lung / Q000378:metabolism*; D008969:Molecular Sequence Data; D014132:Trachea / Q000378:metabolism*
C120066:CLCA2 protein, human; D018118:Chloride Channels; D018076:DNA, Complementary