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Physiol. Genomics 3: 101-111, 2000;
1094-8341/00 $5.00
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Received 16 August 1999; accepted in final form 2 June 2000.
Physiological Genomics 3:101-111 (2000)
1094-8341/00 $5.00 © 2000 American Physiological Society

Cloning a chloride conductance mediator from the apical membrane of porcine ileal enterocytes

K. J. GASPAR1, K. J. RACETTE1, J. R. GORDON2, M. E. LOEWEN1 and G. W. FORSYTH1

1 Department of Veterinary Physiological Sciences
2 Department of Veterinary Microbiology, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada S7N 5B4

Gaspar, K. J., K. J. Racette, J. R. Gordon, M. E. Loewen, and G. W. Forsyth. Cloning a chloride conductance mediator from the apical membrane of porcine ileal enterocytes. Physiol Genomics 3: 101–111, 2000.—Attempts to attribute ileal brush-border chloride conductance to specific proteins were pursued by screening a porcine intestinal cDNA library. A 0.94-kb clone was identified on expression screening with a monoclonal antibody that inhibited enterocyte brush-border chloride conductance. Further screening approaches led to the isolation of a 3.1-kb full-length sequence called pCLCA1, consistent with the identification of a 2.9-kb transcript through Northern analysis. This sequence had significant homology to the CLCA gene family of calcium-regulated chloride channels, especially to hCLCA1. However, a strong A-kinase consensus phosphorylation site in a predicted cytoplasmic loop of the protein was a notable difference from the hCLCA1 gene product. Several porcine exocrine epithelial tissues, including ileum, trachea, and the major salivary glands express pCLCA1 mRNA. In situ hybridization studies localized the expression of pCLCA1 mRNA to the crypt and villus epithelia of porcine ileum, whereas tracheal expression was observed in both surface epithelium and submucosal glands. In situ expression of pCLCA1 in mouse 3T3 cells induces an ionomycin-dependent chloride conductance activity in these cells.

chloride channel; cystic fibrosis transmembrane conductance regulator; brush-border membrane; intestinal secretion; ileum




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