Ksp-cadherin gene promoter. I. Characterization and renal epithelial cell-specific activity

Dilys A. Whyte, Congyi Li, R. Brent Thomson, Stagey L. Nix, Reza Zanjani, Sharon L. Karp, Peter S. Aronson, Peter Igarashi

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Abstract

Kidney-specific cadherin (Ksp-cadherin, cadherin 16) is a novel, kidney- specific member of the cadherin superfamily that is expressed exclusively in the basolateral membrane of renal tubular epithelial cells. To characterize the Ksp-cadherin gene promoter, a λ bacteriophage clone containing 3.7 kb of the proximal 5' flanking region of the mouse Ksp-cadherin gene was isolated. The transcription initiation site was mapped by RNase protection assays and 5' rapid amplification of cDNA ends, and a 709-bp intron was identified within the 5' untranslated region. The proximal 5' flanking region was 'TATA- less' but contained other consensus promoter elements including an initiator (Inr), GC boxes, and a CAAT box. Potential binding sites were identified for transcription factors that are involved in tissue-specific gene expression including activator protein-2 (AP-2), hepatocyte nuclear factor-3 (HNF-3), basic helix-loop-helix (bHLH) proteins, CCAAT/enhancer-binding protein (C/EBP), and GATA factors. Transfection of luciferase reporter plasmids containing 2.6 kb of the 5' flanking region markedly increased luciferase activity in renal epithelial cells (MDCK and mIMCD-3) but not in mesenchymal cells (NIH 3T3 and MMR1). Deletion analysis identified an 82-bp region from - 31 to -113 that was essential for promoter activity in transfected renal epithelial cells. Electrophoretic mobility-shift assays showed that mIMCD-3 cells contain nuclear proteins that bind to this region of the promoter. Mutational analysis showed that sequences within the HNF-3 consensus site and CAAT box were involved in protein binding and promoter activity. We conclude that the proximal 5' flanking region of the mouse Ksp-cadherin gene contains an orientation-dependent promoter that is kidney epithelial cell specific. The region of the promoter from -113 to -31 is required for transcriptional activity and contains binding sites for nuclear proteins that are specifically expressed in renal epithelial cells.

Original languageEnglish (US)
JournalAmerican Journal of Physiology - Renal Physiology
Volume277
Issue number4 46-4
Publication statusPublished - Oct 1999

Cite this

Whyte, D. A., Li, C., Thomson, R. B., Nix, S. L., Zanjani, R., Karp, S. L., ... Igarashi, P. (1999). Ksp-cadherin gene promoter. I. Characterization and renal epithelial cell-specific activity. American Journal of Physiology - Renal Physiology, 277(4 46-4).