Ksp-cadherin gene promoter. I. Characterization and renal epithelial cell-specific activity
| Autor: | Congyi Li, Peter S. Aronson, Peter Igarashi, Stacey L. Nix, Sharon L. Karp, R. Brent Thomson, Reza Zanjani, Dilys A. Whyte |
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| Rok vydání: | 1999 |
| Předmět: |
Transcription
Genetic Physiology DNA Mutational Analysis Molecular Sequence Data 5' flanking region CAAT box Gene Expression Biology Kidney Mice Gene expression Animals Cloning Molecular Promoter Regions Genetic Transcription factor Epithelial polarity Base Sequence Activator (genetics) Cadherin Chromosome Mapping Nuclear Proteins Epithelial Cells Promoter Cadherins Molecular biology Gene Deletion |
| Zdroj: | Experts@Minnesota |
| ISSN: | 1522-1466 1931-857X |
| DOI: | 10.1152/ajprenal.1999.277.4.f587 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
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