The long-term objectives of this project are to elucidate the molecular mechanisms of kidney-specific gene expression and renal epithelial cell differentiation, and to understand how perturbations of these mechanisms cause abnormal kidney development in humans. In the previous project period, we cloned the promoter of a novel kidney-specific cadherin gene (Ksp-cadherin) and showed that a short region of the promoter correctly recapitulates the complete expression pattern of the gene in transgenic mice. The Ksp-cadherin promoter is active exclusively in tubular epithelial cells in the kidney and the developing genitourinary tract. In addition, the kidney-enriched transcription factors, HNF-l and HNF-3, were shown to bind to the promoter and stimulate transcriptional activity. The proposed studies will test the hypothesis that transcription factors that regulate tissue-specific expression of Ksp-cadherin play important roles in kidney-specific gene expression and renal epithelial cell differentiation. In the first specific aim, we wil develop a general method for kidney-specific gene targeting that will be used to inactivate the hepatocyte nuclear factor 113 gene (HNF-1b) in transgenic mice. Mutations of the human HNF-1b gene cause maturity-onset diabetes of the young (MODY) and renal developmental abnormalities. The proposed studies will test whether I-INF-1 b is required for renal tubulogenesis and kidney-specific gene expression, and will examine the effects of a dominant gain-of-function mutation of HNF- b on kidney development. The second specific aim is to identify and characterize additional transcription factors that regulate kidney-specific gene expression. The region of the Ksp-cadherin promoter that is required for tissue-specific expression will be narrowed down, and renal nuclear proteins that bind to the region will be identified. Binding proteins that are co-expressed with Ksp-cadherin in renal tubular epithelial cells will be characterized further. Taken together, these studies will provide new insights into the genetic regulation of renal epithelial cell differentiation and the pathogenesis of kidney abnormalities in MODY. Moreover, these studies will generate unique reagents that will be widely applicable to studies involving cell lineage analysis in the kidney and kidney-specific gene targeting.
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