The purpose of this project is to study the roles of two transcription factors, Cux-1 and Pod-1, in kidney development and renal cyst formation. Cux-1 is a homeodomain protein that functions as a transcriptional repressor of differentiation- specific genes. Studies using in situ hybridization have shown that Cux-1 is developmentally regulated in the kidney. Cux-1 is highly expressed during early stages of nephrogenesis then is down-regulated as nephrons mature. Expression of Cux-1 remains persistently elevated in polycystic kidneys from homozygous cpk mice. Down-regulation of Cux-1 may be required for normal nephrogenesis, and dysregulated expression of Cux-1 may contribute to cyst formation by interfering with nephron differentiation. To test these hypotheses, we will examine whether the expression of Cux-1 is increased in other forms of polycystic kidney disease. Genetic crosses will be performed to determine whether mutations of Cux-1 reduce cyst formation in cpk mutant mice. Cux-1 will be over-expressed in transgenic mice to determine whether increased expression is sufficient to cause cyst formation.
The second aim i s to study the role of a novel basic-helix-loop-helix protein, named Pod-1, in kidney development. Basic-helix-loop-helix (bHLH) proteins are transcription factors that are important in cell differentiation and tissue-specific gene expression. Pod-1, which is the first tissue-restricted bHLH protein that has been identified in the kidney, is expressed in developing podocytes and in mesenchymal cells at sites of epithelial-mesenchymal interaction. Antisense inhibition of Pod-1 blocks mesenchymal cell condensation and inhibits ureteric branching. Pod-1 deficient mice exhibit abnormalities in branching morphogenesis, glomerular and tubular differentiation, and vascular development. The proposed studies will characterize the biochemical and functional properties of Pod-1 in the developing kidney. Specific antibodies generated against Pod-1 will be used to determine the size and subcellular localization of the protein. The DNA-binding properties, protein-protein interactions, and transcriptional activity of Pod-1 will be determined. Studies will be performed to evaluate whether mesenchymal expression of Pod-1 affects branching morphogenesis. Collectively, these studies should provide fundamental insights into transcriptional regulation of kidney morphogenesis and cystogenesis.
