The long term objectives of this proposal are to understand how the T- box gene family of transcription factors regulate gene expression in the kidney and how they contribute to kidney development and function. The focus of the current proposal is TBX2 and TBX3, two closely related members of the T-box family that are highly expressed in the kidney. TBX3 mutations result in ulnar-mammary syndrome, a human disorder associated with renal malformations. Based on their sequence conservation and expression pattern, it is hypothesized that TBX2 and TBX3 either serve a similar function or act to differentially regulate the same target genes. The following complementary in vitro and in vivo approaches are designed to test these hypotheses.
AIM 1. TO DEFINE THE ROLE OF tbx2 and TBX3 in RENAL GENE REGULATION WE WILL (i) IDENTIFY DOWNSTREAM TARGETS BY MEASURING ALTERED GENE EXPRESSION IN STABLE TRANSFECTANTS AND (ii) DETERMINE IF TBX2 AND TBX3 FUNCTION MAY BE MODULATED BY INTERACTIONS WITH OTHER T-BOX PROTEINS. (i) Transfectants in which TBX2 or TBX3 expression has been either induced or inactivated will be analyzed using cDNA arrays. (ii) Structural studies have shown that at least one T-box protein binds DNA as a dimer. The ability of other T-box proteins expressed in the kidney to heterodimerize with TBX2 or TBX3 will be tested.
AIM 2. TO DETERMINE THE ROLE OF TBX2 AND TBX3 IN NEPHROGENESIS, WE WILL CHARACTERIZE PROTEIN EXPRESSION IN MURINE KIDNEYS AND ASSESS THE AFFECT OF ALTERING TBX2 EXPRESSION. The pattern of TBX2 and TBX3 protein expression in mature and immature kidneys will be important in determining whether these genes function cooperatively or antagonistically and will be analyzed by immunocytochemistry. Transgenics expression human TBX2 will be analyzed for alterations in gene expression or kidney morphology using cDNA arrays and other molecular and biochemical markers. The proposed studies will extend our knowledge of the T-box family of developmental regulatory proteins and the role these genes play in the kidney, as well as provide further insight into the molecular mechanisms governing the process of nephrogenesis and the maintenance of normal kidney function.
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