organogenesis involves a programmed pattern of gene expression which controls cellular differentiation and morphogenesis. The developing kidney is one of the best characterized models for the study of the molecular control of organogenesis. The WT-1 gene, originally identified as a tumor suppressor gene associated with Wilms' tumors. A tumor of the kidney which occurs during childhood, is required during the inductive events at the beginning of kidney formation, and is also required for gonad formation. WT-1 is a zinc finger transcription factor with a proline-glutamine rich trans- regulatory region. The defined phenotype of the original WT-1 mutation, combined with a detailed knowledge of WT-1 action in vitro, provides an opportunity to genetically dissect at an organismic level how transcription factors regulate developmental processes in mammals. This grant proposes to undertake a more detailed mutagenic analysis to further our understanding of WT-1 function during development. Novel gene targeting approaches will be used to express mutated forms of WT-1 from the WT-1 genomic locus. Abnormal developmental phenotypes obtained will be interpreted based on morphological changes, alterations in expression of other known markers of urogenital development, and on previous studies of WT-1 function in vitro, where similar WT-1 mutations have altered its action as a transcriptional regulator. Recently, an interaction has been defined between WT-1 and the p53 tumor suppressor gene, yet p53 mutant mice have no obvious defects in urogenital development. The developmental role of p53 remains unclear; most p53 -/- embryos are normal, but a subset have neural tube defects, partially dependent on genetic background. Crossing the WT-1 and p53 mutations provides a means for determining the in vivo significance of the p53-WT-1 interaction, and provides a route for determining a developmental function of p53. Embryos carrying mutations in the WE-1 gene that lead to abnormal urogenital phenotypes will be analyzed by in situ hybridization, immunohistology, and organ culture. RNA subtraction libraries will be constructed to find novel genes expressed in urogenital development.
