EARLY EVENTS IN RENAL DEVELOPMENT

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The kidneys develops when the ureteric bud invades the nephrogenic mesenchyme and begins to secrete survival factors for the mesenchyme and induces it to differentiate into the epithelial of the nephron. We have generated a panel of monoclonal antibodies to induced metanephric mesenchyme and found that two of these antibodies inhibit kidney development. The first antibody blocks tubulogenesis and produces widespread apoptosis of the induced mesenchyme. We have purified its antigen, gp- 100 a membrane protein that is expressed in induced mesenchyme but whose expression fades in adult life. The cDNA of this protein shows that it is a novel gene that has important extracellular and intracellular signal transduction domains. During epithelial development it is expressed in the basal surface in a focal junction. Our First Aim is to complete its cDNA sequence, examine the type of junction it forms, clone the mouse and human homologues and identify its chromosomal location. Using the mouse genomic clone we will delete the gene and generate mutant mice and examine their phenotype. In biochemical studies, we will purify its extracellular ligand. The second antibody recognized an extracellular matrix protein that is initially produced by the ureteric bud and induced mesenchyme but later is restricted to mesangial and smooth muscle cells. The antibody inhibited localization of endothelial cells around the ureteric bud. We have purified and cloned the 46 kDa antigen for this antibody and generated new polyclonal antibodies. The protein has previously been cloned as a marker for aggressive carcinomas including lung, breast and colon. In our Second Aim, we will develop a functional assay for interaction between endothelial and mesangial cells to examine the role of this protein in migration, capillary network formation and angiogenesis. In the Third Aim, we will develop a three dimensional model of ureteric bud branching and renal morphogenesis based on confocal scanning microscopy. We will first generate transgenic mice that express gene fluorescent protein in their ureteric bud lineage. This less invasive approach will allow us to identify critical new morphological structures and the role of known inducing factors and their inhibitors in the generation of the three dimensional architecture of the kidney tubules and their relation to their individual body supply.