Malformations of urogenital-anorectal structures during embryonic development are frequently observed birth defects in humans. For instance, hypospadias, which is due to a failure of the urethral epithelial cells to tubularize, occurs in approximately 1 out of every 320 male births. The genes, proteins, and cell signaling networks that control development of urogenital and anorectal structures are poorly understood. The Eph family of receptor tyrosine kinases and their membrane- anchored ephrin ligands are highly conserved molecules that function in diverse cell-cell recognition events, including those required for axon pathfinding in the nervous system, in migration of neural crest cells and in vascular organization. Preliminary data is provided that show gene-targeted mice mutant for certain Eph receptors and ephrins display hypospadias and delayed septation of the cloaca and cloacal membrane, two common failures in midline fusion characteristic of abnormal urogenital-anorectal development. To explore these preliminary findings in greater detail, three specific aims are proposed: 1) To fully characterize the abnormal urogenital-anorectal development observed in the Eph and ephrin mutant mice; 2) To study and perturb the activities of Eph/ephrin signals in the urogenital system by developing in vitro and in vivo assays; and 3) To delineate the importance of Eph receptors and ephrins in human urogenital-anorectal development by documenting the expression of these molecules and by screening for mutations in ephrin genes in individuals born with hypospadias and other hindgut abnormalities. By combining the powers of mouse molecular genetics, biochemical analysis and clinical-based studies, the overall goal of this collaborative research project is to characterize in detail the cell-cell signal transduction events that are utilized in mammalian urogenital-anorectal development.