In vivo functions of possible tumor suppressor Dab2

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DESCRIPTION (provided by applicant): The Disabled-2 (Dab2) gene inhibits tumor growth through unknown mechanisms. There are 2 Dab2 protein forms, p96 and p67 that appear to be adaptor proteins. Some adaptor proteins regulate endocytosis and others function in signal transduction. One exciting idea that we'll test for Dab2 is that some proteins do both. Our overall goal is to define the molecular mechanisms underlying the in vivo functions of Dab2. We've found that p96 but not p67 binds to endocytic proteins and localizes to clathrin-coated pits. In addition, both p96 and p67 contain a PTB/PID domain that associates with the endocytosis signals of lipoprotein receptors, and a separate domain that binds to a non-muscle myosin, myosin VI, that has been implicated in endocytosis. Indeed, we've shown that lack of Dab2 in the kidney causes protein transport defects similar to those of mice lacking the lipoprotein receptor, megalin. Thus we hypothesize that Dab2 normally regulates megalin traffic. In addition, we've found that Dab2 has another important function prior to gastrulating, when p67 is the predominant form. Dab2 mutant embryos have a defect in an extraembryonic epithelium, the visceral endoderm, and this defect prevents induction of the anterior posterior axis. The phenotype appears to result from impaired signaling by Nodal, a TGFbeta family member. We hypothesize that p67 is involved in signaling in the visceral endoderm. Since p67 lacks signals for coated pit localization, it is possible it is directly involved in signal transduction. We will identify the defective signaling pathways in Dab2-mutant visceral endoderm, and identify the step requiring Dab2. We will test whether p96 is specialized to regulate kidney transport and p67 is specialized for signal transduction, or whether each form is multifunctional. Coupled with studies on subcellular localization of Dab2 proteins in kidney and visceral endoderm, this approach will provide evidence whether the embryonic requirement for Dab2 is an indirect consequence of a role in receptor trafficking, or whether p67 has a signaling function. We will investigate the mechanism by which Dab2 regulates megalin traffic in the kidney, especially the importance of binding to components of clathrin-coated pits and myosin VI. We've also found that Dab2 is phosphorylated in mitogen-stimulated cells by MAP kinase, and will investigate the significance of Dab2 phosphorylation in regulating biological responses.