The major goal of this research project is to elucidate the function of the small GTPase Rap1 and the molecular mechanism(s) by which this GTPase exerts its effects on epithelial morphogenesis. Recent studies in Drosophila and in mammalian systems have implicated Rap1proteins in the control of cell spreading, adhesion and morphogenesis, but little is known about the molecular mechanisms that mediate Rap1 function. The mammalian junctional protein AF-6 was identified as a Rap1-interacting protein, but functional analyses of mammalian Rap1 and AF-6 proteins, and of their interaction, have been hampered by the fact that these proteins are largely refractory to loss- and/or gain-of-function analysis. Hence, we decided to first gain insights into their functions using a genetically more tractable Drosophila system. Our preliminary results from Drosophila implicate Canoe (the Drosophila homologue of AF-6) as an effector of Drap1 (the Drosophila homologue of Rapl) in dorsal closure (DC), a morphogenetic process that relies on elongation and migration of epithelial sheets in the embryo. Furthermore, we have recently identified a putative novel regulator of Drap1 in DC, designated DPDZ-GEF, whose vertebrate homologue is associated with epithelial adherens junctions, just as Rap1 and AF-6/Canoe are. Notably, the Drosophila homologues of Rap1 and AF-6 (Drap1 and Canoe) share a similar physical interaction, and the mammalian proteins can functionally substitute for their Drosophila counterparts. This proposal aims to define the function of Drap1, Canoe, and DPDZ-GEF in regulating cell shape changes, cell-cell adhesion, and coordinated movement of an epithelial sheet during DC. Our specific aims are: 1) To analyze the function of Drap1 and Canoe in DC and cell-cell adhesion, and 2) To characterize the function of DPDZ-GEF. Specific aim 1 is designed to provide a thorough assessment of the mechanisms that underlie the Drap1/Canoe interaction and the role of Canoe as a junctional protein in the process of DC. In aim 2, the biochemical and biological functions of the DPDZ-GEF and its epistatic relationship to Drap1 will be assessed. These studies offer a comprehensive set of approaches to define the roles of Drap1, Canoe, and DPDZ-GEF in regulating cell-cell adhesion and coordinated epithelial cell movements, and will likely lead to a better mechanistic understanding of these processes and facilitate the design of experiments addressing the role of the mammalian homologues in similar processes. Ultimately, our studies are relevant to a number of medically important issues, including wound healing and the homeostasis of epithelial cells and sheets.