BIOCHEMISTRY AND REGULATION OF CADHERIN ACTIVITY

点击翻译按钮获取中文摘要

The cadherin cell adhesion molecules play very important roles in the development and maintenance of solid tissues. The regulation of cadherin adhesion activity at the cell surface is an important mechanism for controlling tissue morphogenesis. The cadherin cytoplasmic tail, the associated catenin polypeptides, and the actin cytoskeleton are important for cadherin-mediated adhesion, but how these elements control the adhesive activity of the extracellular homophilic binding domain is not understood. The overall goals of the proposed project are to understand the mechanisms that determine how cadherin cell adhesion proteins assemble to form a functional adhesive complex at the cell surface and how their adhesive functions at the cell surface are regulated by cytoplasmic factors. The specific aims are designed to address these issues at increasing levels of complexity; first to define the basic biochemical and homophilic-binding properties of cadherin extracellular domains, then to determine the contributions of oligomerization and cytoskeletal anchoring to cadherin adhesion activity, and finally to analyze the mechanisms underlying the regulation of cadherin binding and cell adhesion. Recent findings of the high resolution three dimensional structure of the N-terminal cadherin extracellular domain (EC1), demonstrating the existence of a parallel "strand" dimer with two adhesive interfaces, provide a framework for many of the proposed studies. C-cadherin, the major adhesion protein in the very early Xenopus embryo, will be the primary subject of this study. We have overexpressed and purified milligram amounts of a functionally active, soluble extracellular domain of C-cadherin. This protein and mutated forms of it will be used in many of the proposed experiments. The specific aims are to: 1) Analyze the homophilic binding properties of C-cadherin; 2) Determine the role of the parallel "strand dimer" interaction in the binding properties and cell adhesion functions of C- cadherin; 3) Determine which of the 5 extracellular cadherin domains of C- cadherin contribute to its homophilic binding interactions; 4) Investigate the roles of cytoskeletal anchoring, oligomerization, and catenin-binding in the control of the adhesive function of C-cadherin at the cell surface; 5) Investigate the mechanisms underlying the regulation of the adhesive function of C-cadherin in Xenopus blastomeres and cultured cells; 6) Determine whether the specific requirement for E-cadherin in the integrity of the ectoderm in Xenopus is determined by its extracellular domain or its cytoplasmic tail. The information gained from these studies should improve our understanding of the functions of the cadherins in tissue morphogenesis. The findings will have implications for understanding changes in the adhesive behavior of cells during tumor progression in morphogenetic defects associated with human congenital diseases.