INTERACTIONS OF NEURAL CAMS WITH EXTRACELLULAR PROTEINS

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Cell adhesion molecules (CAMs) and extracellular matrix (ECM) molecules play important roles in cell adhesion and migration during neural histogenesis. The long-term objective of this work is to understand the roles of these proteins in adhesion and migration of neural cells during development. Ng-CAM is expressed on neurons and Schwann cells but not on astroglia. Perturbation experiments with specific antibodies indicate that Ng-CAM is involved in neuron.astroglia adhesion and migration of neurons along Bergmann glia. Its function is also important for neuron-neuron adhesion, axonal growth, and axonal fasciculation. Ng-CAM binds to Ng-CAM on neurons and to distinct ligands on astroglia (heterophilic binding). Recent studies indicate that Ng-CAM interacts with ECM molecules that are associated with astroglia including the 1D1 and 3F8 chondroitin sulfate proteoglycans and laminin. The specific aims of thiN proposal are a) to characterize heterophilic ligands for Ng-CAM and other neural CAMs, b) to identify and analyze regions of the Ng-CAM molecule that are involved in heterophilic binding, and c) to analyze the potential roles of these interactions in adhesion and migration of neural cells. The specificity of interactions between neural CAMs and these ECM molecules will be further investigated using other ECM molecules including proteoglycans, laminin, cytotactin, and fibronectin. Specific antibodies against ECM ligands for Ng-CAM will be prepared and used to compare their localization in tissues with Ng-CAM and N-CAM, and to determine their cellular origin using biosynthesis experiments in culture. To map different structural and functional regions within the Ng-CAM molecule, protein chemical, molecular genetic, and immunological techniques will be used to identify domains in Ng-CAM that bind to specific ligands including the 1D1 and 3F8 proteoglycans, and laminin. To evaluate potential roles of specific domains of Ng-CAM and the heterophilic ligands, protein fragments, for example, that represent defined regions of Ng-CAM and bind to certain ligands, as well as specific antibodies against Ng-CAM and ECM proteins, will be used in assays for neuronal adhesion, neurite growth, and migration of granule cells in cerebellar explants. The recent discovery that the 1D1 and 3F8 proteoglycans bind to neural CAMs and inhibit neuronal adhesion raises the possibility that CAMs on the cell surface function as receptors for proteoglycans and may be involved in signal transduction. Therefore, binding of radiolabeled proteoglycans to cells will be measured and the involvement of neural CAMs will be tested using specific antibodies against the CAMs; the results may provide clues for further studies of molecular mechanisms of cell "repulsion." The findings of these and related studies will form a basis for evaluating the role of interactions between neural CAMs and ECM proteins during normal development, and some of the reagents generated may be useful in therapeutic protocols to improve neuronal regeneration.