The major goal of this project is to further understand the role of cell surface sulfogluguronyl carbohydrate (SGC) on glycolipids (SGGLs) and glycoproteins (SGGPs) in neural cell-cell recognition processes. Our previous studies indicate that SGC bearing molecules are transiently expressed on neural cells in a stage-specific manner, during development of the nervous system, and that the SGC itself appears to participate directly in the precisely choreographed processes of cell adhesion and migration. A malfunction in these processes could lead to improper nervous system development and mental retardation, as identified in several neuronal cell migration disorders. The present proposal is focused upon the expression, regulation and function of SGC and its endogenous lectins which recognize the carbohydrate on neural cell surfaces.
The specific aims are: 1. To characterize the SGC binding proteins (SBPs), (lectins) in the developing nervous system. Three major SBPs from brain have been isolated and purified. These proteins will be further characterized and their binding specificity with respect to the carbohydrate on SGGLs and SGGPs will be determined. The expression and regulation of the SBPs in different areas of the nervous system during development will be established by biochemical and immunocytochemical techniques. The role of SBPs in specific cellular adhesion processes will be evaluated with probing tools, such as appropriate antibodies and modified ligands to the proteins, in cerebellar cell culture systems. 2. To understand the regulation of expression of SGGLs and their biological function, the key regulatory enzyme GlcNAc-Transferase will be purified by novel photoaffinity labeling techniques. The antibodies to the enzyme and mRNA probes will be used for the precise localization of the synthesis of SGGLs at single cell level. 3. The genes for GlcNAc-Tr will be cloned for future studies on gene transfection in vivo. Immunocytochemical and biochemical analyses of the SGC and its lectins will determine which molecules are involved in the process of migration and maturation of the neuronal cells. Co-cultures of the immature cerebellar granule cells and astroglia will be used as model system to define the role of the carbohydrate in this process. Several mouse mutants with abnormal cellular migration and developmental defects in the nervous system will be used as in vivo models. The results will generate fundamental information on the role of SGC and its lectins in cellular interaction and migration in the developing brain and will define the molecular mechanisms involved in the processes of cell-cell interaction and cell differentiation.
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