The ultimate goal of our project is to understand the underlying molecular logic of nervous system development. We will approach this goal by studying glycoproteins containing carbohydrate epitopes which are expressed specifically by all Drosophila neurons (and perhaps even neuronal precursor cells). The two glycoproteins we will focus on are among the best candidates for genes which mark the neuronal cellular phenotype. The proteins have been identified initially as antigens by staining Western blots of tissue extracts with antibodies to horse radish peroxidase (HRP). Since these heterologous antibodies recognize only the carbohydrate moieties of the antigens we have purified the corresponding proteins from both adult and larval stage Drosophila, We have recently generated several different monoclonal antibodies which recognize the protein core of the larval antigen. These antibodies immunocytochemically stain only neurons in Drosophila embryo cultures indicating that the larval antigen is indeed a neuron specific gene. The monoclonal antibodies and/or amino acid sequence information to be derived from the purified proteins will be used to clone the cDNAs and genes for the antigens. We have completed studies describing the developmental appearance of the carbohydrate epitope(s) and now, with the availability of anti-protein core antibodies, we will correlate this information with the cellular distribution of protein(s) using immunocytochemistry. Once nucleic acid probers are obtained we plan to extend our observations to the mRNA level using in situ hybridization and Northern analysis. We also plan to test the functional significance of these genes by cloning and mapping their cis regulatory control elements. We also plan to analyze mutants for neurodevelopmental phenotypes. Information gained from our proposed studies should be helpful in understanding normal nervous system development and ultimately in providing a background for interpreting defective development which results from abnormal neuronal genes or gene expression patterns.
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