The proposed experiments will elucidate functions of two pan- neurally expressed proteins, ELAV and APPL, in differentiation and maintenance of all neurons. The long term objective of studies related to ELAV is to understand mechanism of neuron-specific alternative splicing. Mechanisms of neuron-specific splicing will be investigated using ELAV, a neuron-specific RNA binding protein, and it~s downstream identified target, the regulated intron of the gene neuroglian. Specifically, these studies will elucidate the mechanism of neuron-specific alternative splicing of the nrg pre-mRNA by defining cis-elements involved in the regulated intron that bind ELAV and other nuclear factors, and assess their in vivo significance. Other neuron-specific factors that regulate this process will be identified. These studies will use reporter transgenes in vivo and in cell culture assays to assess splicing, biochemical approaches to study RNA-protein interactions, and genetic methods to identify other factors. The Drosophila APPL protein is a member of the beta amyloid precursor protein (APP) family of proteins. The first identified member of this family, APP, is implicated to have a role in Alzheimer~s disease. Although this family of proteins is widely expressed within the nervous system, the normal physiological roles of these proteins are not well understood. The hypothesis that the APPL protein in Drosophila neuron function sas a cell surface receptor to regulate neuronal processes and synapses will be tested. Other proteins that are involved in APPL-mediated processes will be identified. The experimental design will make extensive use of molecular and genetic approaches possible in Drosophila: use of wild type and in vitro mutagenized transgenes and transgenic animals, and targeted expression, to assay in vivo function. Immunocytochemical techniques and confocal microscopy will be used to study neuronal arbors and neurophysiological studies will be conducted study synaptic function. Since ELAV and APPL both belong to evolutionarily conserved protein families this analysis will contribute to the mechanistic understanding of the function of related proteins in humans.
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