The PI is investigating the function of neurturin, a novel neurotrophic factor purified from Chinese hamster ovary cell conditioned media on the basis of its ability to promote the survival of sympathetic neurons in vitro. Subsequent to obtaining partial amino acid sequence of the purified protein, a cDNA encoding neurturin was isolated. Sequence analysis revealed that it is related to a distant member of the TGF-B superfamily, glial cell line-derived neurotrophic factor (GDNF), which promotes survival of midbrain dopaminergic and motor neurons. Comparison of the neurturin and GDNF sequences has revealed several highly conserved domains which may allow us to identify additional neurotrophic factors, just as the discovery of BDNF allowed the identification of NT-3 and NT-4/5. To pursue the biological functions of neuturin, recombinant neurturin will be produced and characterized with regard to its survival promoting activities. To characterize the interaction of neurturin with its receptor, binding and crosslinking studies will be performed. The identification of the neurturin receptor will be approached by first determining whether it is a known or structurally related member of the Ser/Thr kinase receptor family, such as those which interact with other TGF-B family members. If it is not, the receptor will be isolated by ligand-affinity chromatography or, by expression cloning. The effects of neurturin on sympathetic neuron neurotransmitter phenotype will be examined by monitoring expression of genes encoding neuropeptides and enzymes involved in neurotransmitter synthesis. Chimeric mutants generated by swapping domains between neurturin and GDNF will be analyzed to identify the particular neurturin domains required for promoting sympathetic neuronal survival and, potentially, for interacting with its receptor (s). To elucidate the physiological role of neurturin, the phenotypes of transgenic mice either deficient in neurturin, or in which neurturin is ectopically expressed, will be examined. Special attention will be given to the development and function of the nervous system in these mice, both in its normal and injured state. Because of the wide range of activities noted for members of the TGF-B family, we will also perform a global analysis of these animals as neurturin may influence other cell types as well. The studies outlined in this and the accompanying proposal of the IRPG will provide a solid foundation of basic information regarding the biological role of neurturin. As neurotrophic factors are now considered as potential therapeutic agents for neurodegenerative diseases, such as Alzheimer's as well as for more acute conditions, understanding the function of neurturin could have a major impact on the future treatment of these afflictions.
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