The long term objectives of this project are to characteize the structure and biochemical mechanism of action of the nerve growth factor receptor, and to establish a thorough understanding of the distribution of NGF receptors in various mammalian tissues and of the biological roles of those receptors. Sepecific aims are to produce polyclonal antibodies reactive to various regions of the NGF receptor to be used for immunocytochemical localization of receptor. Antibodies would be made against synthetic peptides corresponsing to various portions of the receptors sequence, and/or to proteins of NGF receptors cDNA expressed in E. coli. Another aim is to produce in vitro mutated NGF receptor cDNA's and examine the consequence of expression of these in transfected cell lines, in order to define receptor regions involved in NGF binding, cytoskeletal linkage, interconversion between high and low affinity sites, glycosylation, etc. These studies would be complemented by biochemical characterization of specific receptor peptide fragments. Another aim is to determine whether various unorthodox sites of NGF receptor expression, such as lymph node follicular dendritic cells, myoepithelial cells, and fetal Purkinje cells are biologically important. This would be accomplished by examining the effects of NGF on isolated cell cultures and on permanent myoeipthelial cell lines. In addition to immunochemical studie sof NGF receptor distribution, receptor expression in various tissues would be examined by qunatiating NGF receptor mRNA levels, using NGF receptor cRNA or synthetic oligonucleotide probes for Northern blot analysis and or in situ hydridization. Also a cDNA encoding an NGF receptor cRNA synthetic oligonucleotide probes for Northern blot analysis and or in situ hybridization. Also a cDNA encoding an NGF receptor structural homolog would be sequence, and the possibility investigated that some apparent sites of NGF receptor expression may involve this homolog rather than the authentic NGF receptor.
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