Studies will examine the signaling mechanisms and function of the 75 kDa neurotrophin receptor, p75/NTR and several structural homologs of P75/NTR. Experiments will test the hypothesis that neurotrophins activate NF-kappaB by a signaling pathway involving p75/NTR- dependent activation of the small G protein Rac, which stimulates superoxide production. A putative novel ligand for p75/NTR will be cloned and functionally characterized. The possible involvement of t he protease TACE in release of a soluble p75/NTR ectodomain from the cell surface will be assessed, by comparing p75/NTR processing in wildtype mouse fibroblasts and fibroblasts from mice lacking the TACE gene. Two structural homologs of p75/NTR will be characterized, expressing these proteins by transfection of various mammalian cell lines with suitable expression vectors. Homolog NRH1 will be expressed in CHO cells and assessed for ability to bind neurotrophins. The capacity of NRH1- mediated signaling to activate NF-kappaB and/or Rac will be assessed,, and the capacity of NRH1 to act as a subunit of a heteromeric p75/NTR receptor complex will be assessed, and the capacity of NRH1 to act as a subunit of a heteromeric p75/NTR receptor complex will be assessed. The possible existence of a mammalian homolog of Xenopus NRH1 will be sought by cDNA library screening. NRH2 encodes a protein resembling the product of the putative TACE-mediated cleavage of P75/NTR. The capacity of NRH2 to act as a dominant negative inhibitor of p75rt/NTR signaling will be assessed. Functions of NRH2 (and NRH1 if present in mice) will be assessed by gene-targeted mutations in mice. The possible existence of a NRH2 splice variant possessing a p75/NTR- like ligand binding will be sought by cDNA library screening. The patterns of developmental expression of P75/NTR, NRH1 and NRH2 will be examined in Xenopus and mice, to assess whether the opportunity for functional interactions exist.
