NGF binds to and activates the protein tyrosine kinase gp 140prototrk. Expression of this receptor is required for at least some responses to NGF. Three outstanding issues are addressed in the present work. First, we determined whether expression of gp 140prototrk is required for all neuronal NGF responses. Second, we examined the role of gp 140prototrk in NGF binding and internalization. Third, we addressed the utility of NGF-nonresponsive PC12nnr5 cells for study of the NGF mechanism. In contrast to wild-type PC12 cells, PC12nnr5 cells do not express endogenous gp 140prototrk. We therefore asked whether they possess other defects that compromise NGF signaling pathways. To answer these questions, we transfected PC12nnr5 cells with a cDNA encoding full- length human gp 140prototrk and isolated cell lines permanently expressing the receptor. Introduction of trk rescued all of the many and varied NGF responses assessed, including enhanced protein tyrosine phosphorylation, induction of immediate-early and neural-specific genes, neurite outgrowth and regeneration, maintenance of survival in serum-free medium, and stimulation of AChE activity. In contrast to PC12nnr5 cells, the trk-transfected lines also bind and internalize NGF with wild-type PC12 cell characteristics. These findings indicate that gp 140prototrk is required for many, if not all, responses of neuronal cells to NGF and is necessary for proper NGF binding and internalization. Additionally, as no signaling defect other than the absence of trk expression was revealed in PC12nnr5 cells, this work supports the utility of this line for genetic dissection of the NGF mechanism of action.