Neurotrophins are a group of molecules involved in the development, maintenance, and death of cells comprising the nervous system. There are two classes of receptors for the family of neurotrophins, the trk (tropomyosin-receptor kinase) family of tyrosine kinases and the p75 NTR receptor. It is the role of p75NTR in regulating trk activation, a process termed receptor transmodulation, that is the subject of this thesis. We discovered that p75NTR activation can inhibit trkA signaling. Following published work that p75NTR activation can lead to the production of intracellular ceramide (Dobrowsky et al., 1994; Dobrowsky et al., 1995), an intracellular signaling molecule, we determined that exogenous ceramide mimicked these effects of BDNF. The data suggest that BDNF may use the ceramide signaling cascade to inhibit trkA activation. To help elucidate the nature by which these results may occur, we examined the phosphoamino acid content of trkA. This analysis revealed that BDNF and C2 ceramide both induce an increase in the serine phosphorylation of the trkA intracellular domain when compared to a lack of either agent. Based on our findings, the presence of a signaling mechanism where trkA activity can be modulated by p75NTR has been postulated. After completing dose-response experiments with C2-ceramide, a time-course experiment was proposed. It was discovered that incubation of PC12 cells with greater than 20 muM C2-ceramide caused an increase in trkA receptor activation. Furthermore, this effect was observed in the absence of ligand, and found to be the result of an increase in trkA dimerization. These data suggest that chronic exposure to exogenous ceramide causes an increase in trkA dimerization, and leads to the activation of downstream targets. The revelation that GM1-ganglioside, a sialic acid containing glycosphingolipid and product of the sphingomyelin cycle, increases trkA dimerization correlates with the temporal nature of our observed changes in trkA activation. It is possible that either ceramide becomes metabolized into gangliosides or that gangliosides and ceramide share some intermediate molecule that mediates the effect. (Abstract shortened by UMI.)

Barker, Philip A. (Supervisor)