Toxic Neuronal Death by Glyceraldehyde-3-Phosphate Dehydrogenase and Mitochondria

This proposal was designed to examine the role of glyceraldehyde-3-phosphate dehydrogenase GAPDH, mitochondrial permeability and mitochondrial membrane potential Delta PSI M in toxic neuronal apoptosis. Mitochondria have been shown to play a critical decisional role in some forms of apoptotic cell death. Determining the signaling pathways for specific forms of toxin induced apoptosis might reveal potential pharmacological targets which could be manipulated to slow or alleviate the apoptosis. To address these issues, we proposed a series of experiments which include an in vitro study of neurons and neuron-like cells that will be immunoreacted to determine specific toxic insults that involve changes in GAPDH levels or subcellular distribution, changes in Delta PSI M and increased mitochondrial membrane permeability with the release of factors that signal for apoptotic degradation. We proposed to examine cell lines of PCI2 and 3T3 cells transfected so as to produce inducible GAPDH upregulation. The direct and joint actions of GAPDH and NAD will be examined using a cell free system of nuclear, mitochondrial and cytosolic subfractions. Finally we proposed to examine whether GAPDH alters the transcription andor translation of specific subsets of genes andor their RNA targets using a lysate system.