Proteolipid protein (PLP) is a highly conserved, integral myelin protein of the central nervous system. Modest over expression of the normal protein is, puzzlingly, lethal in humans and in animals at early ages. But absence of this major myelin protein, surprisingly, produces a very mild phenotype in null mice and in humans with PLP gene deletions. Mutations of the protein, even conservative amino acid substitutions, are usually lethal in humans and animals. What are the cellular and molecular factors that produce these different phenotypes? At the cellular level, the severity of the disease in the PLP mutants correlates quite well with the number of dying oligodendrocyte (Ols). At the molecular level, it has been postulated that defective intracellular transport of the mutant PLPs proteins is the underlying factor that induces OL death but the molecular mechanism(s) and signals that induce OL death have not been investigated. We hypothesize that the ubiquity-proteasome complex is involved in the degradation of PLP. The ubiquitin-proteasome system then triggers the programmed cell death (PCD) cascade. We will investigate using the in vitro systems how mutant and normal PLP proteins are processed by the ubiquitin-proteasome system and how OL death is induced. In vivo studies of the PCD cascade in Ols will be performed correlated with the in vitro data to assure that tissue culture mechanisms are an accurate reflection of the in vivo processes.
Specific Aim 1 : Determine if mutations of PLP protein induce PCD and if mutations of DM20 protein (an alternative splice product of the PLP gene) are as lethal as PLP; determine if both proteins are modified with ubiquitin in the endoplasmic reticulum; study PCD cascade in Ols.
Specific Aim 2 : Determine why over expression of normal PLP and normal DM20 proteins induce PCD and if these proteins are modified by ubiquitin. Examine the transport of PLP to the cell membrane by DM20.
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