p25α is an oligodendroglial protein that can induce aggregation of α-synuclein and accumulates in oligodendroglial cell bodies containing fibrillized α-synuclein in the neurodegenerative disease multiple system atrophy (MSA). We demonstrate biochemically that p25α is a constituent of myelin and a high-affinity ligand for myelin basic protein (MBP), and in situ immunohistochemistry revealed that MBP and p25α colocalize in myelin in normal human brains. Analysis of MSA cases reveals dramatic changes in p25α and MBP throughout the course of the disease. In situ immunohistochemistry revealed a cellular redistribution of p25α immunoreactivity from the myelin to the oligodendroglial cell soma, with no overall change in p25α protein concentration using immunoblotting. Concomitantly, an ∼80% reduction in the concentration of full-length MBP protein was revealed by immunoblotting along with the presence of immunoreactivity for MBP degradation products in oligodendroglia. The oligodendroglial cell bodies in MSA displayed an enlargement along with the relocalization of p25α, and this was enhanced after the deposition of α-synuclein in the glial cytoplasmic inclusions. Overall, the data indicate that changes in the cellular interactions between MBP and p25α occur early in MSA and contribute to abnormalities in myelin and subsequent α-synuclein aggregation and the ensuing neuronal degeneration that characterizes this disease.