Myelin is a dynamic, functionally active membrane, sensitive to its environment including axons. During myelinogenesis, it is produced in prodigious quantities. It is a polarized membrane, with both macro- and microdomain organization. It actively maintains its unique structure and special relationship with axons. Among its interesting biochemical properties is a high concentration of specific glycosphingolipids, in particular galactocerebroside (GalC) and sulfatide. The galactolipids have been strongly implicated in the regulation of oligodendrocyte differentiation and myelin maintenance. In combination with cholesterol, current evidence indicates that these lipids are able to form microdomains in which a discrete set of proteins become associated in order to effect specialized functions often related to signal transduction. While a rather small number of """"""""major"""""""" myelin proteins are recognized, there is a plethora of poorly identified quantitatively """"""""minor"""""""" proteins. The function of both classes is poorly understood. This long term goal of this application is to develop a data base of the total complement of myelin proteins the Myelin Proteome, and to begin the task of associating function with each of these structural/metabolic components. This will be accomplished by first displaying the total myelin protein complement as a two-dimensional gel electrophoretic map, and identifying the individual members by immunolabeling, non-Edman mass spectrometry, and bioinformatics. This information will provide a database both for this project and for the myelin biology community. Second, a detailed analysis will be made of the proteins in the glycosphinogolipid-cholesterol microdomains of myelin, relating them to the total myelin proteome. A study will be made of their organization, and a hypothesis will be tested that proposes that there is a mosaic of distinguishable microdomains. Third, this information will be used to investigate the functional biology of these microdomains. A mouse model in which galactosylcerebroside and sulfide are not produced (the CGT-null mouse) will be analyzed in terms of its microdomains. In this mouse the nodes of Ranvier are altered; in what we propose are the molecular correlates of these morphological changes, we have shown that the association of a discrete set of proteins with the glycosphinogolipid microdomains is altered. Finally, we have shown that myelin-oligodendrocyte glycoprotein (MOG) partitions into these microdomains, and the experimental stimulation its entry in maturing oligodendrocytes results in the rapid tyrosine phosphorylation of specific proteins. The relationship of this to antiOMOG mediated allergic encephalomyelitis will be considered. It is expected that concepts derived from these investigations will help to better understand myelin biogenesis, maintenance and normal function, and help to design paradigms to promote myelin stability and remyelination in multiple sclerosis.
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