Differential expression and MAL-dependent targeting of the L-MAG and S-MAG isoforms to myelin membranes

Many degenerative diseases of the nervous system, including Multiple Sclerosis and peripheral neuropathies, are triggered by an impaired interaction between the axons and their surrounding myelin sheaths. The cause for this disturbed axon-myelin interaction, and the secondary neuronal damage that produces the clinical symptoms, lies in a primary defect of the myelin sheath. The myelin sheath itself is formed and maintained by oligodendrocytes and Schwann cells, the myelinating glial cells of the central and peripheral nervous system, respectively. The isoforms of the myelin-associated glycoprotein (MAG) are thought to be potential key elements of axon-myelin interaction, since these immunoglobulin-like cell signalling proteins are known to be localized in the periaxonal and paranodal myelin membranes. The MAG isoforms each display one of two possible intracellular C-termini as a result of alternative mRNA splicing. The C-terminus of the large isoform (L-MAG) has been shown to mediate downstream signals via the non-receptor tyrosine kinase Fyn, while the Cterminus of the short isoform (S-MAG) is thought to interact with the glial cytoskeleton. We have investigated the regulation and differential expression of L- and S-MAG in oligodendroglial cells and in transgenic mice by the use of genomic constructs that encode individually green fluorescent protein-tagged MAG isoforms. In the oligodendroglial cells LMAG was the dominant isoform prior to the stimulation of cells with cyclicAMP, whereas upon cyclicAMP stimulation, S-MAG was predominantly expressed in cells exhibiting advanced morphological differentiation. The investigation of our transgenic mice revealed that the two MAG isoforms are differentially expressed in distinct fibre tracts of the striatum and that S-MAG seems to be predominantly expressed in the long projecting fibres of the corpus callosum. Thus, the two MAG isoforms appear not only to be differentially expressed during development and in the adult, but they seem to mediate isoform-specific aspects of the axon-myelin interaction in distinct regions of the adult brain. A major question in the formation and maintenance of the myelin-axon interaction concerns the coordinated targeting of myelin signalling molecules and lipids to the different myelin compartments. Recent results suggest that glycolipid-enriched microdomains, socalled 'lipid-rafts', are involved in special sorting and trafficking mechanisms of membrane proteins and lipids. Furthermore, they are thought to serve as platforms for signal transduction processes. This makes them to interesting candidates for axon-myelin interactions, as well as for interactions between the apposed myelin membranes. The integral membrane protein 'Myelin and Lymphocyte Protein' (MAL) is suggested to be involved in lipid-raft-mediated protein targeting and signalling in myelinating cells. Our investigation of adult brain tissue of MAL-deficient mice showed that the incorporation of particular myelin components, such as MAG, into myelin membranes was significantly reduced. Thus, the targeting of L- and S-MAG to the myelin membranes appears to be dependent on the lipid-raft protein MAL. Furthermore, the MAL-deficient mice showed several ultra structural alterations comparable to those of the MAG-deficient mice and that reflect an impaired axon-myelin interaction. Our data supports the idea that-raft mediated trafficking of myelin constituents, such as MAG, to the different myelin compartments is a major task of adult oligodendrocytes in the context of maintaining the axonal contact of the myelin sheath. With the use of the isoform specific tagged MAG expressing mice, it will be possible for the first time, to investigate their differential function in axon-glia interaction as well as their dependence on MAL in vivo.