Gangliosides mediate axon-myelin stability and inhibit neurite outgrowth.

摘要

As the major nerve cell sialoglycoconjugates, certain gangliosides (notably GD1a and GT1b) are functional ligands for myelin-associated glycoprotein (MAG), which is a sialic acid-dependent Ig-family member lectin (Siglec) that is highly enriched at the myelin-axon interface. This dissertation constitutes an investigation of the roles of ganglioside-MAG interactions to maintain long-term axon-myelin integrity and to inhibit axon outgrowth.; To study the functional roles of ganglioside-MAG binding, mice engineered to lack a key enzyme in complex ganglioside synthesis, GM2/GD2 synthase, were studied. These mice do not express GD1a, GT1b (or other complex gangliosides), instead expressing only two simple gangliosides, GM3 and GD3. They displayed decreased central myelination, axonal degeneration in both the central and peripheral nervous systems, and demyelination in peripheral nerves. Furthermore, they displayed progressive and selective loss of MAG in the central and peripheral nervous systems whereas the expression of other major myelin proteins was unaltered. At 6 weeks of age, the expression level of MAG in brain myelin prepared from GM2/GD2 synthase knockout mice was indistinguishable from control mice. However MAG expression fell to 30% of control in 1 year old mice. Northern blot analysis of MAG mRNA revealed no difference between knockout and control mice at any age, indicating that decreased MAG expression in knockout mice was not due to transcriptional regulation, and might possibly be due to increased degradation after the failure of target ganglioside binding.; Consistent with biochemical and morphological findings, GM2/GD2 synthase knockout mice displayed progressive behavioral neuropathies including deficits in reflexes, strength, coordination and balance. One-year old null mice walked in small labored movements and displayed tremor and catalepsy. Our findings establish an essential role for complex gangliosides in maintaining the integrity of axon-myelin structure, biochemistry and normal neural physiology.; We also propose that gangliosides GD1a and GT1b mediate MAG inhibition of axon outgrowth. Antibody-induced crosslinking of GD1a or GT1b mimics MAG inhibition. To study changes in gene expression after neurite outgrowth inhibition, gene expression was analyzed using DNA microarray technology. Cerebellar granule cells were treated under two inhibitory protocols including MAG-containing detergent extract of myelin and antibody-induced clustering of GD1a. Transcription of a limited number of genes was found to be altered using both inhibitory protocols. The ultimate goal is to provide targets for further biological study. This dissertation contains a full description of the approaches used to analyze and confirm altered gene expression.