In view of the importance of the myelin sheath in the maintenance of a functionally normal brain, studies into the mechanism by which the membrane is formed are particularly important. Yet the mechanism of myelin assembly and the regulation of this process is an important neurobiological problem which is poorly understood. The overall objective of the proposed work is to examine, in detail, the expression of myelin basic protein (MBP) genes in normal mice and in dysmyelinating mutants which we have shown to have altered MBP metabolism. We propose to isolate and sequence cDNAs which correspond to the four mouse MBPs in order to establish the structural relationship among the 4 MBP mRNAs as well as the proteins themselves. Isolation and subcloning of the 4 MBP cDNAs will also provide probes with which to study MBP mRNA metabolism in the nucleus following transcription of the genes in normal and mutant mice. Several lines of evidence suggest that the MBP mRNAs are translated with different efficiencies and experiments are proposed to examine this in greater detail using inhibitors of specific steps of protein synthesis. Any differences in translational efficiencies will be related to structural differences noted in the primary sequences of the 4 MBP cDNAs. Since so little is known about MBP mRNA metabolism, experiments are also proposed to measure the levels of MBP mRNA precursors in the nuclei of normal and mutant mice, and to determine whether there occurs a build-up of precursor molecules in these mutants so that MPB mRNA does not reach cytoplasmic ribosomes. Finally using immunocytochemical and in situ hybridization techniques the mechanisms by which MBPs are transported from their sites of synthesis to their sites of assembly into myelin will be examined in normal mice and in quaking mice to determine where MPB assembly is blocked.
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