This application proposes to identify and characterize proteins that are specific to glia. Previous studies have focussed on the structure and genetics of proteolipid protein (PLP), a major component of CNS myelin that is expressed specifically in oligodendrocytes. We have isolated full-length cDNA clones for two rat pLp mRNAs and determined their nucleotide sequences. Characterization of cDNA clones for a second myelin proteolipid, DM-20, indicated that the sequence of DM-20 is identical to that of PLP, except for the absence of 35 amino acids from the major internal hydrophilic domain of PLP. The mRNAs encoding PLP and DM- 20 are derived from the single PLP gene by alternative RNA splicing. Studies of the dysmyelinating mouse mutant jimpy demonstrated that jimpy PLP mRNA contains a deletion of 74 nucleotides relative to wild type PLP mRNAs and that this defect is generated by aberrant RNA processing rather than by deletion of genomic sequences. We have also begun selection of clones corresponding to other glial-specific proteins, from cDNA libraries constructed from cultured astrocytes and from optic nerve. It is the aim of the proposed studies to: 1. Determine the structure of the PLP gene in the rat, analyse its regulation, and investigate the alternative splicing of PLP gene transcripts. 2. Characterize the mutation in the jimpy allele of the PLP gene. 3. Investigate the expression of the PLP gene in the dysmyelinating mutant mouse jimpy-msd and in the myelin deficient (md) rat. 4. Analyse the cellular expression of PLP and DM-20 in normal and mutant animals by in situ hybridization. 5. Determine the topological organization of the PLP polypeptide and the mechanism of its insertion in the oligodendrocyte membrane. 6. Select and characterize cDNA clones corresponding to further glial-specific or glial enriched mRNA expressed in oligodendroyctes and astrocytes. Analysis of the expression of the PLP gene in normal and mutant animals will lead to an understanding of the regulation of this gene, particularly during myelination. Characterization of PLP and other glial proteins will allow us to define the extent of genetic similarity between glial cells and provide a basis for determining the functions of glia, a means for classifying glia and their subtypes and ultimately may improve our understanding of the involvement of glia in neurological disease.
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