Proteolipid protein (PLP) and PO are the major intrinsic proteins of CNS and PNS myelin respectively. These proteins account for 50-60% of the total myelin membrane proteins. We have been able to acylate PLP and PO with [3H]-palmitic acid in vivo and provided firm evidence that the labeled fatty acids are covalently linked to these proteins by ester bonds. These studies will be extended to examine a) the specificity of acylation in subcellular fractions isolated from rat brain, b) whether acylated PLP is produced by undifferentiated and/or differentiated oligodendrocytes, c) whether PLP is acylated in the cytosol of olidogendrocytes prior to its incorporation into the myelin membrane, d) whether all four fatty acids are present in oligodendrocytes and/or the myelin sheath during the critical period of myelination, e) the turnover of the fatty acid moiety and peptide backbone of PLP in developing and mature rat brain. Since we have developed a new procedure for the preparation of water-soluble acylated and deacylated forms of PLP, we will be able to determine what percent of total PLP is acylated, turnover rate of acylated and non-acylated forms of PLP. Furthermore, it will now be possible to generate monoclonal antibodies to the acylated and deacylated forms of PLP. This will enable us to explore the possibility that the peptide backbone, to which the fatty acid is linked, contains the antigenic site(s) and (f) the peptides and subsequently, the specific amino acids to which these labeled fatty acids are linked. Myelin isolated from neurologically-deficient quaking mouse brain contains a substantially reduced concentration of PLP; studies are designed to ascertain if the lack of PLP in myelin is due to the absence of the acylated form of PLP. These studies will be extended to other myelin-deficient mutants. Recently we secured the first evidence that PO is acylated in vivo, and methyl esters of palmitate, stearate, oleate and linoleate are covalently linked to PO. These studies will be extended to ascertain whether or not acylated PO is produced by undifferentiated and/or differentiated Schwann cells prior to the formation of compact myelin and the linkage of fatty acids to specific amino acids in PO. Since PO has been show to be glycosylated, sulfated, phosphorylated, and acylated, we will determine if sulfation, phosphorylation and acylation is inhibited without affecting protein synthesis by monensin, tunicamycin, and cerulenin.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS013464-08
Application #
3395189
Study Section
Neurology B Subcommittee 1 (NEUB)
Project Start
1976-12-01
Project End
1987-03-31
Budget Start
1985-04-01
Budget End
1986-03-31
Support Year
8
Fiscal Year
1985
Total Cost
Indirect Cost
Name
Washington University
Department
Type
Schools of Medicine
DUNS #
062761671
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Agrawal, H C; Agrawal, D (1991) Proteolipid protein and DM-20 are synthesized by Schwann cells, present in myelin membrane, but they are not fatty acylated. Neurochem Res 16:855-8
Agrawal, H C; Agrawal, D; Strauss, A W (1990) Cleavage of the P0 glycoprotein of the rat peripheral nerve myelin: tentative identification of cleavage site and evidence for the precursor-product relationship. Neurochem Res 15:993-1001
Agrawal, H C; Sprinkle, T J; Agrawal, D (1990) 2',3'cyclic nucleotide-3'-phosphodiesterase in peripheral nerve myelin is phosphorylated by a phorbol ester-sensitive protein kinase. Biochem Biophys Res Commun 170:817-23
Agrawal, H C; Noronha, A B; Agrawal, D et al. (1990) The myelin-associated glycoprotein is phosphorylated in the peripheral nervous system. Biochem Biophys Res Commun 169:953-8
Agrawal, H C; Sprinkle, T J; Agrawal, D (1990) 2',3'-cyclic nucleotide-3'-phosphodiesterase in the central nervous system is fatty-acylated by thioester linkage. J Biol Chem 265:11849-53
Agrawal, H C; Agrawal, D (1989) Effect of cycloheximide on palmitylation of PO protein of the peripheral nervous system myelin. Biochem J 263:173-7
Agrawal, H C; Agrawal, D (1989) Tumor promoters accentuate phosphorylation of PO: evidence for the presence of protein kinase C in purified PNS myelin. Neurochem Res 14:409-13
Bourdette, D N; Seil, F J; Meshul, C K et al. (1988) Antisera to an axolemma-enriched fraction have antiaxon and antimyelin effects in vitro. Ann N Y Acad Sci 540:423-6
Yoshimura, T; Agrawal, D; Agrawal, H C (1987) Cell-free acylation of rat brain myelin proteolipid protein and DM-20. Biochem J 246:611-7
Agrawal, H C; Agrawal, D; Yoshimura, T et al. (1987) In vitro acylation of myelin PLP and DM-20 in the quaking mouse brain. Neurochem Res 12:783-6

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