Synthesis Myelin and Incorporation of Myelin Polypeptides into CNS

The distribution of newly synthesized proteolipid protein (PLP, 23 kdaltons) and myelin basic proteins (MBPs, 14-21.5 kdaltons) was determined in microsomal and myelin fractions prepared from the brainstems oi 10-30 d-old rats sacrificed at different times after an intracranial injection of 3~S-methionine. Labeled MBPs were found in the myelin fraction 2 min after the injection, whereas PLP appeared first in the rough microsomal fraction and only after a lag of 30 rain in the myelin fraction. Cell-free translation experiments using purified mRNAs demonstrated that PLP and MBPs are synthesized in bound and free polysomes, respectively. A mechanism involving the cotranslational insertion into the ER membrane and subsequent passage of the polypeptides through the Golgi apparatus is consistent with the lag observed in the appearance of the in vivo-labeled PLP in the myelin membrane. Newly synthesized PLP and MBPs are not proteolytically processed, because the primary translation products synthesized in vitro had the same electrophoretic mobility and N-terminal amino acid sequence as the mature PLP and MBP polypeptides. It was found that crude myelin fractions are highly enriched in mRNAs coding for the MBPs but not in mRNA coding for PLP. This suggests that whereas the bound polysomes synthesizing PLP are largely confined to the cell body, free polysomes synthesizing MBPs are concentrated in oligodendrocyte processes involved in myelination, which explains the immediate incorporation of MBPs into the developing myelin sheath. The myelin sheath has long been recognized as a rich source of plasma membranes with a relatively simple biochemical composition. In the central nervous system the sheath is derived from the plasma membrane of the oligodendrocyte (c.f. reference 44), a cell that during myelinogenesis greatly expands its surface while developing cytoplasmic processes that wrap around neighboring axons. Compact myelin is formed when the oligodendrocyte cytoplasm is extruded from the myelinating process, thus allowing for a close apposition of adjacent plasma membrane surfaces. In the resulting periodic multilamellar structure, apposed cytoplasmic aspects of the oligodendrocyte plasma membrane form what, in thin-section electron microscopy, is recognized as a major dense line, whereas the extracellular membrane faces form a thinner intraperiod line. The major integral membrane protein of CNS myelin is the myelin proteolipid protein (PLP, or lipophilin, Mr 23 kdaltons) (21, 23) a proteolipid apoprotein that comprises -50% of the total protein (19) and can only be released from the membrane bilayer by treatment with strong detergents or certain organic solvents (23). Myelin also contains a comparable amount of one or a set of peripheral membrane proteins that are easily extracted by acids or high salt treatment, and are known as myelin basic proteins (MBPs), (c.f. reference 11). In the mouse, four different MBP polypeptides have been identified (4). The two major ones (18.5 kdaltons and 14 kdaltons), which were previously identified in certain rodent species (39) have been designated L and S (for large and small) and have the same amino acid sequence, except for the deletion of 40 amino acids near the carboxy terminal of S (17, 40). The two other species (21.5 kdaltons and 17 kdaltons) differ from the L and S proteins only by the presence of an additional polypeptide segment common to both (4). Although the disposition of PLP in the bilayer is not known, and the nature of the association of MBP with the myelin membrane is not well understood, it seems likely that specific interactions between the major myelin proteins play a role in myelinogenesis and that the regular arrangement of these polypeptides within the mature myelin sheath has important functional implications. THE JOURNAL OF CELL BIOLOGY. VOLUM£ 95 NOVEMBER 1982 598-608 5 9 8 © The Rockefeller University Press • 0021-9525/82/11/0598/11 $1.00 on July 8, 2017 jcb.rress. org D ow nladed fom