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Keywords:

  • Myelin basic protein;
  • Oligodendrocyte;
  • Expression vector;
  • Membrane localization;
  • Subcellular fractionation

Abstract: Myelin basic protein (MBP) is a major structural component of myelin. It is expressed exclusively in myelinating glia (oligodendrocytes in the CNS and Schwann cells in the PNS) and is localized to the cytoplasmic surface of the plasma membrane and myelin membrane produced by these cells. The work described here concerns the mechanism of plasma membrane localization of MBP in myelinating glial cells and whether it involves differentiated functions specific to these cells or general functions of plasma membrane assembly common to all cells. To this end, the subcellular localization of endogenous MBP in mouse oligodendrocytes was compared with that of transiently expressed MBP in monkey fibroblasts (Cos-1 cells) transfected with an MBP expression vector containing cDNA for rat 14K MBP. The steady-state levels of MBP-specific RNA and of MBP poly-peptide expressed in the transfected fibroblasts were comparable to the levels expressed in oligodendrocytes in primary culture. MBP localization was analyzed in whole cells by immunofluorescence and in specific intracellular compartments by subcellular fractionation. The results show that MBP expressed in wild-type oligodendrocytes is localized to the plasma membrane. In contrast, MBP expressed in transfected fibroblasts appears dispersed in the cytoplasm and is distributed uniformly among the various subcellular fractions. Purified MBP added prior to subcellular fractionation to ho-mozygous shiverer oligodendrocytes (which express no MBP endogenously) is recovered predominantly in the plasma membrane fraction, whereas purified MBP added to mock-transfected fibroblasts exhibits a dispersed distribution among subcellular fractions. On the basis of these results, we propose a model whereby localization of MBP to the plasma membrane in oligodendrocytes involves interaction between the newly synthesized MBP polypeptide and specific MBP receptors which are present in oligodendrocyte plasma membrane, but not in fibroblast plasma membrane.