Szuchet S, Polak P E, Yim S H, Arvanitis D
Department of Neurology, University of Chicago, Illinois 60637.
Glia. 1988;1(2):141-50. doi: 10.1002/glia.440010206.
We have compared highly purified fractions of oligodendrocyte plasma membrane to myelin by one- and two dimensional gel electrophoresis and found them to be distinct. The major myelin proteins--proteolipid protein (PLP), DM-20, and myelin basic protein (MBP), which dominate the sodium dodecyl sulfate polyacrylamide gel electrophoresis pattern of myelin--were minor components of the plasmalemma. However, 2',3', cyclic nucleotide phosphodiesterase (CNPase) and myelin-associated glycoprotein (MAG) were represented equally in both membranes. Labeling the cells with various precursors followed by isolation of plasmalemma revealed that newly synthesized PLP, DM-20, CNPase, and MAG were incorporated into the plasma membrane of "floating" oligodendrocytes (i.e., nonattached to substratum). This was not so with MBP. Nevertheless, scattered patches of MBP were localized on the plasma membrane of intact cells using the immunogold method at the electron microscopic level. The data are consistent with the notion that MBP is not a constituent of the plasma membrane of mature oligodendrocytes (the MBP patches on intact cells are likely remnants from past association with myelin) but is rapidly associated with the plasmalemma of myelinating oligodendrocytes (i.e., attached cells). It is suggested that phosphorylation of MBP provides the triggering signal for plasma membrane association. In order to analyze the minor proteins in myelin and compare them to the plasma membrane by two-dimensional gel electrophoresis, myelin was extracted with chloroform:methanol to remove PLP, DM-20, and MBP. Even in the absence of PLP, DM-20, and MBP the pattern of extracted myelin still differed from that of plasmalemma indicating that their minor protein compositions were not the same. Myelin was characterized by a group of proteins that clustered at pI 5.5-6.5 and Mr 40,000-60,000 of which alpha-tubulins, beta-tubulins, and actin are part: the plasmalemma had tubulins and actin but in different proportions. Our findings indicate that in addition to PLP, DM-20, and MPB, myelin is also enriched relative to the plasmalemma in another group of proteins.
我们通过一维及二维凝胶电泳，将高度纯化的少突胶质细胞质膜组分与髓磷脂进行了比较，发现它们截然不同。髓磷脂的主要蛋白质——蛋白脂蛋白（PLP）、DM - 20和髓磷脂碱性蛋白（MBP），在髓磷脂的十二烷基硫酸钠聚丙烯酰胺凝胶电泳图谱中占主导地位，但在质膜中却是次要成分。然而，2',3',环核苷酸磷酸二酯酶（CNPase）和髓磷脂相关糖蛋白（MAG）在两种膜中的含量相当。用各种前体标记细胞，随后分离质膜，结果显示新合成的PLP、DM - 20、CNPase和MAG被整合到“漂浮”少突胶质细胞（即未附着于基质的细胞）的质膜中。MBP则并非如此。不过，在电子显微镜水平上，使用免疫金法可观察到完整细胞的质膜上有散在的MBP斑块。这些数据与以下观点一致：MBP不是成熟少突胶质细胞质膜的组成成分（完整细胞上的MBP斑块可能是过去与髓磷脂结合的残余物），但会迅速与正在形成髓鞘的少突胶质细胞（即附着细胞）的质膜结合。有人提出，MBP的磷酸化提供了质膜结合的触发信号。为了分析髓磷脂中的次要蛋白质，并通过二维凝胶电泳将它们与质膜进行比较，用氯仿：甲醇提取髓磷脂以去除PLP、DM - 20和MBP。即便没有PLP、DM - 20和MBP，提取的髓磷脂图谱仍与质膜图谱不同，这表明它们的次要蛋白质组成并不相同。髓磷脂的特征是一组聚集在pH值5.5 - 6.5、相对分子质量40,000 - 60,000的蛋白质，其中包括α - 微管蛋白、β - 微管蛋白和肌动蛋白：质膜中有微管蛋白和肌动蛋白，但比例不同。我们的研究结果表明，除了PLP、DM - 20和MPB外，相对于质膜，髓磷脂在另一组蛋白质中也更为丰富。
