Schoenfeld T A, McKerracher L, Obar R, Vallee R B
Worcester Foundation for Experimental Biology, Shrewsbury, Massachusetts 01545.
J Neurosci. 1989 May;9(5):1712-30. doi: 10.1523/JNEUROSCI.09-05-01712.1989.
Five high-molecular-weight microtubule-associated proteins (MAPs) were identified in brain tissue in previous work from this laboratory (Bloom et al., 1984). These proteins were termed MAP 1A, 1B, 1C, 2A, and 2B. The MAP 1's differed from the MAP 2's, and showed little evidence of interrelationship on the basis of immunological and biochemical comparison. We report here that MAP 1A and MAP 1B are, in fact, related at the level of subunit composition. Immunoprecipitation of the individual MAPs showed that both contained low-molecular-weight subunits of Mr 30,000 and Mr 19,000 (light chains 1 and 3). An additional subunit, light chain 2 (Mr 28,000), was primarily found in preparations of MAP 1A. The light chains co-sedimented with microtubules after chymotryptic digestion of the MAPs. This suggested an association of the light chains with the microtubule binding domains of the MAPs, which are identified here as distinct fragments of Mr 60,000 for MAP 1A and 120,000 for MAP 1B. A panel of monoclonal anti-MAP 1A and anti-MAP 1B antibodies, including one that reacts with a common phosphorylated epitope, was used to examine the distribution of these proteins in the developing rat brain and spinal cord. MAP 1B was found to be abundant in the newborn brain and to decrease with development, in contrast to MAP 1A which increased with development. By immunohistochemistry MAP 1B was found to be highly concentrated in developing axonal processes in the cerebellar molecular layer, the corticospinal tract, the mossy fibers in the hippocampus, and the olfactory nerve. Of particular interest, the mossy fiber and olfactory nerve staining persisted in the adult, indicating continued outgrowth of the mossy fibers as well as olfactory nerve axons. MAP 1A staining was, in contrast, weak or absent in developing axonal fibers but moderate in mature axons and intense in developing and mature dendritic processes. Our results indicate that MAP 1A and MAP 1B are structurally related components of the neuronal cytoskeleton with complementary patterns of expression.
本实验室先前的研究工作（布鲁姆等人，1984年）在脑组织中鉴定出了五种高分子量微管相关蛋白（MAPs）。这些蛋白被命名为MAP 1A、1B、1C、2A和2B。MAP 1类蛋白与MAP 2类蛋白不同，基于免疫学和生化比较，几乎没有显示出相互关系的证据。我们在此报告，MAP 1A和MAP 1B实际上在亚基组成水平上是相关的。对单个MAPs进行免疫沉淀显示，两者都含有分子量为30,000和19,000的低分子量亚基（轻链1和3）。另一个亚基，轻链2（分子量28,000），主要存在于MAP 1A的制剂中。在对MAPs进行胰凝乳蛋白酶消化后，轻链与微管一起沉降。这表明轻链与MAPs的微管结合结构域有关联，在此处确定MAP 1A的微管结合结构域为分子量60,000的不同片段，MAP 1B的为120,000。一组抗MAP 1A和抗MAP 1B单克隆抗体，包括一种与常见磷酸化表位反应的抗体，用于检测这些蛋白在发育中的大鼠脑和脊髓中的分布。结果发现，与随发育增加的MAP 1A相反，MAP 1B在新生脑中含量丰富，并随发育而减少。通过免疫组织化学发现，MAP 1B高度集中在小脑分子层、皮质脊髓束、海马体中的苔藓纤维和嗅神经的发育中的轴突过程中。特别有趣的是，苔藓纤维和嗅神经的染色在成体中持续存在，表明苔藓纤维以及嗅神经轴突持续生长。相比之下，MAP 1A染色在发育中的轴突纤维中较弱或不存在，但在成熟轴突中中等强度，在发育中和成熟的树突过程中较强。我们的结果表明，MAP 1A和MAP 1B是神经元细胞骨架的结构相关成分，具有互补的表达模式。
