MICROTUBULES AND MYELINATION IN A MUTANT RAT

突变大鼠的微管和髓鞘形成

• 批准号: 6188018
• 负责人: IAN DAVID DUNCAN
• 金额: $ 26.59万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-07-01 至 2003-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6188018
• 关键词: axon; cellular polarity; disease /disorder model; glia; in situ hybridization; laboratory rat; microtubule associated protein; microtubules; mutant; myelin; myelination; myelinopathy; nerve /myelin protein; nervous system transplantation; northern blottings; oligodendroglia; protein biosynthesis; protein transport; tissue /cell culture; western blottings

Myelination of the central nervous system (CNS) is a complex developmental process, requiring exquisitely timed interactions between axons and the myelin forming cells, the oligodendrocyte, and the appropriate temporal expression of an array of myelin genes. The initial elaboration of the myelin sheath in the developing CNS, and the long-term maintenance of compact myelin, each place different demands on the myelin forming cell. However, both processes are likely to be influenced by the structural and transport properties of the oligodendrocyte cytoskeleton. This proposal will study a new and unique rat myelin mutant, the taiep rat, which develops a chronic progressive neurological disease as a consequence of a CNS myelin disorder, associated with a microtubular accumulation in oligodendrocyte. The long-term goals are to characterize this mutant and its molecular defect and use it to define the role of microtubule and associated proteins in myelin formation and maintenance. To achieve these long-term goals, five specific aims have been planned: 1) The phenotype of the mutant will be characterize using morphological and morphometric approaches to study gla, cell kinetics, myelin, and the onset of the microtubule accumulation. 2) To determine whether environmental factors in the taiep CNS play any role in the development of the microtubule accumulation, cross transplantation experiments will be carried out. These will study the structure and function of taiep oligodendrocyte transplanted into the CNS of other myelin mutants, and of normal cells transplanted into the taiep CNS. 3) The polarity, stability and organizing centers of microtubule in the taiep rat oligodendrocyte will be compared to normal cells to determine whether an abnormal organization of tubules might interfere with transport events within the cell. 4) The role of microtubule regulatory proteins in the taiep mutation will be studied using antibodies to these proteins by Western blots and immunolabelling. Absence, or an over-or under- expression, or ectopic expression, of such a protein will be determined. Messenger RNAs for these proteins will also be examined by Northern blots and in situ hybridization. Finally, 5) To study the effects of accumulation of microtubule in taiep oligodendrocyte, cultures of these cells will be immunolabelled for myelin proteins known to co-localize with the cytoskeleton, and myelin protein mRNAs will be localized by in situ hybridization. Similar experiments will be performed on normal oligodendrocyte in which the cytoskeleton has been pharmacologically manipulated.

中枢神经系统（CNS）的髓鞘形成是一个复杂的过程， 发展过程，需要精确的时间互动之间 轴突和髓鞘形成细胞，少突胶质细胞，以及 髓鞘基因阵列的适当时间表达。 初始 在发育中的CNS中髓鞘的加工，以及长期的 维持紧密的髓鞘，每种对髓鞘的需求不同 形成细胞 然而，这两个过程都可能受到 少突胶质细胞骨架的结构和运输特性。 这项提案将研究一种新的和独特的大鼠髓鞘突变体，taiep 大鼠，其发展为慢性进行性神经系统疾病， 中枢神经系统髓鞘疾病的后果，与微管 在少突胶质细胞中积累。 长期目标是描述 这种突变体及其分子缺陷，并用它来定义 微管和相关蛋白在髓鞘形成和维持中的作用。 为实现这些长期目标，规划了五个具体目标： 1)突变体的表型将使用形态学方法表征。 和形态计量学的方法来研究gla，细胞动力学，髓鞘， 微管积累的开始。 2)以确定是否 环境因素在taiep CNS的发展中起任何作用 的微管积累，交叉移植实验将 执行。 这些将研究taiep的结构和功能 移植到其他髓鞘突变体CNS中的少突胶质细胞，和 移植到大脑中枢神经系统的正常细胞。 3)极性， taiep大鼠微管的稳定性和组织中心 将少突胶质细胞与正常细胞进行比较，以确定 肾小管的异常组织可能会干扰运输事件 在细胞内。4)微管调节蛋白在细胞凋亡中的作用 TAIEP突变将使用针对这些蛋白质的抗体进行研究， 蛋白质印迹和免疫标记。 缺席，或超过或低于- 将测定这种蛋白质的表达或异位表达。 这些蛋白质的信使RNA也将通过北方印迹进行检查 和原位杂交。 （5）研究 Taiep少突胶质细胞中微管的积累，这些细胞的培养物 细胞将免疫标记已知共定位的髓磷脂蛋白 髓磷脂蛋白mRNA将被定位在细胞骨架中， 原位杂交 类似的实验将在正常的 少突胶质细胞，其中细胞骨架已被破坏 被操纵了