Mechanisms and Functions of MBP mRNA Transport in Oligodendrocytes

描述(申请人提供)：在中枢神经系统中，少突胶质细胞将轴突包裹在许多同心圆层的髓鞘中，这一过程对于轴突中有效的电信号是必不可少的。在多发性硬化症和脑瘫等疾病中， 少突胶质细胞不能形成致密的髓鞘，髓鞘碱性蛋白(MBP)缺乏适当的远端定位。MBP是少突胶质细胞中表达最高的mRNA，是少突胶质细胞表达水平的10倍以上。它也是一种特殊的蛋白质，它的mRNA必须被运输到少突胶质细胞的远端突起才能被翻译。然而，MBP mRNA转运的机制和体内功能仍不清楚。该项目旨在确定MBP mRNA在货物水平的运输调节是否对髓鞘形成是必要的。为了观察MBP mRNA在原代原代少突胶质细胞中的转运，将发展现代成像工具，如RNA结合报告蛋白MS2，以检测和分析MBP mRNA的运动性。此外，将使用两种独立的质谱学方法来识别MBP mRNA颗粒中参与其运输的蛋白质。运输调节因子通常是支架蛋白，具有与多种马达蛋白结合的能力。因此，MBP mRNA转运的候选调节因子将通过它们与发动机蛋白以及siRNA敲除策略的结合能力来确认。最后，将制作两种不同的小鼠模型，以确定MBP mRNA转运是否对体内髓鞘形成是必要的。这项拟议的项目将阐明运动蛋白如何与信使核糖核酸结合，并确定在少突胶质细胞成熟和髓鞘形成过程中关键过程的分子机制，并将对神经胶质细胞生物学、分子马达、RNA蛋白质生物学和髓鞘疾病等领域具有重要意义。重要的是，由于远端MBP蛋白表达和致密髓鞘形成在髓鞘疾病中失败，这些发现可能揭示刺激疾病髓鞘形成的新的治疗靶点。

DESCRIPTION (provided by applicant): In the central nervous system, oligodendrocytes ensheath and wrap axons in many concentric layers of myelin, a process that is essential for efficient electrical signaling in axons. In diseases such as multiple sclerosis and cerebral palsy, oligodendrocytes fail to form compact myelin and proper distal localization of myelin basic protein (MBP) is absent. MBP is the most highly expressed mRNA in oligodendrocytes by over 10-fold. It is also a special protein who's mRNA must be transported to distal processes of oligodendrocytes before it can be translated. The mechanism and in vivo function of MBP mRNA transport, however, has remained elusive. This project aims to determine whether regulation of MBP mRNA transport at the cargo level is necessary for myelination. In order to visualize MBP mRNA transport in living primary oligodendrocytes, modern imaging tools, such as the RNA-binding reporter protein MS2, will be developed for the detection and analysis of MBP mRNA motility. Furthermore, two independent mass spectrometry approaches will be used to identify proteins in the MBP mRNA granule that are involved in its transport. Regulators of transport are often scaffolding proteins that have the ability to bind to multiple motor proteins. Thus, candidate regulators of MBP mRNA transport will be confirmed via their ability to associate with motor proteins as well as with siRNA knockdown strategies. Finally, two different mouse models will be made to determine whether MBP mRNA transport is necessary for myelination in vivo. The proposed project will elucidate how motor proteins associate with mRNA cargos and identify the molecular mechanism underlying a pivotal process in oligodendrocyte maturation and myelination and will have important implications for the fields of glial cell biology, molecular motors, RNA protein biology, and myelin diseases. Importantly, since distal MBP protein expression and compact myelin formation fail in myelinating diseases, these findings may reveal novel therapeutic targets for stimulating myelination in disease.