mTOR complexes in oligodendrocyte differentiation

描述(申请人提供)：少突胶质细胞在中枢神经系统中用髓鞘包裹轴突，以促进动作电位传导。发育过程中轴突髓鞘的形成包括少突胶质细胞的增殖、分化和髓鞘的产生。控制这一过程的细胞内信号机制才刚刚开始被阐明。我们实验室以前的数据表明，哺乳动物靶标雷帕霉素(MTOR)对少突胶质细胞的分化是必不可少的。MTOR是PI3K/Akt的下游靶点，通过与接头蛋白Raptor(MTORC1)或Rictor(MTORC2)的复合体调节多种细胞类型的细胞生长和增殖。MTORC1和mTORC2存在于少突胶质细胞前体细胞中，先前的数据表明每个复合体在分化过程中具有不同的功能。我的假设是，这两个mTOR复合体在少突胶质细胞分化过程中具有基本而不同的功能，并且mTOR信号对重新髓鞘形成和发育髓鞘形成都是必不可少的。通过在原代少突胶质细胞培养中使用siRNA敲除复杂的特定蛋白，将在体外表征mTORC1和mTORC2的敲除效果。携带FLOXED-mTOR和CNP-Cre转基因或可诱导的PLP-Cre转基因的双基因小鼠将在体内为少突胶质细胞特异性敲除mTOR创造条件。这些系统将被用来研究mTOR特异性缺失对分化和髓鞘形成的影响。这些研究的完成将阐明mTOR在体内少突胶质细胞分化、髓鞘形成和再髓鞘形成中的作用。第一个目的是研究mTORC2在调节分化过程中整合的转录因子中的作用。AIM 2将评估mTORC2对细胞骨架的调节并处理副产物。AIM 3将利用体内模型来确定mTOR基因敲除在发育过程中和脱髓鞘损伤后对分化和髓鞘形成的影响。在新生儿缺氧缺血和多发性硬化症中，少突胶质细胞的分化被破坏，这说明需要详细了解相关的机制过程。这些研究将进一步加深对少突胶质细胞分化的了解，并为影响少突胶质细胞和髓鞘形成的神经病理的治疗提供靶点。 公共卫生相关性：本申请建议确定mTOR信号在少突胶质细胞分化和髓鞘产生中的作用。轴突的髓鞘包裹对于轴突的传导是必不可少的，在新生儿缺氧缺血和多发性硬化症中会受到破坏。了解mTOR的作用将为治疗提供靶点，以促进这些中枢神经系统病理中的少突胶质细胞分化和促进重新髓鞘形成。

DESCRIPTION (provided by applicant): Oligodendrocytes envelop axons with a myelin sheath in the central nervous system to facilitate action potential conduction. The establishment of axon myelination during development involves oligodendrocyte proliferation, differentiation, and production of myelin. The intracellular signaling mechanisms governing this process are only beginning to be elucidated. Previous data in our laboratory demonstrated mammalian target of rapamycin (mTOR) as essential for oligodendrocyte differentiation. mTOR, a downstream target of PI3K/Akt, regulates cell growth and proliferation in a number of cell types through complexes with adaptor proteins raptor (mTORC1) or rictor (mTORC2). mTORC1 and mTORC2 are present in oligodendrocyte progenitors and previous data indicate different functions for each complex during differentiation. My hypothesis is that the two mTOR complexes have essential and distinct functions during oligodendrocyte differentiation and that mTOR signaling is essential for remyelination as well as developmental myelination. Through the use of siRNA knockdown of complex specific proteins in primary oligodendrocyte cultures the effects of knockdowns of mTORC1 and mTORC2 will be characterized in vitro. Bigenic mice carrying floxed-mTOR and either a CNP-Cre transgene or an inducible PLP-Cre transgene will be created for oligodendrocyte specific knockdown of mTOR in vivo. These systems will be used to investigate the effects of specific deletion of mTOR on differentiation and myelination. Completion of these studies will elucidate how mTOR functions in oligodendrocyte differentiation, myelination and remyelination in vivo. The first aim will investigate the role of mTORC2 in the regulation of transcription factors integral to the differentiation process. Aim 2 will assess mTORC2 regulation of the cytoskeleton and process outgrowth. Aim 3 will utilize in vivo models to determine the effects of mTOR knockdown on differentiation and myelination during development and following a demyelinating injury. Oligodendrocyte differentiation is disrupted in neonatal hypoxia-ischemia and in Multiple Sclerosis, illustrating a need for a detailed understanding of the mechanistic processes involved. These studies will further the understanding of oligodendrocyte differentiation and provide targets to pursue for therapeutic treatments of neuropathologies that impact oligodendrocytes and myelin formation. PUBLIC HEALTH RELEVANCE: This application proposes to determine the role of mTOR signaling in oligodendrocyte differentiation and production of myelin. Myelin wrapping of axons is essential for their conductance and is disrupted or damaged in neonatal hypoxia-ischemia and in multiple sclerosis. Understanding the effects of mTOR will provide targets for therapeutic treatment to enhance oligodendrocyte differentiation and promote remyelination in these central nervous system pathologies.