Nucleocytoplasmic transport and nuclear pore disruption in ALS/FTD

项目总结 C9ORF72的GGGGCC六核苷酸重复序列扩展(HRE)是最常见的遗传原因 家族性肌萎缩侧索硬化症(ALS)和额颞部痴呆(FTD)，尽管基础疾病 机制定义不明确。多项研究，包括我们自己的，支持一种功能获得机制 HRE介导的神经毒性。扩展的重复序列可能产生隔离RNA结合的有毒RNA 蛋白质。它们也可以通过重复相关的非ATG翻译(RANT)被翻译成有毒的二肽 重复蛋白(Dprs)。HRE RNA和dprs都被假设为介导C9-ALS/FTD的神经毒性。 我们实验室和其他实验室最近的多项研究表明，核孔和/或核孔的破坏 核质转运是酵母、苍蝇和诱导多能细胞神经变性的主要原因 (IPS)C9-ALS/FTD模型。此外，我们最近的研究，使用C9-ALS/FTD果蝇模型，人类 来自C9-ALS患者的iPS神经元和人类C9-ALS CNS组织表明，核质 转运缺陷可能是ALS/FTD发病机制可治疗的基本途径。 该提案将全面调查C9ORF72 HRE破坏的机制 利用包括C9-ALS Fly在内的几种互补模型进行核质转运和核孔 以及小鼠模型和来自C9 ALS/FTD患者的iPS神经元和脑组织，并调查 调节核浆转运可能是ALS/FTD的一种治疗策略。(1)我们会决定 运动神经元和神经胶质细胞核孔复合体的形态和生化组成， 并在Fly、iPS、小鼠模型和人脑中用C9-ALS/FTD研究鼻咽癌的病理特征。鲜为人知 关于CNS鼻咽癌，包括细胞类型之间的差异，以及最终鼻咽癌成分如何， 在C9-ALS/FTD模型中，核孔蛋白表达异常。因此，理解和理解 鼻咽癌和核质转运在中枢神经系统和疾病模型中是根本重要的 剖析病理学的本质。(2)然后我们将研究核质转运的机制 C9-ALS/FTD的中断。我们假设干扰了鼻咽癌和/或核质转运功能 由于核输出序列的核丢失和/或胞浆积聚而导致神经变性 (NE)装载C9-ALS的苍蝇、小鼠和iPS型号的货物。(3)因此，我们将决定 C9-ALS/FTD模型中核质转运中断的后果。(4)最后，我们将确定 如果恢复核质转运挽救C9-ALS/FTD的神经退行性变 可能对人体有用的新型化合物。

PROJECT SUMMARY A GGGGCC hexanucleotide repeat expansion (HRE) in C9ORF72 is the most common genetic cause of familial amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), though the underlying disease mechanism is poorly defined. Multiple studies, including our own, support a gain-of-function mechanism of neurotoxicity mediated by the HRE. Expanded repeats may generate toxic RNAs that sequester RNA-binding proteins. They may also be translated via Repeat-Associated Non-ATG Translation (RANT) into toxic dipeptide repeat proteins (DPRs). Both HRE RNA and DPRs are hypothesized to mediate neurotoxicity in C9-ALS/FTD. Multiple recent studies from our lab and others suggest that disruption of the nuclear pore and/or nucleocytoplasmic transport is a primary cause of neurodegeneration in yeast, fly, and induced pluripotent cell (iPS) models of C9- ALS/FTD. In addition, our recent studies, using a C9-ALS/FTD Drosophila model, human iPS neurons derived from C9-ALS patients, and human C9-ALS CNS tissues, suggest that nucleocytoplasmic transport defects may be a fundamental pathway for ALS/FTD pathogenesis amenable to therapy. This proposal will comprehensively investigate the mechanism by which the C9ORf72 HRE disrupts nucleocytoplasmic transport and nuclear pores utilizing several complementary models including C9-ALS fly and mouse models and iPS neurons and brain tissue from C9 ALS/FTD patients, and investigate whether modulation of nucleocytoplasmic transport may be a therapeutic strategy for ALS/FTD. (1) We will determine the morphological and biochemical composition of the nuclear pore complex (NPC) in motor neurons and glia, and characterize NPC pathology in C9-ALS/FTD in fly, iPS, mouse models and human brain. Little is known about CNS NPCs including differences between cell types and ultimately how the NPC constituents, nucleoporins, are dysregulated in C9-ALS/FTD models. Therefore, understanding the basic characteristics of the NPC and nucleocytoplasmic transport in the CNS and in disease models is fundamentally important to dissecting the nature of pathology. (2) We will then investigate the mechanism of nucleocytoplasmic transport disruption in C9-ALS/FTD. We hypothesize that disrupted NPC and/or nucleocytoplasmic transport function causes neurodegeneration due to nuclear loss and/or cytoplasmic accumulation of nuclear export sequence (NES) containing cargo in fly, mouse, and iPS models of C9-ALS. (3) Therefore, we will determine the consequences of nucleocytoplasmic transport disruption in C9-ALS/FTD models. (4) Finally, we will determine if restoration of nucleocytoplasmic transport rescues neurodegeneration in C9-ALS/FTD by employing a series of novel compounds that may have human utility.