Project 2

项目摘要/摘要：项目2 最近的研究记录了核运输和应力颗粒/液体相变的缺陷，如 C9orf72中潜在的早期事件重复了扩张模型系统。然而，人们实际上对此知之甚少 这些初步发现与ALS和FTD之间的关系，这两种疾病是由相同的突变引起的。这个 六核苷酸重复序列扩增导致异常RNA和二肽重复序列(DPR)蛋白 通过非ATG重复关联翻译(RAN翻译)。一项共识的研究表明这种突变 破坏核质运输，但这种缺陷的机制和细胞类型特异性仍不清楚。 这就引出了一个重要的问题：相同的突变如何不同地涉及皮质神经元和 脊髓运动神经元在临床不同疾病中的作用？新出现的数据也将应激颗粒生物学与 非神经细胞的核质转运。然而，无论这是否与中枢神经系统神经元或神经胶质有关， 不清楚。我们将使用一个基于人类诱导多能干细胞(IPSC)来源的皮质的模型系统 明确的C9orf72 FTD、C9orf72 ALS或C9orf72 ALS患者的神经元和运动神经元 和FTD来探索潜在的生物学机制，这可能有助于解释选择性 在这些差异很大的疾病中，神经元和/或神经胶质细胞的类型。具体地说，在目标1中，我们将 全面评估和比较核孔复合体(NPC)和核质的变化 C9orf72 ALS/FTD IPSC来源的运动神经元和皮质神经元的转运。了解以下方面的差异 运动神经元和皮质神经元核孔复合体将定义基本的神经元特异性生物学，并可能有助于 在剖析C9orf72 ALS和FTD的疾病特异性发病机制中。在目标2中，我们建议澄清 C9orf72重复扩增改变了IPSC来源的运动神经元和皮质神经元中的分子通路。 使用组学分析询问人类皮质和脊髓C9orf72神经元将提供对细胞的洞察- 减轻神经元损伤的具体缺陷和机会。最后，在目标3中，我们将调查 C9orf72 ALS/FTD细胞应力与核孔复合体改变的关系 确定应激颗粒对功能障碍的核质转运的贡献将发现新的 修复核孔复合体的治疗干预方法。总而言之，理解 C9orf72的神经元亚型特异性效应与疾病发病机制之间的联系 核孔复合体的重复扩张、核质运输和基因表达 对于我们理解C9orf72 ALS/FTD的发病机制至关重要。

PROJECT SUMMARY/ABSTRACT: PROJECT 2 Recent studies have documented defects in nuclear transport and stress granule/liquid phase transitions as potentially early events in C9orf72 repeat expansion model systems. However, little is actually known about how these initial discoveries relate to ALS vs. FTD, two diseases caused by the same mutation. The hexanucleotide repeat expansion leads to both aberrant RNA as well as dipeptide repeat (DPR) proteins made via non-ATG repeat associated translation (RAN translation). A consensus of studies suggests the mutation disrupts nucleocytoplasmic transport but the mechanism and cell type specificity of this defect remain unclear. This leads to the important question: how does the same mutation differentially involve cortical neurons versus spinal motor neurons in clinically distinct diseases? Emerging data also links stress granule-based biology to nucleocytoplasmic transport in non-neuronal cells. However, whether this is relevant to CNS neurons or glia is unclear. We will employ a model system based on human induced pluripotent stem cell (iPSC)-derived cortical neurons and motor neurons from well-defined patients with either C9orf72 FTD, C9orf72 ALS or C9orf72 ALS and FTD to explore the underlying biological mechanism that may serve to explain the involvement of selective neuronal and/or glial cell types in these widely differing diseases. Specifically, in Aim 1 we will comprehensively assess and compare alterations in the nuclear pore complex (NPC) and nucleocytoplasmic transport in C9orf72 ALS/FTD iPSC-derived motor and cortical neurons. Understanding the differences in motor and cortical neuron nuclear pore complexes will define fundamental neuron-specific biology and may aid in dissecting the disease-specific pathogenesis in C9orf72 ALS and FTD. In Aim 2, we propose to elucidate molecular pathways altered by C9orf72 repeat expansions in iPSC-derived motor and cortical neurons. Interrogating human cortical versus spinal C9orf72 neurons using “omics” analytics will provide insight into cell- specific defects and opportunities for mitigating neuronal injury. Finally, in Aim 3, we will investigate the relationship between cellular stress and alterations in the nuclear pore complex in C9orf72 ALS/FTD. Determining the contribution of stress granules to dysfunctional nucleocytoplasmic transport will uncover novel approaches to therapeutic interventions for repairing the nuclear pore complex. In summary, understanding the connection between disease pathomechanisms and the neuronal subtype-specific effects of the C9orf72 repeat expansion on the nuclear pore complex, nucleocytoplasmic transport, and gene expression are essential to our understanding of C9orf72 ALS/FTD pathogenesis.