Project 2

项目总结/摘要：项目2 最近的研究已经证明了核运输和应力颗粒/液体相变的缺陷， C9orf72重复扩增模型系统中的潜在早期事件。然而，实际上我们对 这些最初的发现与ALS和FTD之间的关系，这两种疾病是由相同的突变引起的。的 六核苷酸重复扩增导致异常RNA以及二肽重复（DPR）蛋白质， 通过非ATG重复相关翻译（RAN翻译）。研究一致认为， 破坏核质转运，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：同样的突变如何不同地涉及皮层神经元， 脊髓运动神经元在临床上不同的疾病？新出现的数据还将基于压力颗粒的生物学与 非神经元细胞中的核质转运。然而，这是否与CNS神经元或神经胶质有关， 不清楚我们将采用基于人诱导多能干细胞（iPSC）衍生的皮质神经干细胞的模型系统。 来自明确定义的C9orf72 FTD、C9orf72 ALS或C9orf72 ALS患者的神经元和运动神经元 和FTD探索潜在的生物学机制，可能有助于解释选择性 神经元和/或神经胶质细胞类型在这些广泛不同的疾病。具体而言，在目标1中，我们将 全面评估和比较核孔复合体（NPC）和核质 C9orf72 ALS/FTD iPSC衍生的运动和皮质神经元中的转运。了解不同的 运动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， 在剖析C9orf72 ALS和FTD的疾病特异性发病机制。在目标2中，我们建议阐明 在iPSC衍生的运动和皮质神经元中通过C9orf72重复扩增改变的分子途径。 使用"组学"分析询问人类皮质与脊髓C9orf72神经元将提供对细胞的深入了解。 特定缺陷和减轻神经元损伤的机会。最后，在目标3中，我们将研究 C9orf72 ALS/FTD中细胞应激与核孔复合物改变之间的关系。 确定应激颗粒对核质转运功能障碍的贡献将揭示新的 用于修复核孔复合体的治疗干预方法。总之，了解 疾病病理机制与C9orf72的神经元亚型特异性作用之间的联系 核孔复合体的重复扩增、核质转运和基因表达是 对于我们理解C9orf72 ALS/FTD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.