Roles for TDP-43 and FUS in ALS Using Motor Neurons from Embryonic Stem Cells

TDP-43 和 FUS 在使用胚胎干细胞运动神经元的 ALS 中的作用

• 批准号: 8127590
• 负责人: Dara Ditsworth
• 金额: $ 4.62万
• 依托单位: LUDWIG INSTITUTE FOR CANCER RES LTD
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8127590
• 关键词: Adult; Affect; Amyotrophic Lateral Sclerosis; Biology; Brain; Cell Line; Cells; Complementary DNA; DNA; DNA-Binding Proteins; Disease; Disease Progression; ES Cell Line; Future; Gene Expression Profile; Genes; Goals; Human; Inherited; Lead; Light; Motor Neurons; Movement; Mus; Muscle; Mutation; Neurodegenerative Disorders; Neuroglia; Neurons; Onset of illness; Paralysed; Pathogenesis; Pathology; Pathway interactions; Patients; Peptides; Phenotype; Prions; Proliferating; Property; Proteins; RNA; RNA Processing; RNA-Binding Proteins; Role; Signal Pathway; Small Interfering RNA; Spinal Cord; Tetracyclines; Therapeutic; Toxic effect; Transgenic Mice; Weight; base; cell type; embryonic stem cell; gain of function; loss of function; mutant; nervous system disorder; premature; promoter; protein TDP-43; protein function; sarcoma; small hairpin RNA; success; therapeutic development

DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disorder in which premature loss of upper and lower motor neurons leads to fatal paralysis with a typical disease course of one to five years. Mutations in two DNA/RNA binding proteins, TAR DNA-binding protein (TDP-43) and Fused in Sarcoma (FUS) have recently been identified as primary causes of inherited ALS and have led to what is likely to be a paradigm shift in efforts to understand the pathogenesis of ALS. Yet there remains a need to examine the functions of these proteins in cell types relevant for ALS progression, both motor neurons and glial cells. While a contribution from surrounding cells is well established in inherited ALS caused by SOD1 mutations, the impact of non-cell autonomous mechanisms in disease progression following mutation in TDP-43 or FUS is completely unknown. This proposal seeks to use embryonic stem cells derived from existing transgenic mice to examine the consequences of either reduction or mutation of TDP-43 or FUS in purified motor neurons. Cell intrinsic consequences of alterations in these genes will be determined by inducing differentiation of embryonic stem cells into motor neurons which either lack or express mutant TDP-43 or FUS. Rather than take a candidate approach to pursue the few genes that have already been identified as targets of TDP-43 or FUS in proliferating cell lines, isolated motor neuron cultures will be used to identify the complete set of RNAs that are altered by loss of TDP-43 or FUS in Aim 1, and then in Aim 2 to ask whether any RNAs are altered upon mutation of TDP-43 or FUS and if so, if they are the same as those observed upon loss of function. By taking such a comprehensive and systematic approach from purified starting material it is likely that the specific targets and signaling pathways affected can be uncovered - discoveries that might elucidate underlying mechanisms for disease and provide a basis for future therapeutic developments. Success in these goals will lead to greater understanding of ALS disease mechanism and will provide rationale for future studies in glial cell types to determine whether manipulation of supporting non-neuronal cells would provide therapeutic benefit in ALS patients with mutations in TDP-43 or FUS. PUBLIC HEALTH RELEVANCE: Amyotrophic lateral sclerosis (ALS) is an adult-onset neurodegenerative disorder that affects neurons of the brain and spinal cord controlling muscle movement and leads to fatal paralysis within the course of one to five years. Mutations in two proteins have recently been identified as a primary cause of ALS, yet the mechanism by which they contribute to disease is unknown. This project seeks to understand the functions of normal or mutant forms of these proteins in purified motor neurons to uncover the specific pathways affected, and may have wider implications for other nervous system diseases with similar pathology.

描述（由申请人提供）：肌萎缩性侧索硬化症（ALS）是一种成人发病的神经退行性疾病，其中上下运动神经元的过早丧失导致致命性瘫痪，典型的病程为1至5年。两种DNA/RNA结合蛋白TAR DNA结合蛋白（TDP-43）和融合肉瘤（FUS）的突变最近被确定为遗传性ALS的主要原因，并导致了理解ALS发病机制的范式转变。然而，仍然需要检查这些蛋白质在与ALS进展相关的细胞类型（运动神经元和神经胶质细胞）中的功能。虽然在由SOD 1突变引起的遗传性ALS中，周围细胞的贡献是明确的，但TDP-43或FUS突变后非细胞自主机制对疾病进展的影响完全未知。 该建议试图使用来自现有转基因小鼠的胚胎干细胞来检查纯化的运动神经元中TDP-43或FUS的减少或突变的后果。通过诱导胚胎干细胞分化为缺乏或表达突变型TDP-43或FUS的运动神经元来确定这些基因改变的细胞内在后果。不是采取候选方法来追求已经在增殖细胞系中被鉴定为TDP-43或FUS的靶标的少数基因，而是将分离的运动神经元培养物用于鉴定在Aim 1中由于TDP-43或FUS的缺失而改变的RNA的完整集合，然后在Aim 2中询问在TDP-43或FUS突变时是否有任何RNA被改变，如果是，如果它们与功能丧失时观察到的相同。 通过从纯化的起始材料中采取这种全面和系统的方法，可能会发现受影响的特定靶点和信号通路-这些发现可能阐明疾病的潜在机制，并为未来的治疗开发提供基础。这些目标的成功将导致对ALS疾病机制的更深入理解，并将为未来在神经胶质细胞类型中的研究提供理论基础，以确定支持非神经元细胞的操作是否会在具有TDP-43或FUS突变的ALS患者中提供治疗益处。 公共卫生关系：肌萎缩性侧索硬化症（ALS）是一种成年发病的神经退行性疾病，其影响控制肌肉运动的大脑和脊髓的神经元，并导致在一到五年内致命的瘫痪。两种蛋白质的突变最近被确定为ALS的主要原因，但它们导致疾病的机制尚不清楚。该项目旨在了解这些蛋白质在纯化的运动神经元中的正常或突变形式的功能，以揭示受影响的特定途径，并可能对具有类似病理的其他神经系统疾病产生更广泛的影响。