Genetic analysis of UBQLN2-associated neurodegeneration in frontotemporal dementia

额颞叶痴呆中UBQLN2相关神经变性的遗传分析

• 批准号: 10157746
• 负责人: Randal Scot Tibbetts
• 金额: $ 170万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10157746
• 关键词: Address; Aging; Alleles; Alzheimer' s disease related dementia; Amyotrophic Lateral Sclerosis; Apoptosis; Applications Grants; Attenuated; Autopsy; Axonal Transport; Binding; Binding Proteins; Biochemical; Cell Culture Techniques; Cell Line; Characteristics; Chromosome Mapping; Chromosomes; Clinical; Colon Carcinoma; Cytology; Defect; Dementia; Dementia With Amyotrophic Lateral Sclerosis; Dependence; Development; Disease; Disease Pathway; Dissection; Dose; Drosophila genus; Drosophila melanogaster; Enhancers; Exhibits; Eye; Family; Frontotemporal Dementia; Functional disorder; Future; Gene Mutation; Genes; Genetic; Genetic Screening; Genetic study; Goals; Heat Stress Disorders; Histopathology; Human; Lead; Link; Longevity; Mammalian Cell; Maps; Mediating; Missense Mutation; Modeling; Molecular Chaperones; Motor; Motor Neurons; Mutation; Natural Immunity; Nerve Degeneration; Neurodegenerative Disorders; Neuromuscular Junction; Neurons; Neuropathogenesis; Orphan; Parkinson Disease; Pathogenesis; Pathologic; Pathway interactions; Patients; Phenotype; Photoreceptors; Pluripotent Stem Cells; Process; Production; Proline; Property; Proteins; Proteomics; Stress; System; Testing; Tissues; Toxic effect; UBQLN1 gene; Ubiquitin; Up-Regulation; axon guidance; cell type; drug development; fly; frontotemporal lobar dementia-amyotrophic lateral sclerosis; genetic analysis; induced pluripotent stem cell; insight; interest; mutant; mutational status; neuron apoptosis; neuronal survival; novel; protein aggregation; protein degradation; protein folding; protein function; proteostasis; proteotoxicity; receptor; stem cells; trafficking; transcriptomics; ubiquilin

Project summary The overarching goal of this study is to use complementary Drosophila and inducible pluripotent stem cells (iPSC)-derived neuronal models to understand how mutations in the Ubiquilin 2 (UBQLN2) gene cause amyotrophic lateral sclerosis/frontotemporal dementia (ALS/FTD). UBQLN2 and closely related UBQLN1 belong to a family of eukaryotic ubiquitin (Ub)-binding proteins that function, in part, as chaperones for proteins that are destined for proteasomal degradation. Missense mutations within a unique, functionally orphan proline-repeat region (PRR) of UBQLN2 cause X-linked, forms of ALS/FTD, with some patients exhibiting a pure dementia onset. In addition, ubiquilin histopathology, comprised of dense aggregates of UBQLN2 and UBQLN1 are observed in most instances of ALS/FTD regardless of UBQLN2 mutation status. To address pathomechanisms of UBQLN2-associated FTD we exploited the upstream activating sequence (UAS)/GAL4 system to generate isogenic Drosophila strains expressing wild-type (WT) and ALS mutant forms of UBQLN2 in different tissues and cell types. We found that UBQLN2ALS mutants elicited dose-dependent phenotypes— including eye degeneration, motor defects, and lifespan shortening—that were more severe than phenotypes caused by equivalent expression of UBQLN2WT. UBQLN2ALSmutants, but not UBQLN2WT, formed intraneuronal aggregates characteristic of ubiquilin inclusions found in ALS/FTD patients. Unbiased genetic screens identified more than 30 genetic intervals that either reduced or enhanced UBQLN2ALS mutant toxicity. Gene mapping studies suggest that endolysosomal pathways are centrally involved in UBQLN2ALS-mediated neurodegeneration and point toward axon guidance genes and neuronal dependence receptors as potentially novel disease modifiers. In this R01 grant proposal we will continue genetic studies of UBQLN2-mediated neurodegeneration in Drosophila, focusing on endolysosomal trafficking and axon guidance as lead pathways for discovery efforts. Modifier pathways identified in Drosophila will, in turn, inform studies of iPSC-derived motor neurons (iMNs) expressing endogenous UBQLN2ALS alleles. The specific objectives of Aim 1 are to: (a) investigate the Rab5 gene and endolysosomal defects in UBQLN2ALS flies; (b) investigate the contributions of the Unc-5 axon guidance pathway to UBQLN2-mediated neurodegeneration; and (c) map and validate of UBQLN2ALS modifier genes in Drosophila. The specific objectives of Aim 2 are to: (a) establish functional defects in UBQLN2ALS iMNs; (b) carry out proteomic analysis of UBQLN2ALS iMNs; and (c) evaluate genes emerging from Drosophila screens in Aim 1 as genetic modifiers of UBQLN2ALS toxicity in iMNs. The combined genetic, cellular, and biochemical studies using Drosophila and mammalian iMN models will provide important new insights into UBQLN2-associated neurodegeneration in ALS/FTD. In addition, pathways identified in our study are likely to overlap with and inform toxicity pathways instigated by other aggregation-prone, ALS/FTD- associated genes.

项目概要 这项研究的总体目标是使用互补的果蝇和诱导多能干细胞 (iPSC) 衍生的神经元模型，用于了解泛素 2 (UBQLN2) 基因突变如何导致 肌萎缩侧索硬化症/额颞叶痴呆（ALS/FTD）。 UBQLN2 和密切相关的 UBQLN1 属于真核泛素 (Ub) 结合蛋白家族，其部分功能是作为蛋白质的伴侣 注定要被蛋白酶体降解。独特的功能性孤儿基因中的错义突变 UBQLN2 的脯氨酸重复区域 (PRR) 导致 X 连锁形式的 ALS/FTD，一些患者表现出 纯痴呆发作。此外，泛素组织病理学由 UBQLN2 和 UBQLN2 的致密聚集体组成 无论 UBQLN2 突变状态如何，大多数 ALS/FTD 病例中都会观察到 UBQLN1。致地址 我们利用上游激活序列（UAS）/GAL4 来研究 UBQLN2 相关 FTD 的病理机制 系统产生表达野生型（WT）和ALS突变型UBQLN2的同基因果蝇菌株 在不同的组织和细胞类型中。我们发现 UBQLN2ALS 突变体引发剂量依赖性表型—— 包括眼睛退化、运动缺陷和寿命缩短——这些比表型更严重 由 UBQLN2WT 的等效表达引起。形成 UBQLN2ALS 突变体，但不是 UBQLN2WT ALS/FTD 患者中发现的泛素内含物的神经元内聚集体特征。公正的遗传 筛选确定了 30 多个降低或增强 UBQLN2ALS 突变体毒性的遗传区间。 基因图谱研究表明，内溶酶体途径主要参与 UBQLN2ALS 介导的过程 神经变性并指出轴突引导基因和神经元依赖性受体可能 新的疾病调节剂。在此 R01 拨款提案中，我们将继续 UBQLN2 介导的遗传学研究 果蝇的神经变性，重点关注内溶酶体运输和轴突引导作为先导途径 用于发现工作。果蝇中发现的修饰途径反过来将为 iPSC 衍生的研究提供信息 表达内源性 UBQLN2ALS 等位基因的运动神经元 (iMN)。目标 1 的具体目标是： (a) 研究 UBQLN2ALS 果蝇的 Rab5 基因和内溶酶体缺陷； (b) 调查以下方面的贡献 Unc-5 轴突引导通路至 UBQLN2 介导的神经变性； (c) 映射和验证 果蝇中的 UBQLN2ALS 修饰基因。目标 2 的具体目标是： (a) 建立职能 UBQLN2ALS iMN 中的缺陷； (b) 对 UBQLN2ALS iMN 进行蛋白质组分析； (c) 评估基因 目标 1 中果蝇筛选中出现的 UBQLN2ALS 对 iMN 毒性的遗传修饰剂。合并后的 使用果蝇和哺乳动物 iMN 模型进行遗传、细胞和生化研究将提供重要的信息。 对 ALS/FTD 中 UBQLN2 相关神经变性的新见解。此外，我们发现的途径 研究可能与其他易聚集的 ALS/FTD 引发的毒性途径重叠并提供信息 相关基因。