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

基本信息

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
项目类别：
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&apos 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

项目摘要

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.

项目摘要这项研究的总体目标是使用果蝇和诱导多能干细胞（IPSC）衍生的神经元模型，以了解泛素2（UBQLN2）基因中的突变如何引起肌萎缩性侧索硬化/额颞痴呆（ALS/FTD）。 UBQLN2和密切相关的UBQLN1 属于真核泛素（Ub）的家族，结合蛋白质，部分作用为蛋白质的伴侣注定是用于蛋白酶体降解的。独特的功能孤儿中的错义突变 UBQLN2的脯氨酸重复区域（PRR）引起X连锁的ALS/FTD形式，一些患者表现出A 纯痴呆症发作。此外，泛素组织病理学由UBQLN2和无论UBQLN2突变状态如何，在大多数ALS/FTD的情况下都可以观察到UBQLN1。解决 UBQLN2相关的FTD的病理力学我们探索了上游激活序列（UAS）/GAL4 产生表达野生型（WT）和ALS突变形式的ubqln2的果蝇菌株的系统在不同的组织和细胞类型中。我们发现UBQLN2AL突变体引起了剂量依赖的表型 - 包括眼睛变性，运动缺陷和寿命缩短 - 比表型更严重由UBQLN2WT的等效表达引起。 UBQLN2AlsMutant，但不是UBQLN2WT形成的在ALS/FTD患者中发现的泛素夹杂物的神经元聚集体特征。公正的通用筛选确定了30多个遗传间隔，可降低或增强UBQLN2ALS突变体毒性。基因映射研究表明，内侧溶血途径集中参与UBQLN2ALS介导的神经变性并指向轴突引导基因和神经元依赖受体可能是潜在的新型疾病修饰符。在此R01赠款提案中，我们将继续对UBQLN2介导的遗传研究果蝇中的神经变性，重点是内溶性贩运和轴突指导作为铅途径进行发现工作。果蝇中鉴定出的修饰符途径将依次为IPSC衍生的研究提供信息表达内源性UBQLN2al等位基因的运动神经元（IMN）。目标1的具体目标是：（a）研究Ubqln2als苍蝇中的Rab5基因和内溶性缺陷；（b）调查 UNC-5轴突引导途径通往UBQLN2介导的神经变性；（c）地图和验证果蝇中的ubqln2als修饰基因。目标2的具体目标是：（a）建立功能 UBQLN2als IMN中的缺陷；（b）对UBQLN2als IMN进行蛋白质组学分析；（c）评估基因 AIM 1中的果蝇筛选出现，作为IMN中UBQLN2als毒性的遗传修饰符。组合使用果蝇和哺乳动物IMN模型的遗传，细胞和生化研究将提供重要的对ALS/FTD中与UBQLN2相关的神经变性的新见解。此外，我们在我们的研究可能与其他容易碰到的ALS/FTD-煽动的毒性途径重叠并为毒性途径重叠相关基因。