FUS Protein Homeostasis in ALS

ALS 中的 FUS 蛋白稳态

• 批准号: 10620292
• 负责人: Haining Zhu
• 金额: --
• 依托单位: SOUTHERN ARIZONA VA HEALTH CARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-10-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10620292
• 关键词: ALS patients; Address; Affect; Agonist; Amyotrophic Lateral Sclerosis; Animal Model; Attenuated; Bioinformatics; Categories; Cell Line; Cell Nucleus; Cell physiology; Cells; Cessation of life; Contracts; Cytoplasm; Cytoplasmic Granules; DNA Binding; Defect; Diagnosis; Disease; Drosophila genus; Endoplasmic Reticulum; Equilibrium; Foundations; Functional disorder; Future; Genes; Genetic; Genetic Models; Health; Healthcare; Hydrogels; Hyperactivity; Impairment; In Vitro; Inclusion Bodies; Intervention; Knock-out; Knowledge; Liquid substance; Mediating; Membrane Proteins; Messenger RNA; Modeling; Molecular; Motor Neurons; Mutation; N-terminal; Nerve Degeneration; Neurodegenerative Disorders; Neuronal Dysfunction; Neurons; Nonsense-Mediated Decay; Pathologic; Pathology; Pathway interactions; Phase; Phenotype; Physiological; Play; Post-Translational Protein Processing; Property; Protein Biosynthesis; Proteins; Proteome; Proteomics; Publishing; RNA; RNA Binding; RNA Decay; RNA-Binding Proteins; Rat Transgene; Reporting; Ribonucleoproteins; Role; Services; Specificity; Testing; Toxic effect; Transgenic Mice; Translations; Veterans; amyotrophic lateral sclerosis therapy; attenuation; care costs; copper zinc superoxide dismutase; cost; design; disability; effective therapy; familial amyotrophic lateral sclerosis; fused in sarcoma; high risk; improved; in vivo; in vivo Model; induced pluripotent stem cell; inhibitor; innovation; insight; mRNA Decay; mRNA Surveillance; military service; military veteran; mutant; neuron loss; neurotoxicity; novel; overexpression; pharmacologic; prion-like; protein TDP-43; proteostasis; self assembly; sporadic amyotrophic lateral sclerosis; tau Proteins; therapeutic development; transcriptome; transcriptomics; translational impact; unpublished works

Amyotrophic lateral sclerosis (ALS) is a poorly understood neurodegenerative disease with no effective treatment. ALS is connected with military service and veterans are at a higher risk of developing this debilitating disease. Mutations in several genes have been discovered to cause a subset of familial ALS, including copper- zinc superoxide dismutase (SOD1), Fused in Sarcoma (FUS), and TAR DNA-binding protein 43 (TDP-43). Studying familial ALS using the well-defined genetic models will provide critical insights into the disease pathology as well as novel targets for future therapeutic development. This project is focused on the RNA binding protein FUS that has been implicated in both familial and sporadic ALS. ALS-related mutations in FUS cause a liquid-liquid phase separation (LLPS) of the FUS protein, forming liquid droplets in vitro. In neurons, mutant FUS accumulates in the cytoplasm, forming ribonucleoprotein granules and inclusions, eventually leading to neurotoxicity. We recently identified cellular proteins in FUS- positive inclusions and a bioinformatics analysis revealed two previously unknown pathways affected by mutant FUS: protein translation and mRNA surveillance. We also demonstrated that ALS FUS mutations indeed suppressed protein translation and hyper-activated the nonsense-mediated decay (NMD) of mRNAs. Our overarching hypothesis is that the dysregulation of protein translation and mRNA nonsense-mediated decay contributes to FUS toxicity and motor neuron dysfunction. Since little is known about how the mutant FUS- induced dysregulation of protein translation and NMD cause toxicity in neurons, discoveries in this project will fill this knowledge gap and advance the field significantly. Three specific aims are designed to test the hypothesis. Aim 1 is to determine what properties of mutant FUS drive the dysregulation of protein translation and NMD. We will use different domain truncations of FUS to manipulate LLPS and examine the relationship between LLPS and mutant FUS dysregulation. In addition, we will manipulate RNA binding of FUS and test whether RNA binding is a significant contributor to mutant FUS dysfunction. We will also use FUS knockout models to examine whether FUS plays a role in in protein translation and NMD under physiological conditions. Aim 2 is to determine whether mutant FUS impairs translation of specific proteins and NMD of specific mRNAs. We will employ specialized proteomic and transcriptomic approaches to identify changes of nascent protein biosynthesis and mRNA turnover rate as a consequence of mutant FUS. Differentially altered proteins and mRNAs will be validated in animal models as well as iPSC and induced motor neurons derived from familial ALS patients. Results from these -omics approaches will be integrated to determine whether mutant FUS impairs specific molecular and cellular function and pathways. Aim 3 is to examine whether attenuation of NMD hyperactivity can restore protein translation and mitigate FUS neurotoxicity. We will use genetic and pharmacological approaches to attenuate NMD hyperactivity and determine whether the intervention can restore the balance and mitigate FUS toxicity in vitro and in vivo models. The proposed studies are highly innovative, both conceptually and technically. The proposal is based on a number of novel observations regarding the impact of ALS mutant FUS on protein translation and mRNA decay. The results from this project are expected to produce new mechanistic insights into FUS ALS, laying a foundation for future therapeutic development of new ALS treatment(s).

肌萎缩性侧索硬化症（ALS）是一种鲜为人知的神经退行性疾病，没有有效 治疗。 ALS与​​兵役有联系，退伍军人面临着更高的风险 疾病。已经发现几种基因的突变引起了一部分家族性ALS，包括铜 锌超氧化物歧化酶（SOD1），在肉瘤（FUS）和焦油DNA结合蛋白43（TDP-43）中融合。 使用明确定义的遗传模型研究家族ALS将为疾病提供关键的见解 病理学以及未来治疗发展的新目标。 该项目的重点是与家族性和家族性和 零星的ALS。 FUS中与ALS相关的突变导致FUS蛋白的液态液相分离（LLP）， 体外形成液滴。在神经元中，突变型FU在细胞质中积聚，形成核糖核蛋白 颗粒和夹杂物，最终导致神经毒性。我们最近确定了FUS-中的细胞蛋白 阳性夹杂物和生物信息学分析揭示了两个先前未知的途径受突变体影响 FUS：蛋白质翻译和mRNA监测。我们还证明了ALS FUS突变确实 抑制蛋白质翻译和过度激活mRNA的胡说八道介导的衰减（NMD）。我们的 总体假设是蛋白质翻译和mRNA废话介导的衰减的失调 有助于FUS毒性和运动神经元功能障碍。由于对突变液的知识知之甚少。 诱导的蛋白质翻译和NMD的失调引起的神经元引起的毒性，该项目的发现将填充 这些知识差距并大大提高了领域。 设计三个特定目标旨在检验假设。目标1是确定突变体的特性 FUS驱动蛋白质翻译和NMD的失调。我们将使用FUS的不同域截断 操纵LLP并检查LLP与突变FUS失调之间的关系。此外，我们将 操纵FUS的RNA结合并测试RNA结合是否是突变FUS的重要原因 功能障碍。我们还将使用FUS敲除模型检查FUS是否在蛋白质翻译中起作用 和NMD在生理条件下。 AIM 2是确定突变FUS是否会损害的翻译 特定mRNA的特定蛋白质和NMD。我们将采用专门的蛋白质组学和转录组学 确定新生蛋白生物合成和mRNA转换率的变化的方法 突变的fus。差异改变的蛋白质和mRNA将在动物模型以及IPSC中得到验证，并且 来自家族性ALS患者的诱导运动神经元。这些 - 组方法的结果将是 集成以确定突变型FUS是否会损害特定的分子和细胞功能和途径。目的 3是检查NMD多动症的衰减是否可以恢复蛋白质翻译并减轻FUS 神经毒性。我们将使用遗传和药理方法来减弱NMD的多动症和 确定干预是否可以恢复平衡并减轻体外和体内模型的FUS毒性。 拟议的研究在概念和技术上都是高度创新的。该提议基于 关于ALS突变FUS对蛋白质翻译和mRNA的影响的许多新颖的观察结果 衰变。该项目的结果有望产生对FUS ALS的新机械见解，并铺设 新ALS治疗的未来治疗发展基金会。

项目成果

ALS mutant SOD1 interacts with G3BP1 and affects stress granule dynamics.

• DOI: 10.1007/s00401-016-1601-x
• 发表时间: 2016-10
• 期刊: Acta neuropathologica
• 影响因子: 12.7
• 作者: Gal J;Kuang L;Barnett KR;Zhu BZ;Shissler SC;Korotkov KV;Hayward LJ;Kasarskis EJ;Zhu H
• 通讯作者: Zhu H