Understanding the sequence and structural determinants of phase behavior of ALS-causing proteins

了解 ALS 致病蛋白相行为的序列和结构决定因素

• 批准号: 10182841
• 负责人: Tanja Mittag
• 金额: $ 64.38万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10182841
• 关键词: Adhesives; Affinity; Aging; Amyotrophic Lateral Sclerosis; Aromatic Amino Acids; Automobile Driving; Behavior; Biochemical; Biophysics; Brain; Degenerative Disorder; Development; Disease; Disease Progression; Elements; Genetic; Goals; Heterogeneous-Nuclear Ribonucleoproteins; Hydrogen Bonding; Investigation; Lead; Life; Link; Liquid substance; Locales; Location; Mediating; Microscopic; Modeling; Molecular; Mutation; Nature; Neurodegenerative Disorders; Organelles; Pathogenesis; Pathologic; Pattern; Phase; Phase Transition; Physics; Polyribosomes; Positioning Attribute; Process; Proteins; RNA; RNA Recognition Motif; RNA-Binding Proteins; Role; Side; Solid; Solvents; Space Models; Spinal Cord; Stress; Structure; System; Temperature; Testing; Therapeutic; Therapeutic Intervention; Translations; Ursidae Family; Vertebral column; Work; arm; base; cohesion; crosslink; design; disease-causing mutation; driving force; effective therapy; experimental study; insight; interest; macromolecule; member; motor neuron degeneration; mutant; organizational structure; predictive modeling; protein TDP-43; protein aggregation; scaffold; simulation; solid state; stress granule; theories

Summary Amyotrophic lateral sclerosis (ALS) is a life-threatening, neurodegenerative disease that causes the degeneration of motor neurons in the brain and spinal cord. There are currently neither a cure nor effective treatments to slow progression. However, recent new genetic, biochemical and biophysical evidence implicates stress granules as crucibles for disease development. Stress granules are membraneless organelles, also called biomolecular condensates, which form via liquid-liquid phase separation (LLPS) of RNA-binding proteins and RNA. Mutations in RNA-binding proteins convert liquid-like stress granules into solid inclusions. Prolonged stress granule assembly can result in similar effects. These observations point to new opportunities for therapeutic interventions if key open questions regarding the nature of liquid vs. solid assemblies can be answered. We will thus test the overarching hypothesis, which is based on above observations, that mutations in RNA-binding proteins change the driving forces for phase separation, the dynamical arrest of the liquid condensates and the ability of the condensates to promote the formation of protein fibrils. Our proposed studies will thus focus on the physics of phase separation of RNA-binding proteins, specifically on their intrinsically disordered low-complexity domains (LCDs) that are sufficient for mediating phase separation and are the typical locations of disease mutations. We will use the LCD of hnRNPA1 as an archetypal member of the class of ALS-associated RNA- binding proteins and will extend our studies also to the LCD of FUS. Mittag and Pappu have recently developed a stickers-and-spacers model that is based on the identification of transient, cohesive interactions amongst aromatic amino acid residues as providing the main driving force for phase separation. The aromatic residues are the stickers in this model, the spacers are the residues that connect the stickers. The model enables the quantitative prediction of full coexistence curves as a function of temperature and, importantly, resulted in a conceptual advancement of our understanding of how phase separation is encoded in LCDs. The complimentary expertise of Mittag and Pappu will now bring to bear a combination of biophysical experiments, computation and theory on the following three specific aims: (1) To extend the stickers-and-spacers model by quantifying the interplay among different types of stickers and spacers. (2) To test the hypothesis that disease causing mutations within LCDs of ALS-causing RNA-binding proteins cause dynamically arrested phase transitions. (3) To uncover the interplay among sidechain and backbone interactions and their contributions to spatial organization of LCDs within dense phases. Our results will enable quantitative predictions of the effects of ALS-associated mutants on phase behavior. We will obtain a clear understanding of how sequence-specific phase diagrams contribute to the dynamics of phase separation and aging phenomena. We will identify the types of interactions underlying liquid-like and solid-like dense phases. These results will have a direct bearing on therapeutic interventions against the functional disruptions that are likely to be caused by dynamically arrested phase separation.

概括 肌萎缩侧索硬化症 (ALS) 是一种危及生命的神经退行性疾病，会导致 大脑和脊髓中运动神经元的退化。目前尚无治愈方法，也没有有效的方法 减缓进展的治疗。然而，最近新的遗传、生化和生物物理证据表明 应激颗粒作为疾病发展的坩埚。应激颗粒是无膜细胞器，也称为 生物分子缩合物，通过 RNA 结合蛋白的液-液相分离 (LLPS) 形成 核糖核酸。 RNA 结合蛋白的突变将液体状应力颗粒转化为固体内含物。长期压力 颗粒组装可以产生类似的效果。这些观察结果为治疗提供了新的机会 如果可以回答有关液体与固体组件性质的关键开放问题，则进行干预。我们将 因此检验基于上述观察的总体假设，即 RNA 结合突变 蛋白质改变相分离的驱动力、液体冷凝物的动态停止以及 缩合物促进蛋白质原纤维形成的能力。因此，我们提出的研究将集中于 RNA 结合蛋白的相分离物理学，特别是其本质上无序的低复杂性 足以介导相分离并且是疾病的典型位置的域（LCD） 突变。我们将使用 hnRNPA1 的 LCD 作为 ALS 相关 RNA 类的原型成员 结合蛋白，并将我们的研究扩展到 FUS 的 LCD。 Mittag 和 Pappu 最近开发了 贴纸和垫片模型基于识别之间的瞬时、内聚相互作用 芳香族氨基酸残基为相分离提供主要驱动力。芳香族残基 是该模型中的贴纸，垫片是连接贴纸的残留物。该模型使得 定量预测完全共存曲线作为温度的函数，重要的是，结果是 我们对 LCD 中相分离编码方式的理解在概念上取得了进步。免费的 米塔格和帕普的专业知识现在将结合生物物理实验、计算和 关于以下三个具体目标的理论：（1）通过量化贴纸和垫片模型来扩展贴纸和垫片模型 不同类型的贴纸和垫片之间的相互作用。 (2) 检验疾病引起突变的假设 LCD 中引起 ALS 的 RNA 结合蛋白会导致动态停滞的相变。 (3) 揭开 侧链和主干相互作用之间的相互作用及其对 LCD 空间组织的贡献 在致密相内。我们的结果将能够定量预测 ALS 相关突变体的影响 关于相行为。我们将清楚地了解特定序列的相图如何发挥作用 相分离和老化现象的动力学。我们将确定潜在的交互类型 液体状和固体状的致密相。这些结果将直接影响治疗干预 防止动态阻止相分离可能引起的功能破坏。