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: 10558635
• 负责人: Tanja Mittag
• 金额: $ 62.4万
• 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/10558635
• 关键词: Acceleration; Adhesives; Affinity; Aging; Amyotrophic Lateral Sclerosis; Aromatic Amino Acids; Automobile Driving; Behavior; Biochemical; Biophysics; Brain; Cytoplasm; Degenerative Disorder; Development; Disease; Disease Progression; Elements; Genetic; Goals; Heterogeneous-Nuclear Ribonucleoproteins; Hydrogen Bonding; Investigation; Life; Link; Liquid substance; Locales; Location; Mediating; Membrane; Microscopic; Modeling; Molecular; Mutation; Nature; Neurodegenerative Disorders; Nodal; Organelles; Pathogenesis; Pathologic; Pattern; Phase; Phase Transition; Physical condensation; Physics; Polyribosomes; Positioning Attribute; Process; Proteins; RNA; RNA Recognition Motif; RNA-Binding Proteins; Role; Side; Solid; Solvents; Spinal Cord; Stress; Structure; System; TAF15 gene; Temperature; Testing; Therapeutic; Therapeutic Intervention; Translations; Vertebral column; Work; arm; crosslink; design; disease-causing mutation; driving force; effective therapy; experimental study; insight; interest; macromolecule; member; motor neuron degeneration; mutant; predictive modeling; protein TDP-43; protein aggregation; scaffold; simulation; solid state; stress granule; structural determinants; 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 肌萎缩性外侧硬化症（ALS）是一种威胁生命的神经退行性疾病，导致 大脑和脊髓中运动神经元的变性。目前既不能治愈也没有有效 治疗以减慢进展。然而，最近的新遗传，生化和生物物理证据暗示 应力颗粒作为疾病发展的坩埚。压力颗粒是无膜细胞器，也称为 生物分子冷凝物通过RNA结合蛋白和 RNA。 RNA结合蛋白中的突变将液样应激颗粒转化为固定夹杂物。长时间的压力 颗粒组件可能会产生类似的影响。这些观察指出了治疗的新机会 干预措施是否可以回答有关液体与固体组件的性质的关键问题。我们将 因此测试了基于上述观测值的总体假设，即RNA结合中的突变 蛋白质改变了相位分离的驱动力，液体冷凝物的动态停滞和 冷凝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 足以介导相分离的域（LCD），是疾病的典型位置 突变。我们将使用HNRNPA1的LCD作为ALS相关RNA类别的原型成员 结合蛋白质，并将我们的研究也扩展到FUS的LCD。 Mittag和Pappu最近开发了 基于识别瞬态，凝聚力相互作用的贴纸和间距模型 芳族氨基酸残基为相分离提供了主要的驱动力。芳香残留物 是该型号的贴纸，垫片是连接贴纸的残基。该模型启用 完全共存曲线的定量预测是温度的函数，重要的是，导致了 我们对LCD中如何编码相位分离的理解的概念发展。免费 Mittag和Pappu的专业知识现在将带来生物物理实验，计算和 以下三个特定目的的理论：（1）通过量化贴纸和间隔者模型来扩展贴纸和间隔模型 不同类型的贴纸和垫片之间的相互作用。 （2）检验导致突变的疾病的假设 在引起ALS的LCD中，引起RNA结合蛋白会导致动态停滞的相变。 （3）发现 Sidechain和骨干互动之间的相互作用及其对LCD的空间组织的贡献 在密集阶段。我们的结果将对ALS相关突变体的影响进行定量预测 关于阶段行为。我们将清楚了解序列特异性相图如何贡献 到相位分离和老化现象的动力学。我们将确定互动的类型 液体状和固体状的致密相。这些结果将直接与治疗干预有关 反对可能由动态停止的相分离引起的功能中断。