Residue-by-residue details of FUS protein phase separation and aggregation

FUS 蛋白相分离和聚集的逐个残基详细信息

• 批准号: 10503674
• 负责人: Nicolas Lux Fawzi
• 金额: $ 41.42万
• 依托单位: BROWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-23 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10503674
• 关键词: Amino Acids; Amyotrophic Lateral Sclerosis; Aromatic Amino Acids; Automobile Driving; Binding; Biological; Biological Assay; Biological Models; Biological Process; Biophysics; C-terminal; Cations; Cell Communication; Cell model; Cell physiology; Cells; Cellular Stress; Characteristics; Charge; Chimeric Proteins; Chromosomal translocation; Complement; Complex; Computer Simulation; Cytoplasmic Granules; DNA Binding Domain; DNA Damage; DNA Polymerase II; DNA Repair; Data; Disease; Explosion; Family; Foundations; Frequencies; Functional disorder; Future; Gene Expression; Genetic Transcription; Goals; Human; In Vitro; Knowledge; Lead; Link; Liquid substance; Malignant Neoplasms; Mediating; Modeling; Molecular; Molecular Probes; Mutation; NMR Spectroscopy; Neurodegenerative Disorders; Neurons; Nuclear Localization Signal; Oncogenic; Organelles; Pathologic; Pattern; Pharmacology; Phase; Physiological; Physiology; Play; Post-Translational Protein Processing; Process; Proteins; RNA; RNA Folding; RNA Polymerase II; RNA Processing; RNA Recognition Motif; RNA Splicing; RNA metabolism; RNA-Binding Proteins; Resolution; Role; Structure; TAF15 gene; Techniques; Testing; Transcription Coactivator; Transcriptional Activation; Work; base; beta pleated sheet; cell behavior; effective therapy; familial amyotrophic lateral sclerosis; frontotemporal lobar dementia-amyotrophic lateral sclerosis; fused in sarcoma; in vivo; insight; leukemia; multimodality; novel therapeutics; paralogous gene; prevent; protein structure; sarcoma; simulation; structural biology; transcription factor

Project Summary RNA-binding proteins are essential components of numerous large complexes that carry out fundamental processes including transcription, splicing, and DNA repair. Many RNA-binding proteins possess regions predicted to be disordered based on low-complexity sequence characteristics that are critical to normal RNA- processing functions, but also drive aberrant protein assembly in various neurodegenerative disease and cancers. The molecular interactions and functional roles of these disordered domains remain incompletely characterized, especially in the context of disease. Fused in Sarcoma (FUS) is one of twenty-nine human RNA-binding proteins that contains both an essential disordered low-complexity domain (LC) with unusually low charged residue composition and a high frequency of aromatic amino acids as well as several RGG motif regions. Despite disorder, these domains are thought to facilitate interactions in normal RNA metabolism by forming dynamic associations, thereby enabling tunable, reversible spatial clustering. Yet, excessive self- association between FUS disordered domains is believed to result in the formation of pathological neuronal inclusions in sub-types of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), irreversible neurodegenerative diseases that lack effective treatments. Moreover, fusion of the FUS disordered domains to several DNA-binding domains through chromosomal translocations results in uncontrolled gene expression leading to a family of aggressive cancers. Though FUS has emerged as the primary model system for understanding biological phase separation, the contacts holding together FUS assemblies and their structures in physiology and disease are currently unknown because they are invisible to traditional techniques in structural biology. However, we have demonstrated that we can visualize dynamic assemblies of FUS with residue-level resolution. This project will apply advanced nuclear magnetic resonance spectroscopy, molecular simulation, and cell models of FUS function to 1) visualize the molecular contacts that mediate LLPS and in- cell interactions, 2) probe the molecular and cellular impact of LC and RGG mutations of FUS found in familial ALS, and 3) determine the functionally relevant atomic details of FUS complexes with RNA and the C-terminal domain (CTD) of RNA polymerase II associated with RNA processing and transcription. These studies of FUS assembly will provide necessary structure/function information on future pharmacological targets for inhibiting pathological protein associations in types of ALS, FTD, leukemia, and sarcoma. Furthermore, because FUS is only one of many essential RNA-binding proteins containing aggregation-prone low complexity domains, the results of the project will serve as a foundation for understanding an entire class of proteins and for correcting their dysfunctions in disease.

项目概要 RNA 结合蛋白是许多大型复合物的重要组成部分，这些复合物执行基本功能 过程包括转录、剪接和 DNA 修复。许多 RNA 结合蛋白具有区域 根据对正常 RNA 至关重要的低复杂性序列特征预测为无序 处理功能，而且还驱动各种神经退行性疾病和疾病中的异常蛋白质组装 癌症。这些无序结构域的分子相互作用和功能作用仍然不完全 特征，特别是在疾病背景下。融合肉瘤 (FUS) 是二十九种人类肉瘤之一 RNA 结合蛋白，包含必需的无序低复杂性结构域 (LC) 和异常的结构域 低电荷残基组成和高频率的芳香族氨基酸以及几个 RGG 基序 地区。尽管无序，但这些结构域被认为通过以下方式促进正常 RNA 代谢中的相互作用： 形成动态关联，从而实现可调节、可逆的空间聚类。然而过度的自我 FUS 紊乱域之间的关联被认为导致病理性神经元的形成 肌萎缩侧索硬化症 (ALS) 和额颞叶痴呆 (FTD) 亚型中的包涵体，不可逆 缺乏有效治疗方法的神经退行性疾病。此外，FUS 无序结构域的融合 通过染色体易位导致多个 DNA 结合域导致基因表达失控 导致家族患上侵袭性癌症。尽管 FUS 已成为主要模型系统 了解生物相分离、将 FUS 组件结合在一起的接触及其结构 生理学和疾病中的一些因素目前尚不清楚，因为它们对于传统技术来说是不可见的。 结构生物学。然而，我们已经证明我们可以用以下方法可视化 FUS 的动态组装 残留水平分辨率。该项目将应用先进的核磁共振波谱、分子 FUS 功能的模拟和细胞模型 1）可视化介导 LLPS 和 in- 的分子接触 细胞相互作用，2) 探究家族中发现的 FUS LC 和 RGG 突变的分子和细胞影响 ALS，以及 3) 确定 FUS 与 RNA 和 C 末端复合物的功能相关原子细节 RNA 聚合酶 II 的结构域 (CTD) 与 RNA 加工和转录相关。 FUS的这些研究 组装将为未来抑制药物靶标提供必要的结构/功能信息 ALS、FTD、白血病和肉瘤类型中的病理蛋白关联。此外，由于 FUS 是 许多必需的 RNA 结合蛋白中只有一种含有易于聚集的低复杂性结构域， 该项目的结果将作为理解整个蛋白质类别和纠正蛋白质的基础 他们在疾病中的功能障碍。