Visualizing the Mechanisms of Protein Quality Control

蛋白质质量控​​制机制的可视化

• 批准号: 9980958
• 负责人: Peter Shen
• 金额: $ 38.13万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9980958
• 关键词: ATP Hydrolysis; ATP phosphohydrolase; Address; Autophagocytosis; Binding; Cell Cycle Progression; Cell physiology; Cells; Clinical; Complex; Cryoelectron Microscopy; Degenerative Disorder; Enzymes; Equilibrium; Familial Amyotrophic Lateral Sclerosis; Functional disorder; Gene Expression; Goals; Human; Image; Inclusion Body Myopathy with Early-Onset Paget Disease; Life; Malignant Neoplasms; Membrane Fusion; Methods; Molecular; Molecular Conformation; Molecular Machines; Mutation; Pathway interactions; Protein Biosynthesis; Proteins; Proteomics; Quality Control; Research; Role; Structure; Therapeutic; Work; cancer therapy; cofactor; inhibitor/antagonist; insight; programs; tool; trafficking; tumor; valosin-containing protein

Project Summary / Abstract Optimal cellular function requires balanced networks that maintain protein synthesis, folding, trafficking, remodeling and degradation. The Cdc48/p97/VCP AAA ATPase is an essential and abundant molecular machine that helps maintain this balance across eukaryotic life and is a critical control point for many cellular pathways. Cdc48 is best characterized for its role in targeting ubiquitylated proteins for proteasomal degradation, but the enzyme also functions in a wide range of other essential pathways, including cell cycle progression, autophagy, membrane fusion, and gene expression. Mutations in human Cdc48 cause a multisystem proteopathy that clinically manifests as a combination of Inclusion Body Myopathy, Paget's Disease of Bone, Frontotemporal Dementia (collectively known as IBMPFD), and familial Amyotrophic Lateral Sclerosis (fALS). Moreover, Cdc48 expression is elevated in several tumors and its inhibitors are an emerging class of therapeutics for cancer treatment. Despite these critical roles, surprisingly little is known about the molecular mechanisms that allow Cdc48 to perform its myriad cellular functions. More than thirty adaptors and cofactors are known to interact with Cdc48, but how cells organize these binding partners into functional complexes is not well understood. My research program aims to define the mechanisms underlying Cdc48 functions by using an integrative approach of endogenous purification, proteomics, cryo-EM imaging, and computational processing methods to visualize and characterize Cdc48 assemblies in an array of its native compositional and conformational states. During the project period, we propose to achieve two major goals: first, we aim to determine the structures of native Cdc48 assemblies and their molecular determinants that functionally separate the enzyme across multiple cellular pathways; second, we seek to resolve how Cdc48 converts energy from ATP hydrolysis into a pulling force that remodels and unfolds its protein substrates. Addressing these questions is essential to understand how Cdc48 drives a wide range of cellular processes and provide insights into how its dysfunction and misregulation contribute to degenerative disease and cancer. The tools we develop to accomplish these goals will likely be broadly applicable in defining the structural landscapes of other challenging molecular machines.

项目概要/摘要 最佳的细胞功能需要平衡的网络来维持蛋白质的合成、折叠、运输、 重塑和退化。 Cdc48/p97/VCP AAA ATPase 是一种重要且丰富的分子 机器有助于维持真核生命的这种平衡，并且是许多细胞的关键控制点 途径。 Cdc48 的最佳特征在于其在蛋白酶体靶向泛素化蛋白中的作用 降解，但该酶还在许多其他重要途径中发挥作用，包括细胞周期 进展、自噬、膜融合和基因表达。人类 Cdc48 突变导致 多系统蛋白质病，临床表现为包涵体肌病、佩吉特病的组合 骨病、额颞叶痴呆（统称为 IBMPFD）和家族性肌萎缩侧索硬化症 硬化症（fALS）。此外，Cdc48 表达在多种肿瘤中升高，其抑制剂是一种新兴的 癌症治疗的一类疗法。尽管发挥了这些关键作用，但令人惊讶的是，人们对它知之甚少。 允许 Cdc48 执行其多种细胞功能的分子机制。三十多个适配器和 已知辅助因子与 Cdc48 相互作用，但细胞如何将这些结合伙伴组织成功能性的 复合体还没有被很好地理解。我的研究项目旨在定义 Cdc48 的潜在机制 通过使用内源纯化、蛋白质组学、冷冻电镜成像和 计算处理方法，以可视化和表征 Cdc48 组件在其原生数组中的特性 组成和构象状态。在项目期间，我们建议实现两大目标： 首先，我们的目标是确定天然 Cdc48 组装体的结构及其分子决定因素 在多个细胞途径中功能性地分离酶；其次，我们寻求解决Cdc48如何 将 ATP 水解产生的能量转化为重塑和展开其蛋白质底物的拉力。 解决这些问题对于了解 Cdc48 如何驱动广泛的细胞过程至关重要 并深入了解其功能障碍和失调如何导致退行性疾病和癌症。 我们为实现这些目标而开发的工具可能会广泛适用于定义结构 其他具有挑战性的分子机器的景观。