Leveraging ubiquitin-dependent regulatory mechanisms to improve proteome quality in health and disease

利用泛素依赖性调节机制提高健康和疾病中的蛋白质组质量

• 批准号: 10552479
• 负责人: Eric J Bennett
• 金额: $ 32.43万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-01 至 2028-02-29
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10552479
• 关键词: Aging; Biochemical; Biogenesis; Cell Line; Cell Proliferation; Cells; Cellular Stress; Chromosome abnormality; Chronic; Complex; DNA biosynthesis; Defect; Detection; Development; Disease; Dissection; Excision; Functional disorder; Genetic Transcription; Goals; Health; Human; Human Pathology; Impairment; Length; Ligase; Link; Longevity; Messenger RNA; Molecular; Mutation; Nature; Neurodegenerative Disorders; Neurologic Dysfunctions; Outcomes Research; Pathology; Pathway interactions; Persons; Prevalence; Process; Production; Proliferating; Proteins; Proteome; Quality Control; Recycling; Research; Ribosomes; Stress; Structure; System; Toxic effect; Translation Initiation; Translations; Triage; Ubiquitin; Variant; combat; fitness; genome editing; healthspan; improved; neoplastic cell; nervous system disorder; proteostasis; proteotoxicity; response; ubiquitin ligase

PROJECT SUMMARY Errors associated with DNA replication, transcription, mRNA processing, and protein biogenesis result in the continuous production of potentially toxic defective proteins. The error-prone nature of these essential processes requires robust quality control (QC) systems to effectively triage and destroy defective translation products. Protein quality control is an essential component within the larger protein homeostasis (proteostasis) system and proteostasis dysfunction has been implicated in human aging-related pathologies. On one hand, elevated protein QC function is needed to enable neoplastic cell proliferation in cells with high mutational burdens or chromosomal abnormalities. Conversely, impaired proteostasis and defects in protein QC function result in the enhanced production of misfolded and toxic aggregation prone proteins that typify many neurodegenerative disorders. These observations suggest that developing molecular strategies to predictably alter QC function to either enhance, or limit QC capacity as needed can improve aging-associated disorders and extend human healthspan. However, there is a surprising and substantial gap in our understanding of not only how QC systems selectively engage their substrates, but also how substrates evade detection during proteostasis dysfunction. To make substantive progress toward the goal of leveraging QC systems to combat aging-associated disorders, it is necessary to identify and characterize cellular and molecular mechanisms that enable detection and degradation of diverse QC substrates. Recent research progress from my lab has identified a spatially restricted QC pathway that acts on stalled and collided ribosomal complexes both before and after translation initiation to target defective translation products for degradation and recycle ribosomal complexes. Further, we have developed a systematic pipeline for biochemical, structural, and cellular interrogation of enigmatic but critical QC ubiquitin ligases that have been implicated in targeting diverse substrates for degradation by unknown mechanisms. We have focused our initial studies on the ubiquitin ligase HUWE1. Our recently described HUWE1 structure represents the first full-length structure of a HECT-domain ligase. We have generated a unique and powerful set of genome-edited cell lines and HUWE1 variants that have and will enable molecular dissection of HUWE1 function, HUWE1 substrate identification, and identification of cellular stress conditions that require HUWE1 for cellular survival and proliferation. Research outcomes achieved by the proposed studies will mechanistically determine how terminally stalled ribosomes are sensed and resolved and how ribosome-associated QC pathways can be manipulated to alter proteostasis function. Further, we will establish mechanisms by which QC ligases engage substrates under normal and stressed conditions. Successful completion of the proposed research will provide substantial progress toward our long-term goal of combating aging-associated human pathology through the development of molecular strategies to modify cellular responses to chronic proteotoxic stress and improve cellular fitness following proteostasis insults.

项目摘要 与DNA复制，转录，mRNA加工和蛋白质生物发生相关的误差导致 连续产生潜在的有毒缺陷蛋白。这些基本过程容易出错的性质 需要健壮的质量控制（QC）系统，以有效分类并破坏缺陷的翻译产品。 蛋白质质量控​​制是较大的蛋白质稳态（蛋白抑制）系统中的重要组成部分和 蛋白毒性功能障碍与人类衰老相关的病理有关。一方面，升高的蛋白质 需要QC功能以在高突起或染色体的细胞中实现肿瘤细胞的增殖 异常。相反，蛋白质QC功能的蛋白质量和缺陷受损导致增强 代表许多神经退行性疾病的代表性蛋白质折叠式和有毒的聚集蛋白的产生。 这些观察结果表明，开发分子策略以可预测的将QC功能更改为任何一个 根据需要增强或限制QC容量可以改善与衰老相关的疾病并扩展人类健康状况。 但是，我们对QC系统的理解有一个令人惊讶的差距 参与其底物，还可以在蛋白质功能障碍期间避免检测。做 实质性进步朝着利用QC系统打击衰老相关的疾病的目标，这是 确定和表征能够检测和降解的细胞和分子机制所必需的 QC底物的不同。我实验室的最新研究进度已经确定了QC途径的空间限制 在转换启动到目标之前和之后，这都作用于失速和碰撞的核糖体配合物 降解和回收核糖体复合物的不良翻译产物。此外，我们已经开发了 神秘但关键QC泛素的生化，结构和细胞询问的系统管道 与未知机制靶向不同的底物降解的连接酶。我们 将我们的初步研究集中在泛素连接酶HUWE1上。我们最近描述的Huwe1结构 代表海域连接酶的第一个全长结构。我们已经产生了独特而强大的集合 基因组编辑的细胞系和HUWE1变体，它们具有并将使Huwe1的分子解剖 功能，HUWE1底物识别以及需要Huwe1的细胞应力条件的鉴定 细胞存活和增殖。拟议的研究实现的研究结果将机械学 确定终末失速核糖体的感测和分辨方式以及核糖体相关的QC 可以操纵途径以改变蛋白质的功能。此外，我们将建立QC的机制 连接酶在正常和压力条件下互动底物。成功完成拟议的 研究将为我们的长期目标提供重大进展，以打击与衰老相关的人类 通过发展分子策略的病理学，以改变对慢性蛋白毒性的细胞反应 压力并改善蛋白质毒化后的细胞适应性。