Mechanistic dissection of eukaryotic protein biogenesis and degradation pathways

真核蛋白质生物发生和降解途径的机制剖析

• 批准号: 10623737
• 负责人: Vladimir Denic
• 金额: $ 57.59万
• 依托单位: HARVARD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-04-01 至 2028-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10623737
• 关键词: 2019-nCoV; 3-Dimensional; Age; Amino Acids; Area; Autophagocytosis; Autophagosome; Biochemical; Biogenesis; Cell Culture Techniques; Cells; Cues; Degradation Pathway; Dissection; Encapsulated; Endoplasmic Reticulum; Genetic Screening; Genomics; Heat-Shock Response; Human; Intracellular Membranes; Link; Microscopy; Molecular Chaperones; Nerve Degeneration; Organelles; Peptide Elongation Factor 1; Pharmaceutical Preparations; Phenotype; Physiological; Process; Production; Proteins; Quality Control; Shapes; Signal Transduction; Starvation; Stress; System; Testing; Theoretical model; Ubiquitin; Yeasts; biological adaptation to stress; interest; multicatalytic endopeptidase complex; mutant; peroxisome; preclinical efficacy; protein degradation; protein structure function; proteostasis; receptor; reconstitution

Abstract My lab currently has two main areas of interest: 1) a new chaperone system (eFOLD) that enables biogenesis of eukaryotic translation elongation factor 1 alpha (eEF1A); 2) endoplasmic reticulum (ER) and peroxisome degradation by selective autophagy. Errors in eEF1A biogenesis result in rapid degradation by the ubiquitin-proteasome system (UPS) thus making protein degradation a natural link between the two areas. Both eFOLD and selective autophagy are controlled by distinct stress responses (e.g. heat shock vs. amino acid starvation) but jointly serve as effectors of protein homeostasis (proteostasis). Both areas raise similar questions regarding substrate selectivity: How does a specific eFOLD chaperone co-translationally recognize an aggregation-prone region of eEF1A nascent chains? How is terminally misfolded eEF1A recognized for degradation by the UPS? What signals on damaged or unwanted organelles are detected by specific autophagy receptors to orchestrate encapsulation of organelle targets into autophagosomes? To answer these questions, we are dissecting biogenesis and degradation mechanisms that select substrates of grossly different sizes, respond to distinct physiological cues, and have widely different temporal dynamics. Using yeast and human cell culture in parallel, we are exploring conserved aspects of eFOLD and selective autophagy mechanisms shared by each species, as well as species-specific adaptations. Broadly speaking, our projects spawn from identification of missing factors by genetic screening or biochemical purification but all seek a deep mechanistic understanding of mutant phenotypes through biochemical reconstitution with purified components and protein structure-function analysis. Along this path, we iteratively test our hypotheses by genomics, quantitative cell microscopy, and theoretical modeling approaches.

摘要 我的实验室目前有两个主要感兴趣的领域：1)一种新的伴侣系统(EFOLD)，它能够实现生物发生 真核细胞翻译延长因子1α(EEF1a)；2)内质网(ER)和过氧化物酶 通过选择性自噬进行降解。EEF1A生物发生中的错误导致快速降解 泛素-蛋白酶体系统(UPS)因此使蛋白质降解成为这两个区域之间的天然联系。 EFOLD和选择性自噬都是由不同的应激反应控制的(例如，热休克和氨基 酸饥饿)，但共同作为eff受体的蛋白质稳态(蛋白稳态)。这两个领域都提出了类似的问题 关于底物选择性的问题：一个特定的ficFOLD伴侣是如何共翻译识别的 EEF1a新生链易聚集区域？如何识别末端错误折叠的eEF1a UPS的降级？SPECIfic检测到损坏或不需要的细胞器上的哪些信号 自噬受体协调细胞器靶向自噬小体的包裹？回答 这些问题，我们正在大体上剖析选择底物的生物发生和降解机制 Diff的大小不同，对不同的生理线索有不同的反应，并具有广泛不同的时间动力学特征(ff)。vbl.使用 酵母和人类细胞培养并行进行，我们正在探索eFOLD和选择性的保守方面 每个物种共有的自噬机制以及物种间的fic适应。一般说来， 我们的项目通过基因筛选或生化纯化fi阳离子从缺失因子的同源fi阳离子衍生而来，但 所有人都通过生化重组寻求对突变表型的深入机制理解 纯化的fi组分和蛋白质结构功能分析。沿着这条路，我们反复测试我们的 通过基因组学、定量细胞显微镜和理论建模方法的假设。