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Translation Kinetics and their Effects on Protein Structure and Function, mRNA half-lives, and Cellular Phenotype

翻译动力学及其对蛋白质结构和功能、mRNA 半衰期和细胞表型的影响

基本信息

批准号：
10552103
负责人：
Edward Patrick O'Brien
金额：
$ 58.24万
依托单位：
PENNSYLVANIA STATE UNIVERSITY, THE
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-08-01 至 2028-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10552103
关键词：
Affect Amino Acids Bacteria Big Data Binding Bioinformatics Biological Catalysis Cells Chemotaxis Circadian Rhythms Codon Nucleotides Cryoelectron Microscopy DNA Data Data Set Disease Drosophila genus Enzymes Gene Expression Genetic Transcription Growth Human Kinetics Lead Link Machine Learning Malignant Neoplasms Mass Spectrum Analysis Messenger RNA Methods Molecular Mutation NUP214 gene Outcome Pattern Phenotype Positioning Attribute Process Proteins Research Research Proposals Ribosomes Statistical Methods Techniques Testing Time Translating Translations cell growth computerized tools enzyme activity follow-up fungus in silico in vivo evaluation insight mRNA Transcript Degradation migration molecular modeling novel predictive modeling promoter protein function protein misfolding protein structure rate of change simulation theories tool transcription factor

项目摘要

Project Summary Translation kinetics critically influences protein structure and function, the rate of mRNA degradation, and cellular phenotype. When synonymous codon mutations are incorporated into an mRNA molecule (which changes the rate at which codon positions are translated by the ribosome but not the amino acids they encode) the specific activity of enzymes changes for long time periods, mRNA degradation rates are altered when these mutations lead to ribosome traffic jams, and the ability of cells to migrate and maintain a circadian rhythm can be affected. These effects occur across species, from bacteria to fruit flies, from fungi to humans. What is missing in this field is a comprehensive, molecular understanding of how translation kinetics influences these processes. Utilizing both computational (theory, simulation, and big data) and experimental (Mass Spec, Cryo- EM, and NMR) methods, this proposal focuses on four fundamental questions: (i) Can the existence of a novel form of protein misfolding, which is suggested to link synonymous mutations and altered protein function, be experimentally demonstrated? (ii) Can gene expression be altered when such protein misfolding occurs in transcription factors? (iii) Is it possible to understand and predict how elongation kinetics give rise to different patterns of ribosome traffic, and how these patterns influence translation-dependent mRNA degradation? (iv) What molecular mechanisms connect synonymous mutations in humans to changes in growth phenotype? Preliminary data suggest clear hypotheses to these questions. For questions (i) and (ii), synonymous mutations are hypothesized to alter the kinetic partitioning of nascent protein molecules into subpopulations of misfolded, soluble, self-entangled states that have reduced functionality, which can affect catalysis in the case of enzymes or DNA promotor binding in the case of transcription factors. For question (iii), application of interpretable machine learning has suggested molecular factors that can alter both ribosome traffic and mRNA degradation – which will form the basis for follow up in silico and in vivo testing. And for question (iv), rigorous statistical methods the PI has used have identified synonymous cancer drivers which provide a unique opportunity to understand and connect synonymous mutations to cellular phenotype. These hypotheses will be tested using computational tools including multi-scale simulation techniques, bioinformatics, and machine learning. And experimentally tested using mass spectrometry, Cryo-EM, NMR, and enzymatic chemotaxis. This research will establish a unifying mechanism by which synonymous mutations can alter soluble protein structure and function over long time periods. It will provide a novel molecular basis by which synonymous mutations can affect gene expression at the transcriptional level. It will result in a predictive model connecting non-linear effects between translation kinetics, ribosome traffic, and translation-dependent mRNA degradation. And finally, it will establish the existence of synonymous cancer drivers affecting human cell growth phenotype and the molecular mechanisms by which this occurs.

项目概要翻译动力学对蛋白质结构和功能、mRNA 降解率以及细胞表型。当同义密码子突变被整合到 mRNA 分子中时（改变核糖体翻译密码子位置的速率，但不改变它们编码的氨基酸）酶的比活性会长时间变化，当这些变化发生时，mRNA 降解率会发生变化突变导致核糖体交通堵塞，细胞迁移和维持昼夜节律的能力可以受到影响。这些影响发生在各个物种之间，从细菌到果蝇，从真菌到人类。什么是该领域缺少对翻译动力学如何影响这些因素的全面的分子理解。流程。利用计算（理论、模拟和大数据）和实验（质谱、冷冻） EM 和 NMR）方法，该提案重点关注四个基本问题：（i）是否可以存在一种新颖的蛋白质错误折叠的形式，被认为将同义突变和蛋白质功能改变联系起来实验证明？ (ii) 当这种蛋白质错误折叠发生在转录因子？ (iii) 是否有可能理解和预测伸长动力学如何产生不同的核糖体运输模式，以及这些模式如何影响翻译依赖性 mRNA 降解？（四）哪些分子机制将人类同义突变与生长表型的变化联系起来？初步数据对这些问题提出了明确的假设。对于问题 (i) 和 (ii)，同义假设突变会改变新生蛋白质分子动态分配为亚群错误折叠、可溶、自缠结状态的功能性降低，可能会影响催化作用在转录因子的情况下，酶或 DNA 启动子的结合。对于问题(iii)，应用可解释的机器学习表明分子因素可以改变核糖体运输和 mRNA 降解——这将构成后续计算机模拟和体内测试的基础。对于问题（iv）， PI 使用的严格统计方法已经确定了同义的癌症驱动因素，这些驱动因素提供了独特的有机会了解同义突变并将其与细胞表型联系起来。这些假设将使用计算工具进行测试，包括多尺度模拟技术、生物信息学和机器学习。并使用质谱、冷冻电镜、核磁共振和酶趋化性进行了实验测试。这项研究将建立一个统一的机制，通过该机制同义突变可以改变可溶性长期的蛋白质结构和功能。它将提供一个新的分子基础同义突变可以影响转录水平的基因表达。它将产生一个预测结果连接翻译动力学、核糖体运输和翻译依赖性之间的非线性效应的模型 mRNA 降解。最后，它将确定影响人类的同义癌症驱动因素的存在细胞生长表型及其发生的分子机制。

项目成果

期刊论文数量（21）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Identifying A- and P-site locations on ribosome-protected mRNA fragments using Integer Programming

DOI：
10.1038/s41598-019-42348-x
发表时间：
2019-04-18
期刊：
SCIENTIFIC REPORTS
影响因子：
4.6
作者：
Ahmed, Nabeel;Sormanni, Pietro;O'Brien, Edward P.
通讯作者：
O'Brien, Edward P.

Pulse labeling reveals the tail end of protein folding by proteome profiling.

脉冲标记揭示了通过蛋白质组分析的蛋白质折叠的尾端。

DOI：
10.1016/j.celrep.2022.111096
发表时间：
2022-07-19
期刊：
CELL REPORTS
影响因子：
8.8
作者：
Zhu, Mang;Kuechler, Erich R.;Wong, Ryan W. K.;Calabrese, Gaetano;Sitarik, Ian M.;Rana, Viraj;Stoynov, Nikolay;O'Brien, Edward P.;Gsponer, Jorg;Mayor, Thibault
通讯作者：
Mayor, Thibault

Mechanical Forces Have a Range of Effects on the Rate of Ribosome Catalyzed Peptidyl Transfer Depending on Direction.