Recognition and degradation of mRNA by nonsense-mediated decay.

通过无义介导的衰变来识别和降解 mRNA。

基本信息

批准号：
10458690
负责人：
Kristian Eileen Baker
金额：
$ 37.9万
依托单位：
CASE WESTERN RESERVE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-08-01 至 2025-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10458690
关键词：
3&apos Untranslated Regions ATP Hydrolysis ATP phosphohydrolase Attenuated Binding Binding Sites Biochemical Biochemical Genetics Biological Biological Process Biotin Cells Complement Complex Coupled Data Data Set Defect Engineering Ensure Enzymes Eukaryotic Cell Event Excision Gene Expression Genetic Diseases Genetic Translation Genomic approach Goals Hereditary Disease Human Genetics Immunoprecipitation Individual Label Left Length Mediating Messenger RNA Modeling Molecular Monitor Nature Nonsense Codon Nonsense Mutation Nonsense-Mediated Decay Open Reading Frames Pathway interactions Poly(A)+ RNA Process Production Proteins Protocols documentation Quality Control RNA RNA Binding RNA Decay RNA-Binding Proteins Reagent Recycling Regulator Genes Research Ribosomes Role Saccharomycetales Seminal Sequence Analysis Signal Transduction Testing Therapeutic Time Transcript Translating Translations Yeast Model System Yeasts cell type cofactor crosslink experimental study genetic approach genetic information in vivo in vivo monitoring innovation insight mRNA Decay mRNA Transcript Degradation mutant novel polypeptide premature prevent protein complex public health relevance tool transcriptome

项目摘要

PROJECT SUMMARY/ABSTRACT Gene expression is closely monitored to ensure fidelity in the conversion of genetic information into biological activity. In eukaryotic cells a specialized quality control checkpoint exists during mRNA translation to evaluate mRNA integrity and rapidly degrade transcripts containing a nonsense codon in their protein-coding region - a signal that causes premature termination of translation and, if left unchecked, the accumulation of truncated polypeptides. This pathway, referred to as nonsense-mediated mRNA decay (NMD), serves a vital biological function by protecting cells from the deleterious consequences of incomplete proteins. In spite of over two decades of study, fundamental questions central to this process remain poorly resolved, including how cells distinguish normal and premature translation termination and what molecular events occur subsequently between the translating mRNA and NMD machinery to incite accelerated degradation of the nonsense codon- containing transcript. The long term objective of my research is to gain a comprehensive molecular understanding of the events underlying the cell's ability to recognize and target nonsense-containing mRNA for rapid decay. In the last decade, we have made seminal discoveries in characterizing RNA features vital for the recognition and efficient targeting of an mRNA to NMD. Recently, we have begun to dissect molecular events subsequent to substrate recognition through identifying the complement of proteins specifically associated with nonsense- containing mRNA and characterizing a catalytically-inactive mutant of the key NMD factor, UFP1. Notably, we have described a functional interaction between the NMD machinery and prematurely terminating ribosomes that exists in vivo and which is critical for dictating the fate of the aberrant transcript. We propose here to capitalize on our past efforts and exploit novel tools and reagents to i) identify UPF1 RNA binding targets globally to provide unprecedented insight into how NMD factor binding dictates the targeting of mRNA to NMD; ii) interrogate further the functional interaction between prematurely terminating ribosomes and the NMD machinery essential for targeting substrates to rapid decay; and iii) monitor NMD mRNP dynamics in vivo using enzyme-catalyzed proximity labeling. Together, these studies will provide unprecedented insight into the molecular events underlying an integral cellular RNA quality control pathway and regulator of gene expression.

项目概要/摘要基因表达受到密切监测，以确保遗传信息转化为生物信息的保真度活动。在真核细胞中，mRNA 翻译过程中存在专门的质量控制检查点来评估 mRNA 完整性并快速降解蛋白质编码区含有无义密码子的转录物 - 导致翻译过早终止的信号，如果不加以控制，会导致截断的累积多肽。该途径被称为无义介导的 mRNA 衰减 (NMD)，具有重要的生物学功能。通过保护细胞免受不完整蛋白质的有害后果而发挥作用。尽管超过两个经过数十年的研究，这一过程的核心基本问题仍未得到解决，包括细胞如何区分正常和过早翻译终止以及随后发生的分子事件翻译 mRNA 和 NMD 机器之间的相互作用，以加速无义密码子的降解包含转录本。我研究的长期目标是获得全面的分子了解细胞识别和靶向含有无义的 mRNA 的能力背后的事件快速腐烂。在过去的十年中，我们在表征对识别至关重要的 RNA 特征方面取得了开创性的发现以及将 mRNA 有效靶向 NMD。最近，我们开始剖析后续的分子事件通过识别与无义特异性相关的蛋白质的补体来进行底物识别含有 mRNA 并表征关键 NMD 因子 UFP1 的催化失活突变体。值得注意的是，我们描述了 NMD 机器和过早终止核糖体之间的功能相互作用它存在于体内，对于决定异常转录本的命运至关重要。我们在此建议利用我们过去的努力并开发新的工具和试剂来 i) 识别 UPF1 RNA 结合靶标在全球范围内提供关于 NMD 因子结合如何决定 mRNA 靶向 NMD 的前所未有的见解； ii) 进一步探究过早终止核糖体和 NMD 之间的功能相互作用使基质快速腐烂所必需的机械； iii) 使用以下方法监测体内 NMD mRNP 动态酶催化的邻近标记。总之，这些研究将为我们提供前所未有的见解。完整细胞RNA质量控制途径和基因表达调节器背后的分子事件。