Recognition and degradation of mRNA by nonsense-mediated decay

无义介导的衰变对 mRNA 的识别和降解

• 批准号: 8499372
• 负责人: Kristian Eileen Baker
• 金额: $ 28.79万
• 依托单位: CASE WESTERN RESERVE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-20 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8499372
• 关键词: Accounting; Address; Belief; Binding; Binding Proteins; Biological Assay; Cell model; Cells; Core Protein; Data; Discrimination; Disputes; Elements; Ensure; Eukaryotic Cell; Gene Expression; Gene Mutation; Genetic; Goals; Grant; Health; Hereditary Disease; Human; Human Genetics; Individual; Lead; Left; Light; Mediating; Messenger RNA; Metabolism; Modeling; Molecular; Mutation; Nonsense Codon; Nonsense-Mediated Decay; Pathway interactions; Patients; Peptide Initiation Factors; Play; Poly(A) Tail; Process; Protein Binding; Proteins; Quality Control; RNA; RNA Binding; RNA-Binding Proteins; Recruitment Activity; Relative (related person); Ribosomes; Role; Saccharomyces cerevisiae; Signal Transduction; Terminator Codon; Testing; Trans-Activators; Translating; Translations; Untranslated Regions; Yeasts; base; innovation; insight; mRNA Decay; messenger ribonucleoprotein; novel; novel strategies; novel therapeutic intervention; polypeptide; premature; prevent; protein protein interaction; response

DESCRIPTION (provided by applicant): Degradation of mRNA plays an important role in regulating gene expression. In eukaryotic cells a specialized pathway has evolved to stimulate the degradation of aberrant mRNAs containing a premature nonsense codon (PTC), a signal that causes early termination of translation and, if left unchecked, the accumulation of truncated polypeptides. This pathway, referred to as nonsense- mediated mRNA decay (NMD), represents a conserved quality control mechanism that upholds fidelity and protects cells from erroneous gene expression. The widely accepted model for NMD posits that PTC-containing mRNA are recognized as aberrant due an impaired interaction between the terminating ribosome and the mRNA poly(A) tail-bound protein, PAB1, as a consequence of translation termination occurring too far away. Our observations challenge this paradigm and indicate that a model invoking a distance between the terminating ribosome and PAB1 as the critical determinant in NMD substrate recognition cannot sufficiently explain how cells discriminate between normal and premature translation termination. The long term goal of this proposal is to elucidate the molecular mechanisms underlying both recognition and rapid degradation of mRNA by the NMD pathway. We will address the important questions of how nonsense-containing mRNA are both recognized and targeted for rapid degradation under three specific aims. First, we will identify and characterize mRNP elements important for targeting PTC-containing mRNA to NMD. In addition, we propose two innovative and unbiased approaches to identify cis- and trans-acting antagonists of NMD. Second, we will investigate the requirements for assembly of NMD proteins with mRNA substrates. In particular, we will examine the order for protein assembly, whether binding targets are ribosomes, mRNA, or both, and particular mRNA features for their importance in NMD mRNP assembly. Finally we will also assess the functional capabilities of individual NMD proteins independent of their RNA binding capacities. Third, we will elucidate how recognition of a PTC- containing mRNA as aberrant is communicated to the cellular decay machinery and leads to accelerated degradation of the mRNA. Our preliminary data dispute a common belief that the NMD machinery directly recruits the decay machinery to the mRNA through conserved protein- protein interactions. We plan to further challenge this model as well as test our hypothesis that rapid mRNA decay is an indirect consequence of restricting translation initiation factor binding to the mRNA 52 cap.

描述（由申请人提供）：mRNA的降解在调节基因表达中起重要作用。在真核细胞中，已经进化出一种专门的途径来刺激含有过早无义密码子（PTC）的异常mRNA的降解，PTC是一种导致翻译提前终止的信号，如果不加抑制，则会导致截短多肽的积累。该途径被称为无义介导的mRNA衰变（NMD），代表了保守的质量控制机制，其维持保真度并保护细胞免于错误的基因表达。广泛接受的NMD模型假定，含有PTC的mRNA被认为是异常的，这是由于终止核糖体和mRNA聚（A）尾结合蛋白PAB 1之间的相互作用受损，这是翻译终止发生得太远的结果。我们的观察结果挑战了这种范式，并表明，调用终止核糖体和PAB 1之间的距离作为NMD底物识别的关键决定因素的模型不能充分解释细胞如何区分正常和过早的翻译终止。该提案的长期目标是阐明NMD途径识别和快速降解mRNA的分子机制。我们将解决的重要问题，如何无义的mRNA都承认和有针对性的快速降解下三个具体目标。首先，我们将鉴定和表征mRNP元件，其对于将含有PTC的mRNA靶向NMD是重要的。此外，我们提出了两个创新的和公正的方法来确定顺式和反式作用的拮抗剂的NMD。第二，我们将研究NMD蛋白与mRNA底物组装的要求。特别是，我们将研究蛋白质组装的顺序，结合靶点是否是核糖体、mRNA或两者，以及特定mRNA特征在NMD mRNP组装中的重要性。最后，我们还将评估独立于其RNA结合能力的单个NMD蛋白的功能能力。第三，我们将阐明如何识别一个PTC-含有mRNA作为异常是沟通的细胞衰变机制，并导致加速降解的mRNA。我们的初步数据质疑一个共同的信念，即NMD机制直接招募衰变机制的mRNA通过保守的蛋白质-蛋白质相互作用。我们计划进一步挑战这一模型，以及测试我们的假设，即快速mRNA衰变是限制翻译起始因子结合mRNA 52帽的间接后果。