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Mechanisms of Translational Surveillance in C. elegans

线虫的转化监测机制

基本信息

批准号：
9932597
负责人：
Joshua Arribere
金额：
$ 10.92万
依托单位：
UNIVERSITY OF CALIFORNIA SANTA CRUZ
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-18 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9932597
关键词：
Address Affect Animals Area Binding Biochemical Biochemistry Biological Assay Caenorhabditis elegans Cells Cleaved cell Complex Data Dependence Event Excision Freezing Functional disorder Gene Abnormality Gene Expression Gene Expression Regulation Genes Genetic Genetic Diseases Genetic Variation Genomics Goals Hereditary Disease High-Throughput Nucleotide Sequencing Human Human Genetics Inherited Investigation Knock-out Knowledge Mediating Messenger RNA Modeling Molecular Monitor Mus Mutate Mutation Nature Non-Stop Decay Nonsense Codon Nonsense-Mediated Decay Pathway interactions Peptides Postdoctoral Fellow Process Production Property Proteins Quality Control RNA RNA Helicase RNA Interference Reporter Repression Ribosomal Interaction Ribosomal RNA Ribosomes Role SKI gene Site Structure Terminator Codon Testing Translations Work assault cofactor design disease-causing mutation exosome field study fly genetic approach human disease in vivo insight mRNA Decay mRNA Expression mRNA Transcript Degradation mutant novel nuclease premature prevent recruit ribosome releasing factor spatiotemporal

项目摘要

Project Summary/Abstract Cells have evolved quality control mechanisms to identify, remove, and mitigate harm from abnormal gene expression and its intermediates. Among these mechanisms is Nonsense-Mediated mRNA Decay (NMD). During NMD, cells identify and then degrade mRNAs with premature stop codons encoding truncated, deleterious, and potentially dominant-acting proteins. NMD is intimately related with human disease: ~11% of all human inherited diseases are caused by premature stop codons. Despite much effort, an understanding of how NMD target mRNAs are identified and then removed remains elusive. Our long-term goal is to illuminate how cells recognize and repress mRNAs via NMD in metazoa (animals). The key focus of this proposal is the second step of this process: the molecular details of how NMD targets are removed from the cell. My lab employs the animal C. elegans, in which the NMD machinery is non-essential, allowing us to exploit powerful genetic approaches not readily available in other animals. Two factors key for NMD target mRNA clearance are the RNA helicase SKI and the ribosome rescue factor PELO. When cells lack SKI and PELO, ribosomes stall on truncated premature stop codons of NMD target mRNAs. We demonstrated that ribosomes stalled on truncated stop codons represent an intermediate of NMD, providing a novel functional landmark in the process of mRNA removal by NMD. Building on this observation, our specific aims are: (Aim 1) We will characterize the nature and timing of RNA cleavage event(s) that give rise to ribosomes stalled on NMD target mRNAs. We will use reporters designed to test distinct models of the spatio-temporal relationship between RNA cleavage and premature translation termination. (Aim 2) We will illuminate the molecular mechanisms by which SKI and PELO remove RNAs and ribosomes during NMD, leveraging our facile ability to mutate SKI and PELO, and to assay their function in vivo. (Aim 3) Finally, we will address a question fundamental to the perceived role of NMD in preventing truncated protein production: How does NMD affect protein production from its target mRNAs? We will determine what protein species are produced during NMD, and determine whether there is repression of nascent protein chains as NMD target mRNAs are degraded. We expect that a clear picture of the cascade of events that comprise target mRNA clearance during NMD will (1) inform models of preceding events, i.e., how premature stop codons are recognized; (2) illuminate the molecular events of co-translational mRNA decay, important for understanding normal gene expression and regulation; and (3) yield a clear picture of what mRNAs and proteins are expressed from genes harboring premature stop codons, with mechanistic implications for NMD’s roles in human diseases.

项目总结/文摘