Structural basis of protein synthesis by the hepatitis C virus IRES RNA

丙型肝炎病毒IRES RNA合成蛋白质的结构基础

• 批准号: 9068943
• 负责人: Jeffrey S Kieft
• 金额: $ 29.3万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-01 至 2017-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9068943
• 关键词: Address; Automobile Driving; Binding; Biochemical; Biochemistry; Biological Assay; Cell Culture Techniques; Cells; Cellular Stress; Crystallization; Data; Development; Engineering; Eukaryotic Initiation Factor-2; Future; Goals; Health; Hepatitis C virus; Human; Immune response; Infection; Internal Ribosome Entry Site; Literature; Messenger RNA; Methods; Modeling; Molecular; Molecular Conformation; Pathway interactions; Peptide Initiation Factors; Process; Production; Protein Biosynthesis; Proteins; RNA; RNA Binding; RNA Viruses; Research; Resolution; Ribosomes; Role; Stress; Structure; Testing; Translation Initiation; Translations; Viral Genome; Viral Proteins; Virus; base; experience; interest; meetings; new therapeutic target; novel strategies; novel therapeutics; pathogen; structural biology; tool; viral RNA; working group

DESCRIPTION (provided by applicant): Hepatitis C virus (HCV) is an example of a virus that uses an internal ribosome entry site (IRES) RNA to drive production of all the viral proteins. The HCV IRES RNA forms a specific structure that binds directly to the 40S ribosomal subunit and in so doing changes the conformation of the subunit. This physical manipulation of the ribosome appears to be a critical component of the mechanism by which the HCV IRES "hijacks" the translation machinery. However, how the IRES accomplishes these structural changes, how they propagate through the ribosome, and how the induced structures compare to canonical ribosome conformations remains unknown. In addition, there is growing evidence that the HCV IRES is one component of the virus' strategy to evade the host cell's innate immune response, in part by driving translation initiation by alternate, eukaryotic initiation factor (eIF) 2-independent pathways operating during cellular stress. While the mechanism by which the HCV IRES operates in unstressed cells is well-studied, the pathway used in stressed cells is very poorly understood. We seek to explore the ability of the IRES to manipulate the host cell's machinery by accessing alternate translation initiation pathways and also through physical manipulation of the ribosome. We propose two aims: 1) Define the IRES domains, factors, and mechanisms that enable the HCV IRES to operate under conditions of cellular stress, and 2) Determine the high-resolution structure of an HCV IRES RNA bound to a ribosome or ribosomal subunit. To accomplish these aims, we will use a combination of biochemistry, cell culture-based assays, and structural biology in an integrated approach focused on developing new models for HCV IRES function at an unprecedented level of detail. Several of the methods and tools we will use have been developed in our lab and thus are unique to our lab. Our studies promise to contribute new discoveries to how IRESs function, how mammalian ribosomes operate, how ribosomes can be manipulated, and how translation occurs during periods of cell stress.

描述（由申请人提供）：丙型肝炎病毒（HCV）是使用内部核糖体进入位点（IRES）RNA来驱动所有病毒蛋白产生的病毒的例子。 HCV IRES RNA 形成直接与 40S 核糖体亚基结合的特定结构，从而改变该亚基的构象。这种对核糖体的物理操作似乎是 HCV IRES“劫持”翻译机器机制的关键组成部分。然而，IRES如何实现这些结构变化、它们如何通过核糖体传播以及诱导的结构如何与规范核糖体构象进行比较仍然未知。此外，越来越多的证据表明，HCV IRES 是病毒逃避宿主细胞先天免疫反应策略的组成部分，部分是通过在细胞应激期间运行的替代真核起始因子 (eIF) 2 独立途径驱动翻译起始。虽然 HCV IRES 在未受应激的细胞中发挥作用的机制已得到充分研究，但在受应激的细胞中使用的途径却知之甚少。我们试图通过访问替代翻译起始途径以及通过对核糖体的物理操作来探索 IRES 操纵宿主细胞机器的能力。我们提出两个目标：1) 定义使 HCV IRES 在细胞应激条件下运行的 IRES 结构域、因素和机制，2) 确定与核糖体或核糖体亚基结合的 HCV IRES RNA 的高分辨率结构。为了实现这些目标，我们将采用生物化学、基于细胞培养的检测和结构生物学的综合方法，重点以前所未有的细节水平开发 HCV IRES 功能的新模型。我们将使用的几种方法和工具是我们实验室开发的，因此是我们实验室独有的。我们的研究有望为 IRES 如何发挥作用、哺乳动物核糖体如何运作、核糖体如何被操纵以及细胞应激期间翻译如何发生等方面做出新的发现。

项目成果

The structures of nonprotein-coding RNAs that drive internal ribosome entry site function.

• DOI: 10.1002/wrna.1105
• 发表时间: 2012-03
• 期刊: WILEY INTERDISCIPLINARY REVIEWS-RNA
• 影响因子: 7.3
• 作者: Plank, Terra-Dawn M.;Kieft, Jeffrey S.
• 通讯作者: Kieft, Jeffrey S.

HCV IRES manipulates the ribosome to promote the switch from translation initiation to elongation.

• DOI: 10.1038/nsmb.2465
• 发表时间: 2013-02
• 期刊: NATURE STRUCTURAL & MOLECULAR BIOLOGY
• 影响因子: 16.8
• 作者: Filbin, Megan E.;Vollmar, Breanna S.;Shi, Dan;Gonen, Tamir;Kieft, Jeffrey S.
• 通讯作者: Kieft, Jeffrey S.

RNA structures that resist degradation by Xrn1 produce a pathogenic Dengue virus RNA.

• DOI: 10.7554/elife.01892
• 发表时间: 2014-04-01
• 期刊: eLife
• 影响因子: 7.7
• 作者: Chapman EG;Moon SL;Wilusz J;Kieft JS
• 通讯作者: Kieft JS

An approach to the construction of tailor-made amphiphilic peptides that strongly and selectively bind to hairpin RNA targets.

• DOI: 10.1021/ja807609m
• 发表时间: 2009-02-18
• 期刊: Journal of the American Chemical Society
• 影响因子: 15
• 作者: Lee SJ;Hyun S;Kieft JS;Yu J
• 通讯作者: Yu J

The structural basis of transfer RNA mimicry and conformational plasticity by a viral RNA.

• DOI: 10.1038/nature13378
• 发表时间: 2014-07-17
• 期刊: NATURE
• 影响因子: 64.8
• 作者: Colussi, Timothy M.;Costantino, David A.;Hammond, John A.;Ruehle, Grant M.;Nix, Jay C.;Kieft, Jeffrey S.
• 通讯作者: Kieft, Jeffrey S.