Mechanisms of viral RNA maturation by co-opting cellular exonucleases

通过选择细胞核酸外切酶使病毒 RNA 成熟的机制

基本信息

批准号：
10814079
负责人：
Jeffrey S Kieft
金额：
$ 53.46万
依托单位：
NEW YORK STRUCTURAL BIOLOGY CENTER
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-06-01 至 2027-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10814079
关键词：
3&apos Untranslated Regions 3-Dimensional 5&apos Untranslated Regions Address Antiviral Response Architecture Attenuated Biochemistry Bioinformatics Biophysics Cells Characteristics Classification Code Complex Coupled Coupling Cryoelectron Microscopy Cultured Cells Dangerousness Data Dengue Dengue Virus Directed Molecular Evolution Disease Elements Environment Enzymes Exonuclease Exoribonucleases Flavivirus Health In Vitro Individual Infection Japanese encephalitis virus Kinetics Knowledge Laboratories Learning Link Location Mechanics Modeling Molecular Outcome Pathogenicity Pattern Powassan virus Process Production Property Proteins RNA RNA Decay RNA Folding RNA Viruses Regulation Research Resistance Resolution Structure Surface Therapeutic Intervention Tick-Borne Encephalitis Virus Untranslated RNA Variant Viral Viral Genome Virus Virus Diseases West Nile virus Work X-Ray Crystallography Yellow fever virus ZIKA Zika Virus arthropod-borne computerized tools genomic RNA human pathogen insight mosquito-borne poly A specific exoribonuclease structural biology success three dimensional structure tick-borne tick-borne flavivirus viral RNA viral genomics virology

项目摘要

PROJECT SUMMARY Flaviviruses are single-stranded positive-sense RNA viruses that include dangerous human pathogens like dengue, West Nile, Yellow Fever, Zika, and many others. During infection, these viruses produce a set of non- coding RNAs called ‘subgenomic flavivirus RNAs’ (sfRNAs) that interact with cellular proteins to manipulate the cellular environment, to include inhibiting the antiviral response. sfRNAs have been directly linked to cytopathic and pathogenic outcomes, and viruses that cannot produce sfRNAs are attenuated, motivating efforts to understand the mechanism of xrRNA production. sfRNAs are made when cellular 5’à3’ exoribonucleases (in particular, Xrn1) processively degrade the viral genomic RNA but then halt at specifically structured RNA elements in the viral 3’ UTR called exoribonuclease resistant RNAs (xrRNAs). By solving the structures of multiple xrRNAs by x-ray crystallography and combining this with biochemistry, biophysics, and virology, we showed that xrRNAs fold into a unique ring-like topology that creates a mechanical block the exoribonuclease cannot pass through. Furthermore, we used our discoveries to classify xrRNAs and to find new examples associated with both non-coding and coding RNAs. These successes now define several new questions. First, xrRNAs are often found in multiple copies ‘in tandem’ where their function is coupled in some way, but the structural basis of this coupling, and the effects of breaking the coupling on both sfRNA formation and viral infection kinetics, are unknown. Second, although we have a good understanding of several classes of xrRNAs, we have yet to solve the structure of an xrRNA from a tick-borne flavivirus, which appear to have interesting and unique properties. Third, although we have found many new examples of xrRNAs, it appears there are many more that are undiscovered, and we also do not understand how the various classes of xrRNA relate evolutionarily. How do these structures diversify and evolve in 3-D given the tight constraints on their folding? Here, we propose to answer these questions in three aims, employing a strategy that combines biochemistry, x- ray crystallography, cryo-EM, virology, and in vitro selections coupled with computational tools. The research described here will contribute significant basic knowledge regarding an important molecular process of broad applicability to viral disease, a necessary step between the discovery of a mechanism and the targeting of it for therapeutic intervention.

项目摘要黄病毒是单链阳性RNA病毒，包括危险的人类病原体登革热，西尼罗河，黄热病，寨卡病毒等。在感染期间，这些病毒产生一组非 - 编码RNA称为“亚基因组黄病毒RNA”（SFRNA），与细胞蛋白相互作用以操纵细胞环境，包括抑制抗病毒反应。 SFRNA已直接与细胞病变有关和致病结果以及无法产生SFRNA的病毒被减弱，激励着努力了解XRRNA产生的机制。当细胞5'à3'exoribonleboneleases（在特别是，XRN1）在过程中降解病毒基因组RNA，但在特定结构的RNA上停止病毒3'UTR中的元素称为埃塞哥利核酸酶耐药RNA（XRRNA）。通过解决结构通过X射线晶体学的多个XRRNA，并将其与生物化学，生物物理学和病毒学结合起来，我们表明Xrrnas折叠成独特的环形拓扑，从而产生机械块，使驱虫夹酶无法通过。此外，我们利用我们的发现来对XRRNA进行分类并找到新的示例与非编码和编码RNA有关。这些成功现在定义了几个新问题。第一的， Xrrnas通常以多个副本“双人”的方式找到，其中其功能以某种方式耦合，但是该耦合的结构基础，以及破坏耦合对SFRNA形成和病毒的影响感染动力学是未知的。其次，尽管我们对几类XRRNA有很好的了解，但我们尚未从tick传播黄病毒中解决Xrrna的结构，该结构似乎很有趣，并且独特的属性。第三，尽管我们发现了许多Xrrnas的新示例，但似乎有很多未发现的更多内容，我们也不了解XRRNA的各种类别如何相关进化。鉴于它们的折叠严重约束，这些结构如何在3D中多样化和发展？在这里，我们建议在三个目标中回答这些问题，采用一种结合生物化学的策略，x- 射线晶体学，冷冻EM，病毒学和体外选择以及计算工具。研究此处描述的将为广泛的重要分子过程提供重要的基础知识适用于病毒疾病，这是发现机制的必要步骤治疗干预。

项目成果

期刊论文数量（12）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Zika virus dumbbell-1 structure is critical for sfRNA presence and cytopathic effect during infection.

DOI：
10.1128/mbio.01108-23
发表时间：
2023-08-31
期刊：
mBio
影响因子：
6.4
作者：
通讯作者：

A New Subclass of Exoribonuclease-Resistant RNA Found in Multiple Genera of Flaviviridae.

DOI：
10.1128/mbio.02352-20
发表时间：
2020-09-29
期刊：
mBio
影响因子：
6.4
作者：
Szucs MJ;Nichols PJ;Jones RA;Vicens Q;Kieft JS
通讯作者：
Kieft JS

Thoughts on how to think (and talk) about RNA structure.

DOI：
10.1073/pnas.2112677119
发表时间：
2022-04-26
期刊：
Proceedings of the National Academy of Sciences of the United States of America
影响因子：
11.1
作者：
通讯作者：

Mechanism and structural diversity of exoribonuclease-resistant RNA structures in flaviviral RNAs.

DOI：
10.1038/s41467-017-02604-y
发表时间：
2018-01-09
期刊：
Nature communications
影响因子：
16.6
作者：
MacFadden A;O'Donoghue Z;Silva PAGC;Chapman EG;Olsthoorn RC;Sterken MG;Pijlman GP;Bredenbeek PJ;Kieft JS
通讯作者：
Kieft JS

Exoribonuclease-Resistant RNAs Exist within both Coding and Noncoding Subgenomic RNAs