Mechanistic insights into multifaceted roles of coronavirus exoribonuclease complex
冠状病毒外核糖核酸酶复合物多方面作用的机制见解
基本信息
- 批准号:10713523
- 负责人:
- 金额:$ 38.25万
- 依托单位:
- 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:2023-08-01 至 2028-05-31
- 项目状态:未结题
- 来源:
- 关键词:2019-nCoVBiochemicalBiological ModelsBiologyBiophysicsCellsComplexCoronavirusCoupledCouplingCryoelectron MicroscopyDevelopmentDigestionDissectionEnzymesExoribonucleasesFutureGenetic TranscriptionGenomeGoalsImmune EvasionLife Cycle StagesLinkMediatingMolecularNucleotidesPhysiologicalPlayPolymeraseProcessProteinsRNARNA BiochemistryRNA CapsRNA ProcessingRNA Proof-ReadingRNA VirusesRNA chemical synthesisRegulationResearchRoleSARS-CoV-2 genomeStructureTherapeutic InterventionViralViral GenomeViral Nonstructural ProteinsViruscofactorcoronavirus treatmentdimerexperimental studyfightinginsightnovel therapeutic interventionnucleotide analogprogramssingle moleculetargeted treatmenttoolviral RNAvirology
项目摘要
As SARS-CoV-2 continues to wreak havoc across the globe, it is imperative to understand the mechanism and
regulation of the viral genome replication and transcription, which are essential processes in coronavirus life
cycle and represent important targets for therapeutic interventions. Coronavirus genome replication and
transcription are carried out by a dynamic replication-transcription complex (RTC), assembled from an array of
viral non-structural proteins (nsps). Within the RTC, a unique proofreading exoribonuclease (ExoN) complex,
nsp14-nsp10, boosts replication fidelity by excising mis-incorporated nucleotides and many antiviral nucleotide
analogs. In addition to its role in proofreading viral RNA synthesis, the ExoN complex is also involved in viral
RNA 5′ capping, which is critical for immune evasion by coronaviruses. Despite extraordinary efforts in studying
coronavirus biology and replication, major gaps remain in our understanding of the key roles ExoN complex
plays in various fundamental aspects of coronavirus life cycle. First, it is unclear how ExoN complex coordinates
with the low-fidelity viral polymerase to proofread RNA synthesis. Second, it is poorly understood how ExoN
complex is modulated by viral cofactors. Third, it is unknown how the two different enzymatic functions, RNA
cleavage and capping activities, of ExoN complex are coupled in the virus life cycle. The central objective of
our proposed experiments is to fill these gaps in understanding through a systematic dissection of the structural
basis and functional roles of ExoN complex and its dynamic interlay with viral cofactors in viral RNA synthesis
and processing. We will use SARS-CoV-2 as a model system and employ a combination of cryo-electron
microscopy, single-molecule biophysics, protein-RNA biochemistry, and cell virology to achieve this central goal
through the following aspects: Project 1, coordination of polymerase and exoribonuclease during mismatch
correction. We will define the mechanism by which RNA mismatches are transferred from polymerase to ExoN
and identify the molecular determinants for their functional interplay. Project 2, modulation of ExoN complex by
viral cofactors. We will elucidate the molecular details of the interaction between ExoN complex and a key RTC
subunit, nsp8, and determine how this interaction modulates the proofreading activity of ExoN complex during
mismatch correction. Project 3, coupling of RNA exonucleolytic digestion and 5′ capping activities of ExoN
complex. Building on our newfound cryo-EM structure of a dimeric form of the ExoN complex in which its RNA
digestion and capping activities are coupled, we will determine the molecular and biochemical underpinnings
and establish the physiological significance of the functional link between the two enzymatic activities of ExoN
complex. Through this research program, we will reveal the principles and molecular details governing the
multifaceted roles of this unique viral RNA proofreader and provide new insights into the mechanisms and
regulation of coronavirus genome replication and transcription. More broadly, the established tools and
experimental platforms are readily applicable to studying other RNA viruses.
随着SARS-CoV-2继续在全球肆虐,了解其发病机制和
病毒基因组复制和转录的调控,这是冠状病毒生命中必不可少的过程
它代表着治疗干预的重要目标。冠状病毒基因组复制和
转录是由动态复制-转录复合体(RTC)执行的,RTC由
病毒非结构蛋白(NSP)。在RTC中,有一个独特的校对外显子复合体,
NSP14-NSP10通过切除错误结合的核苷酸和许多抗病毒核苷酸来提高复制保真度
类比。除了它在校对病毒rna合成方面的作用外,外显子复合体还参与了病毒的
RNA5‘端封顶,这是冠状病毒免疫逃避的关键。尽管在学习上付出了非凡的努力
在冠状病毒的生物学和复制方面,我们对关键角色外显子复合体的理解仍然存在重大差距
在冠状病毒生命周期的各个基本方面发挥作用。首先,尚不清楚外显子是如何复配的。
用低保真病毒聚合酶校对RNA合成。其次,人们对外显子是如何产生的了解甚少
复合体受病毒辅助因子的调节。第三,目前还不清楚这两种不同的酶是如何发挥作用的,RNA
外显子复合体的切割和封端活性在病毒生命周期中是偶联的。的中心目标是
我们提议的实验是通过对结构的系统剖析来填补这些理解上的空白
病毒RNA合成中外显子复合体及其与病毒辅因子的动态相互作用
和处理中。我们将以SARS-CoV-2作为模型系统,并使用冷冻电子组合
显微镜、单分子生物物理学、蛋白质-RNA生物化学和细胞病毒学来实现这一中心目标
通过以下方面:项目1,错配过程中聚合酶和外切核糖核酸酶的协调
更正。我们将定义RNA错配从聚合酶转移到外显子的机制
并确定它们功能相互作用的分子决定因素。项目2,外显子复合体的调控
病毒辅助因子。我们将阐明外显子复合体与一个关键的RTC相互作用的分子细节
亚基,nsp8,并确定这种相互作用如何调节外显子复合体在
失配修正。项目3,核糖核酸外切与外显子5‘端封端活性的耦合
很复杂。建立在我们新发现的冷冻-EM结构的基础上,该结构是外显子复合体的二聚体形式,其RNA
消化和封顶活动是耦合的,我们将确定分子和生化基础
并建立了外显子两种酶活性之间的功能联系的生理意义
很复杂。通过这个研究项目,我们将揭示支配生物多样性的原理和分子细节。
这种独特的病毒RNA校对器的多方面作用,并提供了对机制和
冠状病毒基因组复制和转录的调控。更广泛地说,现有的工具和
实验平台很容易应用于其他RNA病毒的研究。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Yang Yang其他文献
Automatic tagging by exploring tag information capability and correlation
通过探索标签信息能力和相关性来自动标记
- DOI:
10.1007/s11280-011-0132-6 - 发表时间:
2012-05 - 期刊:
- 影响因子:0
- 作者:
Huang Zi;Shen Heng Tao;Yang Yang;Li Zhoujun - 通讯作者:
Li Zhoujun
Photodegradation of Dechlorane Plus in n-nonane under the irradiation of xenon lamp
氙灯照射下正壬烷中 DeChlorane Plus 的光降解
- DOI:
- 发表时间:
2013 - 期刊:
- 影响因子:0
- 作者:
Yang Yang;Gang Yu;Shubo Deng;Bin Wang - 通讯作者:
Bin Wang
Is Ignorance Bliss?
无知是福吗?
- DOI:
10.1016/j.jebo.2019.02.025 - 发表时间:
2019 - 期刊:
- 影响因子:2.2
- 作者:
Yang Yang - 通讯作者:
Yang Yang
Structure-independent conductance of thiophene-based single-stacking junctions
- DOI:
10.1002/anie.201913344. - 发表时间:
2020 - 期刊:
- 影响因子:
- 作者:
Li Xiaohui;Wu Qingqing;Bai Jie;Hou Songjun;Jiang Wenlin;Tang Chun;Song Hang;Huang Xiaojuan;Zheng Jueting;Yang Yang;Liu Junyang;Hu Yong;Shi Jia;Liu Zitong;Lambert Colin J.;Zhang Deqing;Hong Wenjing - 通讯作者:
Hong Wenjing
Yang Yang的其他文献
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{{ truncateString('Yang Yang', 18)}}的其他基金
CK22-008, Building Mathematical Modeling Workforce Capacity to Support Infectious Disease and Healthcare Research - 2022
CK22-008,建立数学建模劳动力能力以支持传染病和医疗保健研究 - 2022
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- 资助金额:
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用于立体选择性自由基生物催化的可进化金属酶平台
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10650814 - 财政年份:2022
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Rapid 3D Ultrasound Tomography Reconstruction Methods for Guided Interventions
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- 批准号:
10670956 - 财政年份:2022
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$ 38.25万 - 项目类别:
Rapid 3D Ultrasound Tomography Reconstruction Methods for Guided Interventions
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10509562 - 财政年份:2022
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Neuronal and Behavioral Deficits Associated with Scn2a Deficiency in Autism Spectrum Disorder
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10599103 - 财政年份:2021
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