Coupling of N7-methyltransferase and 3′-5′ exoribonuclease with SARS-CoV-2 polymerase reveals mechanisms for capping and proofreading

被引：121

作者：

Yan, Liming ^{[1
,2
,3
]}

Yang, Yunxiang ^{[1
,5
]}

Li, Mingyu ^{[1
]}

Zhang, Ying ^{[1
]}

Zheng, Litao ^{[2
,3
,4
]}

Ge, Ji ^{[4
]}

Huang, Yucen C. ^{[1
]}

Liu, Zhenyu ^{[1
]}

Wang, Tao ^{[1
,2
,3
]}

Gao, Shan ^{[1
]}

Zhang, Ran ^{[4
]}

Huang, Yuanyun Y. ^{[6
]}

Guddat, Luke W. ^{[7
]}

Gao, Yan ^{[2
,3
]}

Rao, Zihe ^{[1
,2
,3
,4
,8
,9
,10
,11
]}

Lou, Zhiyong ^{[1
,11
]}

机构：

[1] Tsinghua Univ, Sch Med, MOE Key Lab Prot Sci, Beijing, Peoples R China

[2] ShanghaiTech Univ, Shanghai Inst Adv Immunochem Studies, Shanghai, Peoples R China

[3] ShanghaiTech Univ, Sch Life Sci & Technol, Shanghai, Peoples R China

[4] Tsinghua Univ, Sch Life Sci, Beijing, Peoples R China

[5] Tsinghua Univ, Tsinghua Univ Peking Univ Joint Ctr Life Sci, Sch Life Sci, Beijing, Peoples R China

[6] CALTECH, Div Chem & Chem Engn, Pasadena, CA 91125 USA

[7] Univ Queensland, Sch Chem & Mol Biosci, Brisbane, Qld, Australia

[8] Nankai Univ, Coll Life Sci, State Key Lab Med Chem Biol, Tianjin, Peoples R China

[9] Nankai Univ, Coll Pharm, Tianjin, Peoples R China

[10] Chinese Acad Sci, CAS Ctr Excellence Biomacromol, Inst Biophys, Natl Lab Biomacromol, Beijing, Peoples R China

[11] Guangzhou Lab, Guangzhou, Peoples R China

来源：

CELL | 2021年 / 184卷 / 13期

基金：

中国国家自然科学基金;

关键词：

CRYO-EM STRUCTURE; STRUCTURAL BASIS; RNA-POLYMERASE; HIGH-FIDELITY; REPLICATION; TRANSCRIPTION; VIRUS; NSP14; BACKTRACKING; EXCISION;

D O I：

10.1016/j.cell.2021.05.033

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

The capping of mRNA and the proofreading play essential roles in SARS-CoV-2 replication and transcription. Here, we present the cryo-EM structure of the SARS-CoV-2 replication-transcription complex (RTC) in a form identified as Cap(0)-RTC, which couples a co-transcriptional capping complex (CCC) composed of nsp12 NiRAN, nsp9, the bifunctional nsp14 possessing an N-terminal exoribonuclease (ExoN) and a C-terminal N7-methyltransferase (N7-MTase), and nsp10 as a cofactor of nsp14. Nsp9 and nsp12 NiRAN recruit nsp10/nsp14 into the Cap(0)-RTC, forming the N7-CCC to yield cap(0) ((7Me)GpppA) at 5' end of pre-mRNA. A dimeric form of Cap(0)-RTC observed by cryo-EM suggests an in trans backtracking mechanism for nsp14 ExoN to facilitate proofreading of the RNA in concert with polymerase nsp12. These results not only provide a structural basis for understanding co-transcriptional modification of SARS-CoV-2 mRNA but also shed light on how replication fidelity in SARS-CoV-2 is maintained.

引用

页码：3474 / +

页数：23

共 54 条

[1] New tools for the analysis and validation of cryo-EM maps and atomic models [J].

Afonine, Pavel V. ;

Klaholz, Bruno P. ;

Moriarty, Nigel W. ;

Poon, Billy K. ;

Sobolev, Oleg V. ;

Terwilliger, Thomas C. ;

Adams, Paul D. ;

Urzhumtsev, Alexandre .

ACTA CRYSTALLOGRAPHICA SECTION D-STRUCTURAL BIOLOGY, 2018, 74 :814-840

[2] RNA 3′-end mismatch excision by the severe acute respiratory syndrome coronavirus nonstructural protein nsp10/nsp14 exoribonuclease complex [J].

Bouvet, Mickael ;

Imbert, Isabelle ;

Subissi, Lorenzo ;

Gluais, Laure ;

Canard, Bruno ;

Decroly, Etienne .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2012, 109 (24) :9372-9377

[3] In Vitro Reconstitution of SARS-Coronavirus mRNA Cap Methylation [J].

Bouvet, Mickael ;

Debarnot, Claire ;

Imbert, Isabelle ;

Selisko, Barbara ;

Snijder, Eric J. ;

Canard, Bruno ;

Decroly, Etienne .

PLOS PATHOGENS, 2010, 6 (04) :1-13

[4] Mutagenesis of S-Adenosyl-L-Methionine-Binding Residues in Coronavirus nsp14 N7-Methyltransferase Demonstrates Differing Requirements for Genome Translation and Resistance to Innate Immunity [J].

Case, James Brett ;

Ashbrook, Alison W. ;

Dermody, Terence S. ;

Denison, Mark R. .

JOURNAL OF VIROLOGY, 2016, 90 (16) :7248-7256

[5] Structural Basis for Helicase-Polymerase Coupling in the SARS-CoV-2 Replication-Transcription Complex [J].

Chen, James ;

Malone, Brandon ;

Llewellyn, Eliza ;

Grasso, Michael ;

Shelton, Patrick M. M. ;

Olinares, Paul Dominic B. ;

Maruthi, Kashyap ;

Eng, Edward T. ;

Vatandaslar, Hasan ;

Chait, Brian T. ;

Kapoor, Tarun M. ;

Darst, Seth A. ;

Campbell, Elizabeth A. .

CELL, 2020, 182 (06) :1560-+

[6] High-resolution noise substitution to measure overfitting and validate resolution in 3D structure determination by single particle electron cryomicroscopy [J].

Chen, Shaoxia ;

McMullan, Greg ;

Faruqi, Abdul R. ;

Murshudov, Garib N. ;

Short, Judith M. ;

Scheres, Sjors H. W. ;

Henderson, Richard .

ULTRAMICROSCOPY, 2013, 135 :24-35

[7] Biochemical and Structural Insights into the Mechanisms of SARS Coronavirus RNA Ribose 2′-O-Methylation by nsp16/nsp10 Protein Complex [J].

Chen, Yu ;

Su, Ceyang ;

Ke, Min ;

Jin, Xu ;

Xu, Lirong ;

Zhang, Zhou ;

Wu, Andong ;

Sun, Ying ;

Yang, Zhouning ;

Tien, Po ;

Ahola, Tero ;

Liang, Yi ;

Liu, Xinqi ;

Guo, Deyin .

PLOS PATHOGENS, 2011, 7 (10)

[8] Functional screen reveals SARS coronavirus nonstructural protein nsp14 as a novel cap N7 methyltransferase [J].

Chen, Yu ;

Cai, Hui ;

Pan, Ji'an ;

Xiang, Nian ;

Tien, Po ;

Ahola, Tero ;

Guo, Deyin .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2009, 106 (09) :3484-3489

[9] Structural basis of RNA polymerase II backtracking, arrest and reactivation [J].

Cheung, Alan C. M. ;

Cramer, Patrick .

NATURE, 2011, 471 (7337) :249-253

[10] 2′-O methylation of the viral mRNA cap evades host restriction by IFIT family members [J].

Daffis, Stephane ;

Szretter, Kristy J. ;

Schriewer, Jill ;

Li, Jianqing ;

Youn, Soonjeon ;

Errett, John ;

Lin, Tsai-Yu ;

Schneller, Stewart ;

Zust, Roland ;

Dong, Hongping ;

Thiel, Volker ;

Sen, Ganes C. ;

Fensterl, Volker ;

Klimstra, William B. ;

Pierson, Theodore C. ;

Buller, R. Mark ;

Gale, Michael, Jr. ;

Shi, Pei-Yong ;

Diamond, Michael S. .

NATURE, 2010, 468 (7322) :452-456

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