{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,1]],"date-time":"2026-10-01T16:00:05Z","timestamp":1790870405320,"version":"4.1.0"},"reference-count":49,"publisher":"Oxford University Press (OUP)","issue":"10","license":[{"start":{"date-parts":[[2024,3,5]],"date-time":"2024-03-05T00:00:00Z","timestamp":1709596800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by-nc\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000060","name":"NIAID","doi-asserted-by":"publisher","award":["AI123498"],"award-info":[{"award-number":["AI123498"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"NIAID","doi-asserted-by":"publisher","award":["AI158463"],"award-info":[{"award-number":["AI158463"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"NIAID","doi-asserted-by":"publisher","award":["WIS03099"],"award-info":[{"award-number":["WIS03099"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000002","name":"NIH","doi-asserted-by":"publisher","award":["R35GM118110"],"award-info":[{"award-number":["R35GM118110"]}],"id":[{"id":"10.13039\/100000002","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2024,6,10]]},"abstract":"<jats:title>Abstract<\/jats:title>\n               <jats:p>Coronaviruses are a diverse subfamily of viruses containing pathogens of humans and animals. This subfamily of viruses replicates their RNA genomes using a core polymerase complex composed of viral non-structural proteins: nsp7, nsp8 and nsp12. Most of our understanding of coronavirus molecular biology comes from betacoronaviruses like SARS-CoV and SARS-CoV-2, the latter of which is the causative agent of COVID-19. In contrast, members of the alphacoronavirus genus are relatively understudied despite their importance in human and animal health. Here we have used cryo-electron microscopy to determine structures of the alphacoronavirus porcine epidemic diarrhea virus (PEDV) core polymerase complex bound to RNA. One structure shows an unexpected nsp8 stoichiometry despite remaining bound to RNA. Biochemical analysis shows that the N-terminal extension of one nsp8 is not required for in vitro RNA synthesis for alpha- and betacoronaviruses. Our work demonstrates the importance of studying diverse coronaviruses in revealing aspects of coronavirus replication and identifying areas of conservation to be targeted by antiviral drugs.<\/jats:p>","DOI":"10.1093\/nar\/gkae153","type":"journal-article","created":{"date-parts":[[2024,3,5]],"date-time":"2024-03-05T20:09:38Z","timestamp":1709669378000},"page":"5975-5986","source":"Crossref","is-referenced-by-count":15,"title":["An alphacoronavirus polymerase structure reveals conserved replication factor functions"],"prefix":"10.1093","volume":"52","author":[{"given":"Thomas K","family":"Anderson","sequence":"first","affiliation":[{"name":"Biochemistry Department, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Institute for Molecular Virology, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Center for Quantitative Cell Imaging, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peter J","family":"Hoferle","sequence":"additional","affiliation":[{"name":"Biochemistry Department, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Institute for Molecular Virology, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Center for Quantitative Cell Imaging, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kennan J","family":"Chojnacki","sequence":"additional","affiliation":[{"name":"Biochemistry Department, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Institute for Molecular Virology, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Center for Quantitative Cell Imaging, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kenneth\u00a0W","family":"Lee","sequence":"additional","affiliation":[{"name":"Biomolecular Chemistry Department, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joshua\u00a0J","family":"Coon","sequence":"additional","affiliation":[{"name":"Biomolecular Chemistry Department, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Department of Chemistry, University of Wisconsin-Madison , Madison , WI\u00a0 53715 , USA"},{"name":"Morgridge Institute for Research , Madison , WI\u00a0 53715 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5974-2709","authenticated-orcid":false,"given":"Robert\u00a0N","family":"Kirchdoerfer","sequence":"additional","affiliation":[{"name":"Biochemistry Department, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Institute for Molecular Virology, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"},{"name":"Center for Quantitative Cell Imaging, University of Wisconsin-Madison , Madison , WI\u00a0 53706 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"286","published-online":{"date-parts":[[2024,3,5]]},"reference":[{"key":"2024060923573174700_B1","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1007\/978-1-4939-2438-7_1","article-title":"Coronaviruses: an overview of their replication and pathogenesis","volume":"1282","author":"Fehr","year":"2015","journal-title":"Methods Mol. Biol."},{"key":"2024060923573174700_B2","doi-asserted-by":"crossref","first-page":"991","DOI":"10.1016\/S0022-2836(03)00865-9","article-title":"Unique and conserved features of genome and proteome of SARS-coronavirus, an early split-off from the coronavirus group 2 lineage","volume":"331","author":"Snijder","year":"2003","journal-title":"J. Mol. Biol."},{"key":"2024060923573174700_B3","doi-asserted-by":"crossref","first-page":"1814","DOI":"10.1056\/NEJMoa1211721","article-title":"Isolation of a novel coronavirus from a man with pneumonia in Saudi Arabia","volume":"367","author":"Zaki","year":"2012","journal-title":"N. Engl. J. Med."},{"key":"2024060923573174700_B4","doi-asserted-by":"crossref","first-page":"7790","DOI":"10.1128\/JVI.01244-13","article-title":"Middle East respiratory syndrome coronavirus (MERS-CoV): announcement of the Coronavirus Study Group","volume":"87","author":"de\u00a0Groot","year":"2013","journal-title":"J. Virol."},{"key":"2024060923573174700_B5","doi-asserted-by":"crossref","first-page":"265","DOI":"10.1038\/s41586-020-2008-3","article-title":"A new coronavirus associated with human respiratory disease in China","volume":"579","author":"Wu","year":"2020","journal-title":"Nature"},{"key":"2024060923573174700_B6","doi-asserted-by":"crossref","first-page":"270","DOI":"10.1038\/s41586-020-2012-7","article-title":"A pneumonia outbreak associated with a new coronavirus of probable bat origin","volume":"579","author":"Zhou","year":"2020","journal-title":"Nature"},{"key":"2024060923573174700_B7","first-page":"004","article-title":"Human Respiratory coronaviruses detected In patients with influenza-like illness in Arkansas, USA","volume":"2014","author":"Silva","year":"2014","journal-title":"Virol. Mycol."},{"key":"2024060923573174700_B8","doi-asserted-by":"crossref","first-page":"104391","DOI":"10.1016\/j.jcv.2020.104391","article-title":"A RT-PCR assay for the detection of coronaviruses from four genera","volume":"128","author":"Xiu","year":"2020","journal-title":"J. Clin. Virol."},{"key":"2024060923573174700_B9","doi-asserted-by":"crossref","first-page":"446","DOI":"10.1093\/cid\/ciab456","article-title":"Novel canine coronavirus isolated from a hospitalized patient with pneumonia in East Malaysia","volume":"74","author":"Vlasova","year":"2022","journal-title":"Clin. Infect. Dis."},{"key":"2024060923573174700_B10","doi-asserted-by":"crossref","first-page":"3953","DOI":"10.3390\/ijms23073953","article-title":"Swine Enteric Coronavirus: diverse pathogen-host interactions","volume":"23","author":"Yan","year":"2022","journal-title":"Int. J. Mol. Sci."},{"key":"2024060923573174700_B11","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1007\/s11259-021-09808-0","article-title":"Porcine enteric coronaviruses: an updated overview of the pathogenesis, prevalence, and diagnosis","volume":"45","author":"Liu","year":"2021","journal-title":"Vet. Res. Commun."},{"key":"2024060923573174700_B12","doi-asserted-by":"crossref","first-page":"2434","DOI":"10.3390\/v14112434","article-title":"Porcine Epidemic Diarrhea Virus: an updated overview of virus epidemiology, virulence variation patterns and virus-host interactions","volume":"14","author":"Zhang","year":"2022","journal-title":"Viruses"},{"key":"2024060923573174700_B13","doi-asserted-by":"crossref","first-page":"649","DOI":"10.1177\/1040638713501675","article-title":"Emergence of porcine epidemic diarrhea virus in the United States: clinical signs, lesions, and viral genomic sequences","volume":"25","author":"Stevenson","year":"2013","journal-title":"J. Vet. Diagn. Invest."},{"key":"2024060923573174700_B14","doi-asserted-by":"crossref","first-page":"161","DOI":"10.3201\/eid1801.111259","article-title":"Outbreak of porcine epidemic diarrhea in suckling piglets, China","volume":"18","author":"Sun","year":"2012","journal-title":"Emerg. Infect. Dis."},{"key":"2024060923573174700_B15","first-page":"1","article-title":"Coronavirus genome structure and replication","volume":"287","author":"Brian","year":"2005","journal-title":"Curr. Top. Microbiol. Immunol."},{"key":"2024060923573174700_B16","doi-asserted-by":"crossref","first-page":"7117","DOI":"10.1093\/nar\/gkg916","article-title":"Molecular model of SARS coronavirus polymerase: implications for biochemical functions and drug design","volume":"31","author":"Xu","year":"2003","journal-title":"Nucleic Acids Res."},{"key":"2024060923573174700_B17","doi-asserted-by":"crossref","first-page":"203","DOI":"10.1093\/nar\/gkp904","article-title":"The RNA polymerase activity of SARS-coronavirus nsp12 is primer dependent","volume":"38","author":"te\u00a0Velthuis","year":"2010","journal-title":"Nucleic Acids Res."},{"key":"2024060923573174700_B18","doi-asserted-by":"crossref","first-page":"59","DOI":"10.1016\/bs.aivir.2016.08.008","article-title":"The nonstructural proteins directing coronavirus RNA synthesis and processing","volume":"96","author":"Snijder","year":"2016","journal-title":"Adv. Virus Res."},{"key":"2024060923573174700_B19","doi-asserted-by":"crossref","first-page":"2342","DOI":"10.1038\/s41467-019-10280-3","article-title":"Structure of the SARS-CoV nsp12 polymerase bound to nsp7 and nsp8 co-factors","volume":"10","author":"Kirchdoerfer","year":"2019","journal-title":"Nat. Commun."},{"key":"2024060923573174700_B20","doi-asserted-by":"crossref","first-page":"184","DOI":"10.1016\/j.cell.2020.11.016","article-title":"Cryo-EM structure of an extended SARS-CoV-2 replication and transcription complex reveals an intermediate State in cap synthesis","volume":"184","author":"Yan","year":"2020","journal-title":"Cell"},{"key":"2024060923573174700_B21","doi-asserted-by":"crossref","first-page":"E3900","DOI":"10.1073\/pnas.1323705111","article-title":"One severe acute respiratory syndrome coronavirus protein complex integrates processive RNA polymerase and exonuclease activities","volume":"111","author":"Subissi","year":"2014","journal-title":"Proc. Natl. Acad. Sci. U.S.A."},{"key":"2024060923573174700_B22","doi-asserted-by":"crossref","first-page":"e70968","DOI":"10.7554\/eLife.70968","article-title":"Inhibition of SARS-CoV-2 polymerase by nucleotide analogs from a single-molecule perspective","volume":"10","author":"Seifert","year":"2021","journal-title":"eLife"},{"key":"2024060923573174700_B23","doi-asserted-by":"crossref","first-page":"154","DOI":"10.1038\/s41586-020-2368-8","article-title":"Structure of replicating SARS-CoV-2 polymerase","volume":"584","author":"Hillen","year":"2020","journal-title":"Nature"},{"key":"2024060923573174700_B24","doi-asserted-by":"crossref","first-page":"1499","DOI":"10.1126\/science.abc1560","article-title":"Structural basis for inhibition of the RNA-dependent RNA polymerase from SARS-CoV-2 by remdesivir","volume":"368","author":"Yin","year":"2020","journal-title":"Science"},{"key":"2024060923573174700_B25","doi-asserted-by":"crossref","first-page":"1548","DOI":"10.1016\/j.molcel.2021.01.035","article-title":"Remdesivir is a delayed translocation inhibitor of SARS-CoV-2 replication","volume":"81","author":"Bravo","year":"2021","journal-title":"Mol. Cell"},{"key":"2024060923573174700_B26","doi-asserted-by":"crossref","first-page":"5874","DOI":"10.1038\/s41467-020-19770-1","article-title":"Architecture of a SARS-CoV-2 mini replication and transcription complex","volume":"11","author":"Yan","year":"2020","journal-title":"Nat. Commun."},{"key":"2024060923573174700_B27","doi-asserted-by":"crossref","first-page":"1560","DOI":"10.1016\/j.cell.2020.07.033","article-title":"Structural basis for helicase-polymerase coupling in the SARS-CoV-2 replication-transcription complex","volume":"182","author":"Chen","year":"2020","journal-title":"Cell"},{"key":"2024060923573174700_B28","doi-asserted-by":"crossref","first-page":"e2102516118","DOI":"10.1073\/pnas.2102516118","article-title":"Structural basis for backtracking by the SARS-CoV-2 replication-transcription complex","volume":"118","author":"Malone","year":"2021","journal-title":"Proc. Natl. Acad. Sci. U.S.A."},{"key":"2024060923573174700_B29","doi-asserted-by":"crossref","first-page":"4444","DOI":"10.1128\/JVI.06635-11","article-title":"Nonstructural proteins 7 and 8 of feline coronavirus form a 2:1 heterotrimer that exhibits primer-independent RNA polymerase activity","volume":"86","author":"Xiao","year":"2012","journal-title":"J. Virol."},{"key":"2024060923573174700_B30","doi-asserted-by":"crossref","first-page":"eabf1004","DOI":"10.1126\/sciadv.abf1004","article-title":"Hallmarks of alpha- and betacoronavirus non-structural protein 7+8 complexes","volume":"7","author":"Krichel","year":"2021","journal-title":"Sci. Adv."},{"key":"2024060923573174700_B31","doi-asserted-by":"crossref","first-page":"105455","DOI":"10.1016\/j.compbiomed.2022.105455","article-title":"Conserved protein targets for developing pan-coronavirus drugs based on sequence and 3D structure similarity analyses","volume":"145","author":"Ma","year":"2022","journal-title":"Comput. Biol. Med."},{"key":"2024060923573174700_B32","doi-asserted-by":"crossref","first-page":"e0067122","DOI":"10.1128\/jvi.00671-22","article-title":"The SARS-CoV nsp12 polymerase active site is tuned for large-genome replication","volume":"96","author":"Campagnola","year":"2022","journal-title":"J. Virol."},{"key":"2024060923573174700_B33","doi-asserted-by":"crossref","first-page":"671","DOI":"10.1038\/nmeth.2089","article-title":"NIH image to ImageJ: 25 years of image analysis","volume":"9","author":"Schneider","year":"2012","journal-title":"Nat. Methods"},{"key":"2024060923573174700_B34","doi-asserted-by":"crossref","first-page":"36","DOI":"10.1016\/j.jsb.2005.07.007","article-title":"Automated electron microscope tomography using robust prediction of specimen movements","volume":"152","author":"Mastronarde","year":"2005","journal-title":"J. Struct. Biol."},{"key":"2024060923573174700_B35","doi-asserted-by":"crossref","first-page":"290","DOI":"10.1038\/nmeth.4169","article-title":"cryoSPARC: algorithms for rapid unsupervised cryo-EM structure determination","volume":"14","author":"Punjani","year":"2017","journal-title":"Nat. Methods"},{"key":"2024060923573174700_B36","doi-asserted-by":"crossref","first-page":"107702","DOI":"10.1016\/j.jsb.2021.107702","article-title":"3D variability analysis: resolving continuous flexibility and discrete heterogeneity from single particle cryo-EM","volume":"213","author":"Punjani","year":"2021","journal-title":"J. Struct. Biol."},{"key":"2024060923573174700_B37","doi-asserted-by":"crossref","first-page":"486","DOI":"10.1107\/S0907444910007493","article-title":"Features and development of Coot","volume":"66","author":"Emsley","year":"2010","journal-title":"Acta. Crystallogr. D Biol. Crystallogr."},{"key":"2024060923573174700_B38","doi-asserted-by":"crossref","first-page":"531","DOI":"10.1107\/S2059798318006551","article-title":"Real-space refinement in PHENIX for cryo-EM and crystallography","volume":"74","author":"Afonine","year":"2018","journal-title":"Acta Crystallogr D Struct Biol"},{"key":"2024060923573174700_B39","doi-asserted-by":"crossref","first-page":"519","DOI":"10.1107\/S2059798318002425","article-title":"ISOLDE: a physically realistic environment for model building into low-resolution electron-density maps","volume":"74","author":"Croll","year":"2018","journal-title":"Acta Crystallogr. D Struct. Biol."},{"key":"2024060923573174700_B40","doi-asserted-by":"crossref","first-page":"70","DOI":"10.1002\/pro.3943","article-title":"UCSF ChimeraX: structure visualization for researchers, educators, and developers","volume":"30","author":"Pettersen","year":"2021","journal-title":"Protein Sci."},{"key":"2024060923573174700_B41","doi-asserted-by":"crossref","first-page":"621","DOI":"10.1038\/s41467-022-28113-1","article-title":"A dual mechanism of action of AT-527 against SARS-CoV-2 polymerase","volume":"13","author":"Shannon","year":"2022","journal-title":"Nat. Commun."},{"key":"2024060923573174700_B42","doi-asserted-by":"crossref","first-page":"e00221-18","DOI":"10.1128\/mBio.00221-18","article-title":"Coronavirus susceptibility to the antiviral remdesivir (GS-5734) is mediated by the viral polymerase and the proofreading exoribonuclease","volume":"9","author":"Agostini","year":"2018","journal-title":"mBio"},{"key":"2024060923573174700_B43","doi-asserted-by":"crossref","first-page":"6785","DOI":"10.1074\/jbc.RA120.013679","article-title":"Remdesivir is a direct-acting antiviral that inhibits RNA-dependent RNA polymerase from severe acute respiratory syndrome coronavirus 2 with high potency","volume":"295","author":"Gordon","year":"2020","journal-title":"J. Biol. Chem."},{"key":"2024060923573174700_B44","doi-asserted-by":"crossref","first-page":"4773","DOI":"10.1074\/jbc.AC120.013056","article-title":"The antiviral compound remdesivir potently inhibits RNA-dependent RNA polymerase from Middle East respiratory syndrome coronavirus","volume":"295","author":"Gordon","year":"2020","journal-title":"J. Biol. Chem."},{"key":"2024060923573174700_B45","doi-asserted-by":"crossref","first-page":"53","DOI":"10.1016\/j.bbrc.2022.07.087","article-title":"ATP enhances the error-prone ribonucleotide incorporation by the SARS-CoV-2 RNA polymerase","volume":"625","author":"Pourfarjam","year":"2022","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"2024060923573174700_B46","doi-asserted-by":"crossref","first-page":"279","DOI":"10.1038\/s41467-020-20542-0","article-title":"Mechanism of SARS-CoV-2 polymerase stalling by remdesivir","volume":"12","author":"Kokic","year":"2021","journal-title":"Nat. Commun."},{"key":"2024060923573174700_B47","doi-asserted-by":"crossref","first-page":"2228","DOI":"10.3390\/v13112228","article-title":"Curing cats with feline infectious peritonitis with an oral multi-component drug containing GS-441524","volume":"13","author":"Krentz","year":"2021","journal-title":"Viruses"},{"key":"2024060923573174700_B48","doi-asserted-by":"crossref","first-page":"101923","DOI":"10.1016\/j.jbc.2022.101923","article-title":"Replication of the coronavirus genome: a paradox among positive-strand RNA viruses","volume":"298","author":"Grellet","year":"2022","journal-title":"J. Biol. Chem."},{"key":"2024060923573174700_B49","doi-asserted-by":"crossref","first-page":"5956","DOI":"10.1093\/nar\/gkab370","article-title":"Two conserved oligomer interfaces of NSP7 and NSP8 underpin the dynamic assembly of SARS-CoV-2 RdRP","volume":"49","author":"Biswal","year":"2021","journal-title":"Nucleic Acids Res."}],"container-title":["Nucleic Acids Research"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/academic.oup.com\/nar\/article-pdf\/52\/10\/5975\/58172679\/gkae153.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/academic.oup.com\/nar\/article-pdf\/52\/10\/5975\/58172679\/gkae153.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2024,6,9]],"date-time":"2024-06-09T23:58:15Z","timestamp":1717977495000},"score":1,"resource":{"primary":{"URL":"https:\/\/academic.oup.com\/nar\/article\/52\/10\/5975\/7619545"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,3,5]]},"references-count":49,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2024,3,5]]},"published-print":{"date-parts":[[2024,6,10]]}},"URL":"https:\/\/doi.org\/10.1093\/nar\/gkae153","relation":{},"ISSN":["0305-1048","1362-4962"],"issn-type":[{"value":"0305-1048","type":"print"},{"value":"1362-4962","type":"electronic"}],"subject":[],"published-other":{"date-parts":[[2024,6,10]]},"published":{"date-parts":[[2024,3,5]]}}}