A unifying structural and functional model of the coronavirus replication organelle: Tracking down RNA synthesis

Zoonotic coronavirus (CoV) infections, such as those responsible for the current severe acute respiratory syndrome-CoV 2 (SARS-CoV-2) pandemic, cause grave international public health concern. In infected cells, the CoV RNA-synthesizing machinery associates with modified endoplasmic reticulum membra...

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Bibliographic Details
Published in:	PLoS biology Vol. 18; no. 6; p. e3000715
Main Authors:	Snijder, Eric J, Limpens, Ronald W A L, de Wilde, Adriaan H, de Jong, Anja W M, Zevenhoven-Dobbe, Jessika C, Maier, Helena J, Faas, Frank F G A, Koster, Abraham J, Bárcena, Montserrat
Format:	Journal Article
Language:	English
Published:	United States Public Library of Science 08-06-2020 Public Library of Science (PLoS)
Subjects:	Animals Autoradiography Betacoronavirus - genetics Betacoronavirus - physiology Biology Biology and life sciences Bronchitis Cell Line Cell organelles Cellular structure Chlorocebus aethiops Coronaviridae Coronavirus - classification Coronavirus - physiology Coronavirus Infections - pathology Coronavirus Infections - virology Coronaviruses COVID-19 Electron Microscope Tomography Electron microscopy Endoplasmic reticulum Endoplasmic Reticulum - pathology Endoplasmic Reticulum - ultrastructure Endoplasmic Reticulum - virology Formation Funding Humans Infections Laboratories Medicine and Health Sciences Membrane structures Membrane vesicles Membranes Metabolites Microscopy Middle East respiratory syndrome Middle East Respiratory Syndrome Coronavirus - physiology Pandemics Pneumonia, Viral - pathology Pneumonia, Viral - virology Public health Replication Research and Analysis Methods Respiratory diseases RNA RNA viruses RNA, Viral - metabolism SARS-CoV-2 Severe acute respiratory syndrome Severe acute respiratory syndrome coronavirus 2 Spherules Structure-function relationships Synthesis Therapeutic targets Tomography Transcription Two dimensional analysis Vero Cells Viral diseases Virology Virus Replication Viruses Netherlands
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Summary:	Zoonotic coronavirus (CoV) infections, such as those responsible for the current severe acute respiratory syndrome-CoV 2 (SARS-CoV-2) pandemic, cause grave international public health concern. In infected cells, the CoV RNA-synthesizing machinery associates with modified endoplasmic reticulum membranes that are transformed into the viral replication organelle (RO). Although double-membrane vesicles (DMVs) appear to be a pan-CoV RO element, studies to date describe an assortment of additional CoV-induced membrane structures. Despite much speculation, it remains unclear which RO element(s) accommodate viral RNA synthesis. Here we provide detailed 2D and 3D analyses of CoV ROs and show that diverse CoVs essentially induce the same membrane modifications, including the small open double-membrane spherules (DMSs) previously thought to be restricted to gamma- and delta-CoV infections and proposed as sites of replication. Metabolic labeling of newly synthesized viral RNA followed by quantitative electron microscopy (EM) autoradiography revealed abundant viral RNA synthesis associated with DMVs in cells infected with the beta-CoVs Middle East respiratory syndrome-CoV (MERS-CoV) and SARS-CoV and the gamma-CoV infectious bronchitis virus. RNA synthesis could not be linked to DMSs or any other cellular or virus-induced structure. Our results provide a unifying model of the CoV RO and clearly establish DMVs as the central hub for viral RNA synthesis and a potential drug target in CoV infection.
Bibliography:	new_version ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Current address: Janssen Vaccines and Prevention, Pharmaceutical Companies of Johnson and Johnson, Leiden, the Netherlands The authors have declared that no competing interests exist.
ISSN:	1545-7885 1544-9173 1545-7885
DOI:	10.1371/journal.pbio.3000715