title: Identification and Characterization of a Ribose 2'-O-Methyltransferase Encoded by the Ronivirus Branch of Nidovirales.
abstract: The order Nidovirales currently comprises four virus families: Arteriviridae, Coronaviridae (divided into the subfamilies Coronavirinae and Torovirinae), Roniviridae, and the recently recognized Mesoniviridae RNA cap formation and methylation have been best studied for coronaviruses, with emphasis on the identification and characterization of two virus-encoded methyltransferases (MTases) involved in RNA capping, a guanine-N7-MTase and a ribose-2'-O-MTase. Although bioinformatics analyses suggest that these MTases may also be encoded by other nidoviruses with large genomes, such as toroviruses and roniviruses, no experimental evidence has been reported thus far. In this study, we show that a ronivirus, gill-associated virus (GAV), encodes the 2'-O-MTase activity, although we could not detect 2'-O-MTase activity for the homologous protein of a torovirus, equine torovirus, which is more closely related to coronaviruses. Like the coronavirus 2'-O-MTase, the roniviral 2'-O-MTase harbors a catalytic K-D-K-E tetrad that is conserved among 2'-O-MTases and can target only the N7-methylated cap structure of adenylate-primed RNA substrates. However, in contrast with the coronavirus protein, roniviral 2'-O-MTase does not require a protein cofactor for stimulation of its activity and differs in its preference for several biochemical parameters, such as reaction temperature and pH. Furthermore, the ronivirus 2'-O-MTase can be targeted by MTase inhibitors. These results extend our current understanding of nidovirus RNA cap formation and methylation beyond the coronavirus family.
  
  
  
  Methylation of the 5'-cap structure of viral RNAs plays important roles in genome replication and evasion of innate recognition of viral RNAs by cellular sensors. It is known that coronavirus nsp14 acts as an N7-(guanine)-methyltransferase (MTase) and nsp16 as a 2'-O-MTase, which are involved in the modification of RNA cap structure. However, these enzymatic activities have not been shown for any other nidoviruses beyond coronaviruses in the order Nidovirales In this study, we identified a 2'-O-methyltransferase encoded by ronivirus that shows common and unique features in comparison with that of coronaviruses. Ronivirus 2'-O-MTase does not need a protein cofactor for MTase activity, whereas coronavirus nsp16 needs the stimulating factor nsp10 for its full activity. The conserved K-D-K-E catalytic tetrad is identified in ronivirus 2'-O-MTase. These results extend our understanding of nidovirus RNA capping and methylation beyond coronaviruses and also strengthen the evolutionary and functional links between roniviruses and coronaviruses.
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Cong
authors.initials: C
authors.lastname: Zeng
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Andong
authors.initials: A
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Shan
authors.initials: S
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Yingke
authors.initials: Y
authors.lastname: Tang
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Xu
authors.initials: X
authors.lastname: Jin
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Shilei
authors.initials: S
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Lei
authors.initials: L
authors.lastname: Qin
authors.affiliation: Department of Pathogen Biology, Henan University of TCM, Zhengzhou, Henan, People's Republic of China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Sun
authors.affiliation: Medical Research Institute, School of Basic Medical Sciences, Wuhan University, Wuhan, China.
authors.forename: Chengpeng
authors.initials: C
authors.lastname: Fan
authors.affiliation: Molecular Virology Laboratory, Department of Medical Microbiology, Leiden University Medical Center, RC Leiden, The Netherlands.
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Snijder
authors.affiliation: School of Biological Sciences, University of Reading, Reading, United Kingdom.
authors.forename: Benjamin W
authors.initials: BW
authors.lastname: Neuman
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Department of Food and Environmental Sciences, University of Helsinki, Helsinki, Finland.
authors.forename: Tero
authors.initials: T
authors.lastname: Ahola
authors.affiliation: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, People's Republic of China dguo@whu.edu.cn.|Medical Research Institute, School of Basic Medical Sciences, Wuhan University, Wuhan, China.
authors.forename: Deyin
authors.initials: D
authors.lastname: Guo
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D008745:Methylation; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism*; D028381:Nidovirales / Q000201:enzymology* / Q000235:genetics; D030341:Nidovirales Infections / Q000235:genetics / Q000378:metabolism / Q000821:virology; D017434:Protein Structure, Tertiary; D012315:RNA Caps / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D012266:Ribose / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
chemical_list: D001426:Bacterial Proteins; D012315:RNA Caps; D012367:RNA, Viral; D012266:Ribose; D008780:Methyltransferases; C052744:RNA 2'-O-methyltransferase