title: The Nucleocapsid Protein of Coronaviruses Acts as a Viral Suppressor of RNA Silencing in Mammalian Cells.
abstract: RNA interference (RNAi) is a process of eukaryotic posttranscriptional gene silencing that functions in antiviral immunity in plants, nematodes, and insects. However, recent studies provided strong supports that RNAi also plays a role in antiviral mechanism in mammalian cells. To combat RNAi-mediated antiviral responses, many viruses encode viral suppressors of RNA silencing (VSR) to facilitate their replication. VSRs have been widely studied for plant and insect viruses, but only a few have been defined for mammalian viruses currently. We identified a novel VSR from coronaviruses, a group of medically important mammalian viruses including Severe acute respiratory syndrome coronavirus (SARS-CoV), and showed that the nucleocapsid protein (N protein) of coronaviruses suppresses RNAi triggered by either short hairpin RNAs or small interfering RNAs in mammalian cells. Mouse hepatitis virus (MHV) is closely related to SARS-CoV in the family Coronaviridae and was used as a coronavirus replication model. The replication of MHV increased when the N proteins were expressed in trans, while knockdown of Dicer1 or Ago2 transcripts facilitated the MHV replication in mammalian cells. These results support the hypothesis that RNAi is a part of the antiviral immunity responses in mammalian cells. IMPORTANCE RNAi has been well known to play important antiviral roles from plants to invertebrates. However, recent studies provided strong supports that RNAi is also involved in antiviral response in mammalian cells. An important indication for RNAi-mediated antiviral activity in mammals is the fact that a number of mammalian viruses encode potent suppressors of RNA silencing. Our results demonstrate that coronavirus N protein could function as a VSR through its double-stranded RNA binding activity. Mutational analysis of N protein allowed us to find out the critical residues for the VSR activity. Using the MHV-A59 as the coronavirus replication model, we showed that ectopic expression of SARS-CoV N protein could promote MHV replication in RNAi-active cells but not in RNAi-depleted cells. These results indicate that coronaviruses encode a VSR that functions in the replication cycle and provide further evidence to support that RNAi-mediated antiviral response exists in mammalian cells.
authors.forename: Lei
authors.initials: L
authors.lastname: Cui
authors.forename: Haiying
authors.initials: H
authors.lastname: Wang
authors.forename: Yanxi
authors.initials: Y
authors.lastname: Ji
authors.forename: Jie
authors.initials: J
authors.lastname: Yang
authors.forename: Shan
authors.initials: S
authors.lastname: Xu
authors.forename: Xingyu
authors.initials: X
authors.lastname: Huang
authors.forename: Zidao
authors.initials: Z
authors.lastname: Wang
authors.forename: Lei
authors.initials: L
authors.lastname: Qin
authors.forename: Po
authors.initials: P
authors.lastname: Tien
authors.forename: Xi
authors.initials: X
authors.lastname: Zhou
authors.forename: Deyin
authors.initials: D
authors.lastname: Guo
authors.forename: Yu
authors.initials: Y
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D060565:Argonaute Proteins / Q000235:genetics; D001483:Base Sequence; D002460:Cell Line; D017934:Coronavirus / Q000235:genetics / Q000276:immunology; D000086462:Coronavirus Nucleocapsid Proteins; D053487:DEAD-box RNA Helicases / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D007739:L Cells; D051379:Mice; D006517:Murine hepatitis virus / Q000235:genetics* / Q000254:growth & development / Q000276:immunology; D019590:Nucleocapsid Proteins / Q000096:biosynthesis / Q000235:genetics*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics*; D043244:Ribonuclease III / Q000235:genetics; D016415:Sequence Alignment; D045169:Severe Acute Respiratory Syndrome / Q000235:genetics* / Q000821:virology
chemical_list: C526735:Ago2 protein, mouse; D060565:Argonaute Proteins; D000086462:Coronavirus Nucleocapsid Proteins; D019590:Nucleocapsid Proteins; D034741:RNA, Small Interfering; C509781:Dicer1 protein, mouse; D043244:Ribonuclease III; D053487:DEAD-box RNA Helicases