title: Identification of a noncanonically transcribed subgenomic mRNA of infectious bronchitis virus and other gammacoronaviruses.
abstract: Coronavirus subgenomic mRNA (sgmRNA) synthesis occurs via a process of discontinuous transcription involving transcription regulatory sequences (TRSs) located in the 5' leader sequence (TRS-L) and upstream of each structural and group-specific gene (TRS-B). Several gammacoronaviruses including infectious bronchitis virus (IBV) contain a putative open reading frame (ORF), localized between the M gene and gene 5, which is controversial due to the perceived absence of a TRS. We have studied the transcription of a novel sgmRNA associated with this potential ORF and found it to be transcribed via a previously unidentified noncanonical TRS-B. Using an IBV reverse genetics system, we demonstrated that the template-switching event during intergenic region (IR) sgmRNA synthesis occurs at the 5' end of the noncanonical TRS-B and recombines between nucleotides 5 and 6 of the 8-nucleotide consensus TRS-L. Introduction of a complete TRS-B showed that higher transcription levels are achieved by increasing the number of nucleotide matches between TRS-L and TRS-B. Translation of a protein from the sgmRNA was demonstrated using enhanced green fluorescent protein, suggesting the translation of a fifth, novel, group-specific protein for IBV. This study has resolved an issue concerning the number of ORFs expressed by members of the Gammacoronavirus genus and proposes the existence of a fifth IBV accessory protein. We confirmed previous reports that coronaviruses can produce sgmRNAs from noncanonical TRS-Bs, which may expand their repertoire of proteins. We also demonstrated that noncanonical TRS-Bs may provide a mechanism by which coronaviruses can control protein expression levels by reducing sgmRNA synthesis.
authors.affiliation: Avian Viral Diseases, The Pirbright Institute, Compton Laboratory, Compton, Newbury, Berkshire, United Kingdom.
authors.forename: Kirsten
authors.initials: K
authors.lastname: Bentley
authors.forename: Sarah May
authors.initials: SM
authors.lastname: Keep
authors.forename: Maria
authors.initials: M
authors.lastname: Armesto
authors.forename: Paul
authors.initials: P
authors.lastname: Britton
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D003332:Coronaviridae / Q000235:genetics*; D015967:Gene Expression Regulation, Viral; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000235:genetics; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D012333:RNA, Messenger; D012367:RNA, Viral; D049452:Green Fluorescent Proteins