title: Balance of RNA sequence requirement and NS3/NS3a expression of segment 10 of orbiviruses.
abstract: Orbiviruses are insect-transmitted, non-enveloped viruses with a ten-segmented dsRNA genome of which the bluetongue virus (BTV) is the prototype. Viral non-structural protein NS3/NS3a is encoded by genome segment 10 (Seg-10), and is involved in different virus release mechanisms. This protein induces specific release via membrane disruptions and budding in both insect and mammalian cells, but also the cytopathogenic release that is only seen in mammalian cells. NS3/NS3a is not essential for virus replication in vitro with BTV Seg-10 containing RNA elements essential for virus replication, even if protein is not expressed. Recently, new BTV serotypes with distinct NS3/NS3a sequence and cell tropism have been identified. Multiple studies have hinted at the importance of Seg-10 in orbivirus replication, but the exact prerequisites are still unknown. Here, more insight is obtained with regard to the needs for orbivirus Seg-10 and the balance between protein expression and RNA elements. Multiple silent mutations in the BTV NS3a ORF destabilized Seg-10, resulting in deletions and sequences originating from other viral segments being inserted, indicating strong selection at the level of RNA during replication in mammalian cells in vitro. The NS3a ORFs of other orbiviruses were successfully exchanged in BTV1 Seg-10, resulting in viable chimeric viruses. NS3/NS3a proteins in these chimeric viruses were generally functional in mammalian cells, but not in insect cells. NS3/NS3a of the novel BTV serotypes 25 and 26 affected virus release from Culicoides cells, which might be one of the reasons for their distinct cell tropism.
authors.affiliation: Department of Virology, Central Veterinary Institute of Wageningen UR (CVI), Lelystad, The Netherlands.|Department of Infectious Diseases & Immunology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.
authors.forename: Femke
authors.initials: F
authors.lastname: Feenstra
authors.affiliation: Department of Virology, Central Veterinary Institute of Wageningen UR (CVI), Lelystad, The Netherlands.
authors.forename: René G P
authors.initials: RGP
authors.lastname: van Gennip
authors.affiliation: Department of Virology, Central Veterinary Institute of Wageningen UR (CVI), Lelystad, The Netherlands.
authors.forename: Myrte
authors.initials: M
authors.lastname: Schreuder
authors.affiliation: Department of Biochemistry, Centre for Human Metabolomics, North-West University, South Africa.|Department of Virology, Central Veterinary Institute of Wageningen UR (CVI), Lelystad, The Netherlands.
authors.forename: Piet A
authors.initials: PA
authors.lastname: van Rijn
mesh_terms: D000818:Animals; D002460:Cell Line; D002519:Ceratopogonidae; D015967:Gene Expression Regulation, Viral; D042822:Genomic Instability; D008322:Mammals; D009154:Mutation; D018163:Orbivirus / Q000502:physiology*; D012367:RNA, Viral / Q000235:genetics; D017361:Viral Nonstructural Proteins / Q000096:biosynthesis*; D056189:Viral Tropism; D057074:Virus Release
chemical_list: D012367:RNA, Viral; D017361:Viral Nonstructural Proteins