title: A 6K-deletion variant of salmonid alphavirus is non-viable but can be rescued through RNA recombination.
abstract: Pancreas disease (PD) of Atlantic salmon is an emerging disease caused by Salmonid alphavirus (SAV) which mainly affects salmonid aquaculture in Western Europe. Although genome structure of SAV has been characterized and each individual viral protein has been identified, the role of 6K protein in viral replication and infectivity remains undefined. The 6K protein of alphaviruses is a small and hydrophobic protein which is involved in membrane permeabilization, protein processing and virus budding. Because these common features are shared across many viral species, they have been named viroporins. In the present study, we applied reverse genetics to generate SAV3 6K-deleted (Δ6K) variant and investigate the role of 6K protein. Our findings show that the 6K-deletion variant of salmonid alphavirus is non-viable. Despite viral proteins of Δ6K variant are detected in the cytoplasm by immunostaining, they are not found on the cell surface. Further, analysis of viral proteins produced in Δ6K cDNA clone transfected cells using radioimmunoprecipitation (RIPA) and western blot showed a protein band of larger size than E2 of wild-type SAV3. When Δ6K cDNA was co-transfected with SAV3 helper cDNA encoding the whole structural genes including 6K, the infectivity was rescued. The development of CPE after co-transfection and resolved genome sequence of rescued virus confirmed full-length viral genome being generated through RNA recombination. The discovery of the important role of the 6K protein in virus production provides a new possibility for the development of antiviral intervention which is highly needed to control SAV infection in salmonids.
authors.affiliation: Norwegian University of Life Sciences, Department of Basic Sciences and Aquatic Medicine, Oslo, Norway.
authors.forename: Tz-Chun
authors.initials: TC
authors.lastname: Guo
authors.affiliation: Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Daniel X
authors.initials: DX
authors.lastname: Johansson
authors.affiliation: Norwegian University of Life Sciences, Department of Basic Sciences and Aquatic Medicine, Oslo, Norway.
authors.forename: Øyvind
authors.initials: Ø
authors.lastname: Haugland
authors.affiliation: Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Peter
authors.initials: P
authors.lastname: Liljeström
authors.affiliation: Norwegian University of Life Sciences, Department of Basic Sciences and Aquatic Medicine, Oslo, Norway.
authors.forename: Øystein
authors.initials: Ø
authors.lastname: Evensen
mesh_terms: D000524:Alphavirus / Q000187:drug effects / Q000235:genetics* / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics; D017353:Gene Deletion; D016898:Interferon-alpha / Q000494:pharmacology; D051379:Mice; D050296:Microbial Viability / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017684:Oncorhynchus kisutch / Q000821:virology; D012367:RNA, Viral / Q000235:genetics*; D011995:Recombination, Genetic; D014162:Transfection; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D014771:Virion / Q000187:drug effects / Q000502:physiology
chemical_list: D000906:Antibodies; D018076:DNA, Complementary; D016898:Interferon-alpha; D012367:RNA, Viral; D014764:Viral Proteins