title: Dual Mutation Events in the Haemagglutinin-Esterase and Fusion Protein from an Infectious Salmon Anaemia Virus HPR0 Genotype Promote Viral Fusion and Activation by an Ubiquitous Host Protease.
abstract: In Infectious salmon anaemia virus (ISAV), deletions in the highly polymorphic region (HPR) in the near membrane domain of the haemagglutinin-esterase (HE) stalk, influence viral fusion. It is suspected that selected mutations in the associated Fusion (F) protein may also be important in regulating fusion activity. To better understand the underlying mechanisms involved in ISAV fusion, several mutated F proteins were generated from the Scottish Nevis and Norwegian SK779/06 HPR0. Co-transfection with constructs encoding HE and F were performed, fusion activity assessed by content mixing assay and the degree of proteolytic cleavage by western blot. Substitutions in Nevis F demonstrated that K276 was the most likely cleavage site in the protein. Furthermore, amino acid substitutions at three sites and two insertions, all slightly upstream of K276, increased fusion activity. Co-expression with HE harbouring a full-length HPR produced high fusion activities when trypsin and low pH were applied. In comparison, under normal culture conditions, groups containing a mutated HE with an HPR deletion were able to generate moderate fusion levels, while those with a full length HPR HE could not induce fusion. This suggested that HPR length may influence how the HE primes the F protein and promotes fusion activation by an ubiquitous host protease and/or facilitate subsequent post-cleavage refolding steps. Variations in fusion activity through accumulated mutations on surface glycoproteins have also been reported in other orthomyxoviruses and paramyxoviruses. This may in part contribute to the different virulence and tissue tropism reported for HPR0 and HPR deleted ISAV genotypes.
authors.affiliation: Aquaculture and Fish Health, Marine Scotland Science, Aberdeen, United Kingdom.
authors.forename: Mickael
authors.initials: M
authors.lastname: Fourrier
authors.affiliation: Aquaculture and Fish Health, Marine Scotland Science, Aberdeen, United Kingdom.
authors.forename: Katherine
authors.initials: K
authors.lastname: Lester
authors.affiliation: Epidemiology, Norwegian Veterinary Institute, Oslo, Norway.
authors.forename: Turhan
authors.initials: T
authors.lastname: Markussen
authors.affiliation: Epidemiology, Norwegian Veterinary Institute, Oslo, Norway.
authors.forename: Knut
authors.initials: K
authors.lastname: Falk
authors.affiliation: School of Biological Sciences, University of Aberdeen, Aberdeen, United Kingdom.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Secombes
authors.affiliation: Aquaculture and Fish Health, Marine Scotland Science, Aberdeen, United Kingdom.
authors.forename: Alastair
authors.initials: A
authors.lastname: McBeath
authors.affiliation: Aquaculture and Fish Health, Marine Scotland Science, Aberdeen, United Kingdom.
authors.forename: Bertrand
authors.initials: B
authors.lastname: Collet
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D029941:Fish Proteins / Q000378:metabolism; D006389:Hemagglutinins, Viral / Q000235:genetics*; D046829:Isavirus / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D008969:Molecular Sequence Data; D009154:Mutation; D010447:Peptide Hydrolases / Q000378:metabolism; D012474:Salmon; D014760:Viral Fusion Proteins / Q000235:genetics*; D053586:Virus Internalization
chemical_list: D029941:Fish Proteins; D006389:Hemagglutinins, Viral; D014760:Viral Fusion Proteins; C060814:hemagglutinin esterase; D010447:Peptide Hydrolases