title: Multiple Natural Substitutions in Avian Influenza A Virus PB2 Facilitate Efficient Replication in Human Cells.
abstract: A strong restriction of the avian influenza A virus polymerase in mammalian cells generally limits viral host-range switching. Although substitutions like E627K in the PB2 polymerase subunit can facilitate polymerase activity to allow replication in mammals, many human H5N1 and H7N9 viruses lack this adaptive substitution. Here, several previously unknown, naturally occurring, adaptive substitutions in PB2 were identified by bioinformatics, and their enhancing activity was verified using in vitro assays. Adaptive substitutions enhanced polymerase activity and virus replication in mammalian cells for avian H5N1 and H7N9 viruses but not for a partially human-adapted H5N1 virus. Adaptive substitutions toward basic amino acids were frequent and were mostly clustered in a putative RNA exit channel in a polymerase crystal structure. Phylogenetic analysis demonstrated divergent dependency of influenza viruses on adaptive substitutions. The novel adaptive substitutions found in this study increase basic understanding of influenza virus host adaptation and will help in surveillance efforts.
  
  
  
  Influenza viruses from birds jump the species barrier into humans relatively frequently. Such influenza virus zoonoses may pose public health risks if the virus adapts to humans and becomes a pandemic threat. Relatively few amino acid substitutions-most notably in the receptor binding site of hemagglutinin and at positions 591 and 627 in the polymerase protein PB2-have been identified in pandemic influenza virus strains as determinants of host adaptation, to facilitate efficient virus replication and transmission in humans. Here, we show that substantial numbers of amino acid substitutions are functionally compensating for the lack of the above-mentioned mutations in PB2 and could facilitate influenza virus emergence in humans.
authors.affiliation: Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Mänz
authors.affiliation: Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands.
authors.forename: Miranda
authors.initials: M
authors.lastname: de Graaf
authors.affiliation: Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands.
authors.forename: Ramona
authors.initials: R
authors.lastname: Mögling
authors.affiliation: Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands.
authors.forename: Mathilde
authors.initials: M
authors.lastname: Richard
authors.affiliation: Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands.
authors.forename: Theo M
authors.initials: TM
authors.lastname: Bestebroer
authors.affiliation: Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands.
authors.forename: Guus F
authors.initials: GF
authors.lastname: Rimmelzwaan
authors.affiliation: Department of Viroscience, Erasmus Medical Center, Rotterdam, The Netherlands r.fouchier@erasmusmc.nl.
authors.forename: Ron A M
authors.initials: RAM
authors.lastname: Fouchier
mesh_terms: D000222:Adaptation, Physiological; D019943:Amino Acid Substitution; D000818:Animals; D001717:Birds; D002460:Cell Line; D019295:Computational Biology; D057809:HEK293 Cells; D058507:Host Specificity; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype; D053124:Influenza A Virus, H5N1 Subtype / Q000201:enzymology / Q000235:genetics / Q000502:physiology*; D064766:Influenza A Virus, H7N9 Subtype / Q000201:enzymology / Q000235:genetics / Q000502:physiology*; D005585:Influenza in Birds / Q000821:virology; D007251:Influenza, Human / Q000821:virology; D010802:Phylogeny; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D014779:Virus Replication
chemical_list: C057027:PB2 protein, Influenzavirus A; D014764:Viral Proteins; D012324:RNA-Dependent RNA Polymerase