title: Virulence determinants of avian H5N1 influenza A virus in mammalian and avian hosts: role of the C-terminal ESEV motif in the viral NS1 protein.
abstract: We assessed the prediction that access of the viral NS1 protein to cellular PDZ domain protein networks enhances the virulence of highly pathogenic avian influenza A viruses. The NS1 proteins of most avian influenza viruses bear the C-terminal ligand sequence Glu-Ser-Glu-Val (ESEV) for PDZ domains present in multiple host proteins, whereas no such motif is found in the NS1 homologues of seasonal human virus strains. Previous analysis showed that a C-terminal ESEV motif increases viral virulence when introduced into the NS1 protein of mouse-adapted H1N1 influenza virus. To examine the role of the PDZ domain ligand motif in avian influenza virus virulence, we generated three recombinants, derived from the prototypic H5N1 influenza A/Vietnam/1203/04 virus, expressing NS1 proteins that either have the C-terminal ESEV motif or the human influenza virus RSKV consensus or bear a natural truncation of this motif, respectively. Cell biological analyses showed strong control of NS1 nuclear migration in infected mammalian and avian cells, with only minor differences between the three variants. The ESEV sequence attenuated viral replication on cultured human, murine, and duck cells but not on chicken fibroblasts. However, all three viruses caused highly lethal infections in mice and chickens, with little difference in viral titers in organs, mean lethal dose, or intravenous pathogenicity index. These findings demonstrate that a PDZ domain ligand sequence in NS1 contributes little to the virulence of H5N1 viruses in these hosts, and they indicate that this motif modulates viral replication in a strain- and host-dependent manner.
authors.affiliation: Division of Influenza/Respiratory Viruses, Robert Koch-Institut, Nordufer 20, 13353 Berlin, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Zielecki
authors.forename: Ilia
authors.initials: I
authors.lastname: Semmler
authors.forename: Donata
authors.initials: D
authors.lastname: Kalthoff
authors.forename: Daniel
authors.initials: D
authors.lastname: Voss
authors.forename: Susanne
authors.initials: S
authors.lastname: Mauel
authors.forename: Achim D
authors.initials: AD
authors.lastname: Gruber
authors.forename: Martin
authors.initials: M
authors.lastname: Beer
authors.forename: Thorsten
authors.initials: T
authors.lastname: Wolff
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D002460:Cell Line; D002642:Chick Embryo; D002645:Chickens; D004372:Ducks; D005260:Female; D014644:Genetic Variation; D054884:Host-Pathogen Interactions / Q000235:genetics / Q000276:immunology / Q000502:physiology; D006801:Humans; D053124:Influenza A Virus, H5N1 Subtype / Q000235:genetics* / Q000472:pathogenicity* / Q000502:physiology; D005585:Influenza in Birds / Q000209:etiology / Q000821:virology; D016899:Interferon-beta / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D009976:Orthomyxoviridae Infections / Q000209:etiology / Q000473:pathology / Q000821:virology; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013045:Species Specificity; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics* / Q000502:physiology*; D014774:Virulence / Q000235:genetics / Q000502:physiology
chemical_list: C085820:INS1 protein, influenza virus; D011994:Recombinant Proteins; D017361:Viral Nonstructural Proteins; D016899:Interferon-beta