title: Compatibility among polymerase subunit proteins is a restricting factor in reassortment between equine H7N7 and human H3N2 influenza viruses.
abstract: Reassortment is an important driving force for influenza virus evolution, and a better understanding of the factors that affect this process could improve our ability to respond to future influenza pandemics and epidemics. To identify factors that restrict the generation of reassortant viruses, we cotransfected human embryonic kidney cells with plasmids for the synthesis of viral RNAs of both A/equine/Prague/1/56 (Prague; H7N7) and A/Yokohama/2017/03 (Yokohama; H3N2) viruses together with the supporting protein expression plasmids. Of the possible 256 genotypes, we identified 29 genotypes in 120 randomly plaque-picked reassortants examined. Analyses of these reassortants suggested that the formation of functional ribonucleoprotein (RNP) complexes was a restricting factor, a finding that correlated with the activities of RNP complexes composed of different combinations of the proteins from the two viruses, as measured in a minigenome assay. For at least one nonfunctional RNP complex (i.e., Prague PB2, Prague PB1, Yokohama PA, and Prague NP), the lack of activity was due to the inability of the three polymerase subunit proteins to form a heterotrimer. Adaptation of viruses possessing a gene encoding a chimera of the PA proteins of the two viruses and the remaining genes from Prague virus resulted in compensatory mutations in the PB2 and/or PA protein. These results indicate substantial incompatibility among the gene products of the two test viruses, a critical role for the RNP complex in the generation of reassortant viruses, and a functional interaction of PB2 and PA.
authors.affiliation: Department of Pathobiological Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, 2015 Linden Drive, Madison, WI 53706, USA.
authors.forename: Chengjun
authors.initials: C
authors.lastname: Li
authors.forename: Masato
authors.initials: M
authors.lastname: Hatta
authors.forename: Shinji
authors.initials: S
authors.lastname: Watanabe
authors.forename: Gabriele
authors.initials: G
authors.lastname: Neumann
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Kawaoka
mesh_terms: D001483:Base Sequence; D002478:Cells, Cultured; D005838:Genotype; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000502:physiology; D006801:Humans; D053122:Influenza A Virus, H3N2 Subtype / Q000235:genetics*; D053126:Influenza A Virus, H7N7 Subtype / Q000235:genetics*; D008969:Molecular Sequence Data; D019590:Nucleocapsid Proteins; D021122:Protein Subunits; D016601:RNA-Binding Proteins / Q000502:physiology; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry / Q000502:physiology*; D016865:Reassortant Viruses / Q000235:genetics*; D012261:Ribonucleoproteins / Q000502:physiology; D014758:Viral Core Proteins / Q000502:physiology; D014764:Viral Proteins / Q000502:physiology
chemical_list: D019267:Hemagglutinin Glycoproteins, Influenza Virus; C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; C502893:PB2 protein, influenza virus; D021122:Protein Subunits; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins; D014758:Viral Core Proteins; D014764:Viral Proteins; C077346:influenza virus polymerase basic protein 1; D012324:RNA-Dependent RNA Polymerase