Host-range determinants on the PB2 protein of influenza A viruses control the interaction between the viral polymerase and nucleoprotein in human cells.
Karine Labadie1
Emmanuel Dos Santos Afonso
Marie-Anne Rameix-Welti
Sylvie van der Werf
Nadia Naffakh
Affiliations1 institutions
Unité de Génétique Moléculaire des Virus Respiratoires, URA 1966 CNRS, EA302 Université Paris 7, Institut Pasteur, 25 rue du Dr. Roux, 75724 Paris Cedex 15, France.
The transcription/replication activity of ribonucleoproteins derived from influenza A primary isolates of human (A/Paris/908/97) or avian origin (A/Mallard/Marquenterre/MZ237/83, A/Hong Kong/156/97) was compared upon reconstitution in mammalian or avian cells, using viral-like reporter RNAs synthesized under the control of the human and chicken RNA polymerase I promoters, respectively. In avian cells, transcription/replication activities were in the same range with all ribonucleoproteins tested. In human cells, ribonucleoproteins derived from A/Mallard/Marquenterre/MZ237/83 showed reduced transcription/replication activity and reduced NP binding to the PB1-PB2-PA complex (P) or to the isolated PB2 subunit, as compared to the ribonucleoproteins derived from A/Paris/908/97. Both defects were restored when PB2 residue Glu-627 was changed to a Lys. Ribonucleoproteins derived from the human A/Hong Kong/156/97 H5N1 isolate showed efficient NP-P interaction in human cells, and high levels of activity which were determined mostly by the PB2 and PA proteins. Our data suggest that PB2 might play a pivotal role in molecular interactions involving both the viral nucleoprotein and cellular proteins.
Protein Interaction MappingAnimalsCell LineChickensChloramphenicol O-AcetyltransferaseChlorocebus aethiopsCOS CellsGenes, ReporterHumansInfluenza A virusInfluenza A Virus, H1N1 SubtypeInfluenza A Virus, H3N2 SubtypeInfluenza A Virus, H5N1 SubtypeMolecular Sequence DataPromoter Regions, GeneticRibonucleoproteinsRNA Polymerase IRNA, Viral
Structured evidence records
Evidence records
3 total
Host Range Experiment2 records
Host Range ExperimentExtraction confidence 0.90
Key finding
Influenza A ribonucleoproteins from human and avian isolates showed differential transcription and replication activity depending on whether they were reconstituted in mammalian or avian cells, identifying PB2 residue 627 as a determinant of host range.
The transcription/replication activity of ribonucleoproteins derived from influenza A primary isolates of human (A/Paris/908/97) or avian origin (A/Mallard/Marquenterre/MZ237/83, A/Hong Kong/156/97) was compared upon reconstitution in mammalian or avian cells, using viral-like reporter RNAs synthesized under the control of the human and chicken RNA polymerase I promoters, respectively.
Ribonucleoproteins derived from the avian strain A/Mallard/Marquenterre/MZ237/83 displayed reduced transcription and replication activity in human cells compared to human-derived strains, indicating limited compatibility of avian PB2 in human cells.
In avian cells, transcription/replication activities were in the same range with all ribonucleoproteins tested. In human cells, ribonucleoproteins derived from A/Mallard/Marquenterre/MZ237/83 showed reduced transcription/replication activity and reduced NP binding.
Method
ribonucleoprotein reconstitution; transcription/replication assay; protein interaction assay
Experimental system
in vitro cell culture
Molecular Adaptation1 records
Molecular AdaptationExtraction confidence 0.95
Key finding
Changing PB2 residue Glu-627 to Lys in avian influenza A viruses restored polymerase activity and NP interaction in human cells, showing PB2 E627K mediates adaptation to mammalian hosts.
Both defects were restored when PB2 residue Glu-627 was changed to a Lys. Ribonucleoproteins derived from the human A/Hong Kong/156/97 H5N1 isolate showed efficient NP-P interaction in human cells, and high levels of activity which were determined mostly by the PB2 and PA proteins.
