PB2 Residue 271 Plays a Key Role in Enhanced Polymerase Activity of Influenza A Viruses in Mammalian Host Cells

Abstract

The direct infection of humans with highly pathogenic avian H5N1 influenza viruses has suggested viral mutation as one mechanism for the emergence of novel human influenza A viruses. Although the polymerase complex is known to be a key component in host adaptation, mutations that enhance the polymerase activity of avian viruses in mammalian hosts are not fully characterized. The genomic comparison of influenza A virus isolates has identified highly conserved residues in influenza proteins that are specific to either human or avian viruses, including 10 residues in PB2. We characterized the activity of avian polymerase complexes containing avian-to-human mutations at these conserved PB2 residues and found that, in addition to the E627K mutation, the PB2 mutation T271A enhances polymerase activity in human cells. We confirmed the effects of the T271A mutation using recombinant WSN viruses containing avian NP and polymerase genes with wild-type (WT) or mutant PB2. The 271A virus showed enhanced growth compared to that of the WT in mammalian cells in vitro. The 271A mutant did not increase viral pathogenicity significantly in mice compared to that of the 627K mutant, but it did enhance the lung virus titer. Also, cell infiltration was more evident in lungs of 271A-infected mice than in those of the WT. Interestingly, the avian-derived PB2 of the 2009 pandemic H1N1 influenza virus has 271A. The characterization of the polymerase activity of A/California/04/2009 (H1N1) and corresponding PB2 mutants indicates that the high polymerase activity of the pandemic strain in mammalian cells is, in part, dependent on 271A. Our results clearly indicate the contribution of PB2 amino acid 271 to enhanced polymerase activity and viral growth in mammalian hosts.

Prevalence, genetics, and transmissibility in ferrets of Eurasian avian-like H1N1 swine influenza viruses

“…Several amino acid changes in PB2 are important for the replication or virulence of influenza viruses in mammals (22)(23)(24)(25)(26)(27)(28)(29)(30). A detailed comparison of the amino acid differences among the EAH1N1 SIVs indicated that all of these viruses have mammalian-adapting mutations in their PB2 genes (SI Appendix, Table S3).…”

Biochemical Impact of the Host Adaptation-associated PB2 E627K Mutation on the Temperature-dependent RNA Synthesis Kinetics of Influenza A Virus Polymerase Complex

“…For this test we employed the H3N2 Nanchang strain, which is a well characterized avian strain cloned directly from birds and has never been exposed to humans (21). This avian influenza strain replicates efficiently at high temperatures in avian cells and contains E at the 627 position in PB2 (27). In addition, we introduced the human E627K mutation to the PB2 subunit of the avian Nanchang Pol complex.…”

Adaptive mutations in NEP compensate for defective H5N1 RNA replication in cultured human cells

“…Moreover, the 2009 H1N1 pandemic virus also harbours PB2-E627, but additionally possesses a lysine at position 591 (PB2-E591K), which is able to partially, but not completely, compensate for the lack of the PB2-E627K mutation 10,11 , suggesting that other adaptive factors may account for the efficient replication of these viruses in humans. Other mutations in PB2, known to have a role in host adaptation, are the exchange of aspartic acid with asparagine at position 701 (PB2-D701N), which can confer high pathogenicity to avian H5N1 and H7N7 viruses in mice 4,12 and the substitution of threonine with alanine at position 271 (PB2-T271A), which has been suggested to enhance polymerase activity in mammalian cells 13 . Furthermore, adaptation to human importin α might have a crucial role in interspecies transmission and pathogenicity 15,16 .…”