Article | Published:

Structural insight into cap-snatching and RNA synthesis by influenza polymerase

Nature volume 516, pages 361366 (18 December 2014) | Download Citation

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Abstract

Influenza virus polymerase uses a capped primer, derived by ‘cap-snatching’ from host pre-messenger RNA, to transcribe its RNA genome into mRNA and a stuttering mechanism to generate the poly(A) tail. By contrast, genome replication is unprimed and generates exact full-length copies of the template. Here we use crystal structures of bat influenza A and human influenza B polymerases (FluA and FluB), bound to the viral RNA promoter, to give mechanistic insight into these distinct processes. In the FluA structure, a loop analogous to the priming loop of flavivirus polymerases suggests that influenza could initiate unprimed template replication by a similar mechanism. Comparing the FluA and FluB structures suggests that cap-snatching involves in situ rotation of the PB2 cap-binding domain to direct the capped primer first towards the endonuclease and then into the polymerase active site. The polymerase probably undergoes considerable conformational changes to convert the observed pre-initiation state into the active initiation and elongation states.

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Accessions

Primary accessions

Data deposits

Structure factors and co-ordinates have been deposited in the Protein Data Bank (PDB) under the accessions 4WSA (FluB form 1) and 4WRT (FluB form 2).

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Acknowledgements

We thank the staff of the European Molecular Biology Laboratory (EMBL) eukaryotic expression and high-throughput crystallization facilities within the Partnership for Structural Biology (PSB) and members of the ESRF-EMBL Joint Structural Biology Group for help on European Synchrotron Radiation Facility (ESRF) beamlines. The work was supported by ERC Advanced Grant V-RNA (322586) and EU Grant FLU-PHARM (259751) to S.C. and partially by a Roche Postdoc Fellowship to S.R.

Author information

Author notes

    • Stefan Reich
    • , Delphine Guilligay
    •  & Alexander Pflug

    These authors contributed equally to this work.

Affiliations

  1. European Molecular Biology Laboratory, Grenoble Outstation, 71 Avenue des Martyrs, CS 90181, 38042 Grenoble Cedex 9, France

    • Stefan Reich
    • , Delphine Guilligay
    • , Alexander Pflug
    • , Hélène Malet
    • , Imre Berger
    • , Darren Hart
    • , Thomas Lunardi
    • , Max Nanao
    •  & Stephen Cusack
  2. University Grenoble Alpes-Centre National de la Recherche Scientifique-EMBL Unit of Virus Host-Cell Interactions, 71 Avenue des Martyrs, CS 90181, 38042 Grenoble Cedex 9, France

    • Stefan Reich
    • , Delphine Guilligay
    • , Alexander Pflug
    • , Hélène Malet
    • , Imre Berger
    • , Thibaut Crépin
    • , Darren Hart
    • , Thomas Lunardi
    • , Max Nanao
    • , Rob W. H. Ruigrok
    •  & Stephen Cusack

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Contributions

S.R., D.G. and T.L. did protein expression, purification, crystallization and activity assays. A.P. did crystallographic analysis. H.M. did electron microscopy and fitting to the mini-RNP electron microscopy map. M.N. calculated the first interpretable FluB polymerase electron density map. Using the polyprotein vector designed and provided by I.B., and with the help of D.H., S.C. designed the FluB polymerase construct. T.C., D.H., R.R. and S.C. have long-collaborated on studies of influenza polymerase. S.C. supervised the project, collected data, did crystallographic analysis and wrote the paper with input from S.R., D.G., A.P., H.M. and M.N.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Stephen Cusack.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Information

    This file contains Supplementary Figure 1, Supplementary Text and Supplementary References.

Videos

  1. 1.

    Simulation of the putative rotation of the cap-binding domain (orange) by morphing between the domain positions seen in the FluA and FluB crystal structures.

    The yellow spheres in the cap-binding domain correspond to the bound cap-analogue m7GTP.

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DOI

https://doi.org/10.1038/nature14009

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