Capped RNA primer binding to influenza polymerase and implications for the mechanism of cap-binding inhibitors

Nucleic Acids Res. 2018 Jan 25;46(2):956-971. doi: 10.1093/nar/gkx1210.


Influenza polymerase uses short capped primers snatched from nascent Pol II transcripts to initiate transcription of viral mRNAs. Here we describe crystal structures of influenza A and B polymerase bound to a capped primer in a configuration consistent with transcription initiation ('priming state') and show by functional assays that conserved residues from both the PB2 midlink and cap-binding domains are important for positioning the capped RNA. In particular, mutation of PB2 Arg264, which interacts with the triphosphate linkage in the cap, significantly and specifically decreases cap-dependent transcription. We also compare the configuration of the midlink and cap-binding domains in the priming state with their very different relative arrangement (called the 'apo' state) in structures where the potent cap-binding inhibitor VX-787, or a close analogue, is bound. In the 'apo' state the inhibitor makes additional interactions to the midlink domain that increases its affinity beyond that to the cap-binding domain alone. The comparison suggests that the mechanism of resistance of certain mutations that allow virus to escape from VX-787, notably PB2 N510T, can only be rationalized if VX-787 has a dual mode of action, direct inhibition of capped RNA binding as well as stabilization of the transcriptionally inactive 'apo' state.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Binding Sites / genetics
  • Crystallography, X-Ray
  • HEK293 Cells
  • Humans
  • Indoles / metabolism
  • Indoles / pharmacology
  • Influenza A virus / enzymology
  • Protein Binding
  • Pyridines
  • Pyrimidines
  • Pyrroles
  • RNA / chemistry
  • RNA / genetics
  • RNA / metabolism*
  • RNA Cap Analogs / metabolism*
  • RNA Cap Analogs / pharmacology
  • RNA Caps / chemistry
  • RNA Caps / genetics
  • RNA Caps / metabolism*
  • RNA Polymerase II / chemistry
  • RNA Polymerase II / genetics
  • RNA Polymerase II / metabolism*
  • RNA, Messenger / chemistry
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Transcription, Genetic / drug effects
  • Viral Proteins / chemistry
  • Viral Proteins / genetics
  • Viral Proteins / metabolism*


  • Indoles
  • Pyridines
  • Pyrimidines
  • Pyrroles
  • RNA Cap Analogs
  • RNA Caps
  • RNA primers
  • RNA, Messenger
  • Viral Proteins
  • RNA
  • pimodivir
  • RNA Polymerase II