1887

Abstract

The influenza A virus RNA-dependent RNA polymerase consists of three subunits PB1, PB2 and PA. The 5′ and 3′ terminal sequences of the viral RNA (vRNA) form the viral promoter and are bound by the PB1 subunit. The putative promoter-binding sites of the PB1 subunit have been mapped in previous studies but with contradictory results. The aim of the current study was to investigate the function of two evolutionary conserved regions in PB1 – from aa 233 to 249 and 269 to 281, which lie immediately N- and C-terminal, respectively, of a previously proposed binding site for the 3′ end of the vRNA promoter. The previously proposed binding site extended from aa 249 to 256 and centred on two phenylalanine residues (F251 and F254). However, the fact that F251 is required for polymerase activity was not confirmed here. Instead, it was proposed that the 233–249 region contains a new 5′ vRNA promoter-binding site, and arginine residues crucial for this activity were characterized. However, residues 269–281 were unlikely to be directly involved in promoter binding. These results are discussed in relation to the previous studies and a new model for vRNA promoter binding to the influenza RNA polymerase is presented.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.81453-0
2006-03-01
2019-11-14
Loading full text...

Full text loading...

/deliver/fulltext/jgv/87/3/679.html?itemId=/content/journal/jgv/10.1099/vir.0.81453-0&mimeType=html&fmt=ahah

References

  1. Area, E., Martin-Benito, J., Gastaminza, P., Torreira, E., Valpuesta, J. M., Carrascosa, J. L. & Ortin, J. ( 2004; ). 3D structure of the influenza virus polymerase complex: localization of subunit domains. Proc Natl Acad Sci U S A 101, 308–313.[CrossRef]
    [Google Scholar]
  2. Brownlee, G. G. & Sharps, J. L. ( 2002; ). The RNA polymerase of influenza A virus is stabilized by interaction with its viral RNA promoter. J Virol 76, 7103–7113.[CrossRef]
    [Google Scholar]
  3. Calnan, B. J., Biancalana, S., Hudson, D. & Frankel, A. D. ( 1991; ). Analysis of arginine-rich peptides from the HIV Tat protein reveals unusual features of RNA-protein recognition. Genes Dev 5, 201–210.[CrossRef]
    [Google Scholar]
  4. Carr, S. M., Carnero, E., Garcia-Sastre, A., Brownlee, G. G. & Fodor, E. ( 2005; ). Characterization of a mitochondrial-targeting signal in the PB2 protein of influenza viruses. Virology (in press).
    [Google Scholar]
  5. Crescenzo-Chaigne, B., van der Werf, S. & Naffakh, N. ( 2002; ). Differential effect of nucleotide substitutions in the 3′ arm of the influenza A virus vRNA promoter on transcription/replication by avian and human polymerase complexes is related to the nature of PB2 amino acid 627. Virology 303, 240–252.[CrossRef]
    [Google Scholar]
  6. Crow, M., Deng, T., Addley, M. & Brownlee, G. G. ( 2004; ). Mutational analysis of the influenza virus cRNA promoter and identification of nucleotides critical for replication. J Virol 78, 6263–6270.[CrossRef]
    [Google Scholar]
  7. Deng, T., Sharps, J., Fodor, E. & Brownlee, G. G. ( 2005; ). In vitro assembly of PB2 with a PB1-PA dimer supports a new model of assembly of influenza A virus polymerase subunits into a functional trimeric complex. J Virol 79, 8669–8674.[CrossRef]
    [Google Scholar]
  8. Fechter, P., Mingay, L., Sharps, J., Chambers, A., Fodor, E. & Brownlee, G. G. ( 2003; ). Two aromatic residues in the PB2 subunit of influenza A RNA polymerase are crucial for cap binding. J Biol Chem 278, 20381–20388.[CrossRef]
    [Google Scholar]
  9. Fodor, E. & Brownlee, G. G. ( 2002; ). Influenza virus replication. In Influenza, pp. 1–30. Edited by C. W. Potter. Amsterdam: Elsevier Science.
  10. Fodor, E., Seong, B. L. & Brownlee, G. G. ( 1993; ). Photochemical cross-linking of influenza A polymerase to its virion RNA promoter defines a polymerase binding site at residue 9 to 12 of the promoter. J Gen Virol 74, 1327–1333.[CrossRef]
    [Google Scholar]
  11. Fodor, E., Pritlove, D. C. & Brownlee, G. G. ( 1994; ). The influenza virus panhandle is involved in the initiation of transcription. J Virol 68, 4092–4096.
    [Google Scholar]
  12. Fodor, E., Devenish, L., Engelhardt, O. G., Palese, P., Brownlee, G. G. & García-Sastre, A. ( 1999; ). Rescue of influenza A virus from recombinant DNA. J Virol 73, 9679–9682.
    [Google Scholar]
  13. Fodor, E., Crow, M., Mingay, L. J., Deng, T., Sharps, J., Fechter, P. & Brownlee, G. G. ( 2002; ). A single amino acid mutation in the PA subunit of the influenza virus RNA polymerase inhibits endonucleolytic cleavage of capped RNAs. J Virol 76, 8989–9001.[CrossRef]
    [Google Scholar]
  14. Fodor, E., Mingay, L. J., Crow, M., Deng, T. & Brownlee, G. G. ( 2003; ). A single amino acid mutation in the PA subunit of the influenza virus RNA polymerase promotes the generation of defective interfering RNAs. J Virol 77, 5017–5020.[CrossRef]
    [Google Scholar]
  15. Gonzalez, S. & Ortin, J. ( 1999; ). Characterization of influenza virus PB1 protein binding to viral RNA: two separate regions of the protein contribute to the interaction domain. J Virol 73, 631–637.
    [Google Scholar]
  16. Huarte, M., Falcon, A., Nakaya, Y., Ortin, J., Garcia-Sastre, A. & Nieto, A. ( 2003; ). Threonine 157 of influenza virus PA polymerase subunit modulates RNA replication in infectious viruses. J Virol 77, 6007–6013.[CrossRef]
    [Google Scholar]
  17. Kawaguchi, A., Naito, T. & Nagata, K. ( 2005; ). Involvement of influenza virus PA subunit in assembly of functional RNA polymerase complexes. J Virol 79, 732–744.[CrossRef]
    [Google Scholar]
  18. Lamb, R. A. & Krug, R. M. ( 2001; ). Orthomyxoviridae: the viruses and their replication. In Fields Virology, pp. 1487–1579. Edited by P. M. Howley. Philadelphia: Lippincott Williams & Wilkins.
  19. Leahy, M. B., Pritlove, D. C., Poon, L. L. & Brownlee, G. G. ( 2001; ). Mutagenic analysis of the 5′ arm of the influenza A virus virion RNA promoter defines the sequence requirements for endonuclease activity. J Virol 75, 134–142.[CrossRef]
    [Google Scholar]
  20. Legault, P., Li, J., Mogridge, J., Kay, L. E. & Greenblatt, J. ( 1998; ). NMR structure of the bacteriophage lambda N peptide/boxB RNA complex: recognition of a GNRA fold by an arginine-rich motif. Cell 93, 289–299.[CrossRef]
    [Google Scholar]
  21. Li, M. L., Ramirez, B. C. & Krug, R. M. ( 1998; ). RNA-dependent activation of primer RNA production by influenza virus polymerase: different regions of the same protein subunit constitute the two required RNA-binding sites. EMBO J 17, 5844–5852.[CrossRef]
    [Google Scholar]
  22. Neumann, G., Brownlee, G. G., Fodor, E. & Kawaoka, Y. ( 2004; ). Orthomyxovirus replication, transcription, and polyadenylation. Curr Top Microbiol Immunol 283, 121–143.
    [Google Scholar]
  23. Portela, A., Zurcher, T., Nieto, A. & Ortin, J. ( 1999; ). Replication of orthomyxoviruses. Adv Virus Res 54, 319–348.
    [Google Scholar]
  24. Pritlove, D. C., Fodor, E., Seong, B. L. & Brownlee, G. G. ( 1995; ). In vitro transcription and polymerase binding studies of the termini of influenza A virus complementary RNA: evidence for a cRNA panhandle. J Gen Virol 76, 2205–2213.[CrossRef]
    [Google Scholar]
  25. Puig, O., Caspary, F., Rigaut, G., Rutz, B., Bouveret, E., Bragado-Nilsson, E., Wilm, M. & Seraphin, B. ( 2001; ). The tandem affinity purification (TAP) method: a general procedure of protein complex purification. Methods 24, 218–229.[CrossRef]
    [Google Scholar]
  26. Rao, P., Yuan, W. & Krug, R. M. ( 2003; ). Crucial role of CA cleavage sites in the cap-snatching mechanism for initiating viral mRNA synthesis. EMBO J 22, 1188–1198.[CrossRef]
    [Google Scholar]
  27. Subbarao, K., Chen, H., Swayne, D. & 8 other authors ( 2003; ). Evaluation of a genetically modified reassortant H5N1 influenza A virus vaccine candidate generated by plasmid-based reverse genetics. Virology 305, 192–200.[CrossRef]
    [Google Scholar]
  28. Tiley, L. S., Hagen, M., Matthews, J. T. & Krystal, M. ( 1994; ). Sequence-specific binding of the influenza virus RNA polymerase to sequences located at the 5′ ends of the viral RNAs. J Virol 68, 5108–5116.
    [Google Scholar]
  29. Weiss, M. A. & Narayana, N. ( 1998; ). RNA recognition by arginine-rich peptide motifs. Biopolymers 48, 167–180.[CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.81453-0
Loading
/content/journal/jgv/10.1099/vir.0.81453-0
Loading

Data & Media loading...

Supplements

vol. , part 3, pp. 679 - 688

Primer extension data of steady-state levels of mRNA and cRNA isolated from cells expressing influenza A virus polymerase and vRNA-like CAT-RNA [PDF](31 KB)



PDF

Most Cited This Month

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error