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. 2001 Jul;7(7):979–989. doi: 10.1017/s1355838201002424

The packaging signal of influenza viral RNA molecules.

S Tchatalbachev 1, R Flick 1, G Hobom 1
PMCID: PMC1370150  PMID: 11453070

Abstract

The packaging signal present in influenza viral RNA molecules is shown not to constitute a separate structural element, but to reside within the 5'-bulged promoter structure, as caused by the central unpaired residue A10 in its 5' branch. Upon insertion of two uridine residues in the 3' branch opposite A10, the minus-strand viral RNA (vRNA) promoter is converted into a 3'-bulged structure, whereas the plus-strand cRNA promoter instead adopts the 5'-bulged conformation. In this promoter variant it is exclusively the cRNA that is found packaged in the progeny virions. Upon insertion of only a single uridine nucleotide opposite 5'A10, the two debulged structures of the vRNA and cRNA promoters are rendered identical, and both vRNA and cRNA molecules are packaged indiscriminately, in a 1:1 ratio, but at lower rates. We propose that the binding interactions of viral polymerase with either of the two differently bulged vRNA and cRNA promoter structures result in two different conformations of the enzyme protein. Only the 5' bulged RNA-associated polymerase conformation appears to be recognized for nuclear export, which depends on nuclear matrix protein M1 and nonstructural protein NS2. And the respective wild-type vRNP- or insertion mutant cRNP complex is observed to enter the cytoplasm and hence is included in the viral encapsidation process, which takes place at the plasma membrane.

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Selected References

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  1. Adam S. A., Marr R. S., Gerace L. Nuclear protein import in permeabilized mammalian cells requires soluble cytoplasmic factors. J Cell Biol. 1990 Sep;111(3):807–816. doi: 10.1083/jcb.111.3.807. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ali A., Avalos R. T., Ponimaskin E., Nayak D. P. Influenza virus assembly: effect of influenza virus glycoproteins on the membrane association of M1 protein. J Virol. 2000 Sep;74(18):8709–8719. doi: 10.1128/jvi.74.18.8709-8719.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bui M., Wills E. G., Helenius A., Whittaker G. R. Role of the influenza virus M1 protein in nuclear export of viral ribonucleoproteins. J Virol. 2000 Feb;74(4):1781–1786. doi: 10.1128/jvi.74.4.1781-1786.2000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Flick R., Hobom G. Interaction of influenza virus polymerase with viral RNA in the 'corkscrew' conformation. J Gen Virol. 1999 Oct;80(Pt 10):2565–2572. doi: 10.1099/0022-1317-80-10-2565. [DOI] [PubMed] [Google Scholar]
  5. Flick R., Neumann G., Hoffmann E., Neumeier E., Hobom G. Promoter elements in the influenza vRNA terminal structure. RNA. 1996 Oct;2(10):1046–1057. [PMC free article] [PubMed] [Google Scholar]
  6. Fosmire J. A., Hwang K., Makino S. Identification and characterization of a coronavirus packaging signal. J Virol. 1992 Jun;66(6):3522–3530. doi: 10.1128/jvi.66.6.3522-3530.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Geigenmüller-Gnirke U., Nitschko H., Schlesinger S. Deletion analysis of the capsid protein of Sindbis virus: identification of the RNA binding region. J Virol. 1993 Mar;67(3):1620–1626. doi: 10.1128/jvi.67.3.1620-1626.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. González S., Ortín J. Distinct regions of influenza virus PB1 polymerase subunit recognize vRNA and cRNA templates. EMBO J. 1999 Jul 1;18(13):3767–3775. doi: 10.1093/emboj/18.13.3767. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hsu M. T., Parvin J. D., Gupta S., Krystal M., Palese P. Genomic RNAs of influenza viruses are held in a circular conformation in virions and in infected cells by a terminal panhandle. Proc Natl Acad Sci U S A. 1987 Nov;84(22):8140–8144. doi: 10.1073/pnas.84.22.8140. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ippolito J. A., Steitz T. A. The structure of the HIV-1 RRE high affinity rev binding site at 1.6 A resolution. J Mol Biol. 2000 Jan 28;295(4):711–717. doi: 10.1006/jmbi.1999.3405. [DOI] [PubMed] [Google Scholar]
  12. Jin H., Leser G. P., Zhang J., Lamb R. A. Influenza virus hemagglutinin and neuraminidase cytoplasmic tails control particle shape. EMBO J. 1997 Mar 17;16(6):1236–1247. doi: 10.1093/emboj/16.6.1236. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Johnson A. S., Couto C. G., Weghorst C. M. Mutation of the p53 tumor suppressor gene in spontaneously occurring osteosarcomas of the dog. Carcinogenesis. 1998 Jan;19(1):213–217. doi: 10.1093/carcin/19.1.213. [DOI] [PubMed] [Google Scholar]
  14. Li M. L., Ramirez B. C., Krug R. M. 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. 1998 Oct 1;17(19):5844–5852. doi: 10.1093/emboj/17.19.5844. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Martin K., Helenius A. Nuclear transport of influenza virus ribonucleoproteins: the viral matrix protein (M1) promotes export and inhibits import. Cell. 1991 Oct 4;67(1):117–130. doi: 10.1016/0092-8674(91)90576-k. [DOI] [PubMed] [Google Scholar]
  16. McDougall J. K., Masse T. H., Galloway D. A. Location and cloning of the herpes simplex virus type 2 thymidine kinase gene. J Virol. 1980 Mar;33(3):1221–1224. doi: 10.1128/jvi.33.3.1221-1224.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Mitnaul L. J., Castrucci M. R., Murti K. G., Kawaoka Y. The cytoplasmic tail of influenza A virus neuraminidase (NA) affects NA incorporation into virions, virion morphology, and virulence in mice but is not essential for virus replication. J Virol. 1996 Feb;70(2):873–879. doi: 10.1128/jvi.70.2.873-879.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Murti K. G., Brown P. S., Bean W. J., Jr, Webster R. G. Composition of the helical internal components of influenza virus as revealed by immunogold labeling/electron microscopy. Virology. 1992 Jan;186(1):294–299. doi: 10.1016/0042-6822(92)90084-3. [DOI] [PubMed] [Google Scholar]
  19. Neumann G., Hobom G. Mutational analysis of influenza virus promoter elements in vivo. J Gen Virol. 1995 Jul;76(Pt 7):1709–1717. doi: 10.1099/0022-1317-76-7-1709. [DOI] [PubMed] [Google Scholar]
  20. Neumann G., Hughes M. T., Kawaoka Y. Influenza A virus NS2 protein mediates vRNP nuclear export through NES-independent interaction with hCRM1. EMBO J. 2000 Dec 15;19(24):6751–6758. doi: 10.1093/emboj/19.24.6751. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Neumann G., Zobel A., Hobom G. RNA polymerase I-mediated expression of influenza viral RNA molecules. Virology. 1994 Jul;202(1):477–479. doi: 10.1006/viro.1994.1365. [DOI] [PubMed] [Google Scholar]
  22. O'Neill R. E., Talon J., Palese P. The influenza virus NEP (NS2 protein) mediates the nuclear export of viral ribonucleoproteins. EMBO J. 1998 Jan 2;17(1):288–296. doi: 10.1093/emboj/17.1.288. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Pollack J. R., Ganem D. An RNA stem-loop structure directs hepatitis B virus genomic RNA encapsidation. J Virol. 1993 Jun;67(6):3254–3263. doi: 10.1128/jvi.67.6.3254-3263.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Poon L. L., Pritlove D. C., Fodor E., Brownlee G. G. Direct evidence that the poly(A) tail of influenza A virus mRNA is synthesized by reiterative copying of a U track in the virion RNA template. J Virol. 1999 Apr;73(4):3473–3476. doi: 10.1128/jvi.73.4.3473-3476.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Popov S., Rexach M., Zybarth G., Reiling N., Lee M. A., Ratner L., Lane C. M., Moore M. S., Blobel G., Bukrinsky M. Viral protein R regulates nuclear import of the HIV-1 pre-integration complex. EMBO J. 1998 Feb 16;17(4):909–917. doi: 10.1093/emboj/17.4.909. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Ruigrok R. W., Barge A., Durrer P., Brunner J., Ma K., Whittaker G. R. Membrane interaction of influenza virus M1 protein. Virology. 2000 Feb 15;267(2):289–298. doi: 10.1006/viro.1999.0134. [DOI] [PubMed] [Google Scholar]
  27. Sha B., Luo M. Structure of a bifunctional membrane-RNA binding protein, influenza virus matrix protein M1. Nat Struct Biol. 1997 Mar;4(3):239–244. doi: 10.1038/nsb0397-239. [DOI] [PubMed] [Google Scholar]
  28. Shapiro G. I., Gurney T., Jr, Krug R. M. Influenza virus gene expression: control mechanisms at early and late times of infection and nuclear-cytoplasmic transport of virus-specific RNAs. J Virol. 1987 Mar;61(3):764–773. doi: 10.1128/jvi.61.3.764-773.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Tiley L. S., Malim M. H., Tewary H. K., Stockley P. G., Cullen B. R. Identification of a high-affinity RNA-binding site for the human immunodeficiency virus type 1 Rev protein. Proc Natl Acad Sci U S A. 1992 Jan 15;89(2):758–762. doi: 10.1073/pnas.89.2.758. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wakefield L., Brownlee G. G. RNA-binding properties of influenza A virus matrix protein M1. Nucleic Acids Res. 1989 Nov 11;17(21):8569–8580. doi: 10.1093/nar/17.21.8569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Watanabe K., Handa H., Mizumoto K., Nagata K. Mechanism for inhibition of influenza virus RNA polymerase activity by matrix protein. J Virol. 1996 Jan;70(1):241–247. doi: 10.1128/jvi.70.1.241-247.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Whittaker G., Kemler I., Helenius A. Hyperphosphorylation of mutant influenza virus matrix protein, M1, causes its retention in the nucleus. J Virol. 1995 Jan;69(1):439–445. doi: 10.1128/jvi.69.1.439-445.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Ye Z. P., Pal R., Fox J. W., Wagner R. R. Functional and antigenic domains of the matrix (M1) protein of influenza A virus. J Virol. 1987 Feb;61(2):239–246. doi: 10.1128/jvi.61.2.239-246.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Zhang J., Lamb R. A. Characterization of the membrane association of the influenza virus matrix protein in living cells. Virology. 1996 Nov 15;225(2):255–266. doi: 10.1006/viro.1996.0599. [DOI] [PubMed] [Google Scholar]
  35. Zhao H., Ekström M., Garoff H. The M1 and NP proteins of influenza A virus form homo- but not heterooligomeric complexes when coexpressed in BHK-21 cells. J Gen Virol. 1998 Oct;79(Pt 10):2435–2446. doi: 10.1099/0022-1317-79-10-2435. [DOI] [PubMed] [Google Scholar]
  36. Zheng H., Palese P., García-Sastre A. Nonconserved nucleotides at the 3' and 5' ends of an influenza A virus RNA play an important role in viral RNA replication. Virology. 1996 Mar 1;217(1):242–251. doi: 10.1006/viro.1996.0111. [DOI] [PubMed] [Google Scholar]
  37. Zhou Y., König M., Hobom G., Neumeier E. Membrane-anchored incorporation of a foreign protein in recombinant influenza virions. Virology. 1998 Jun 20;246(1):83–94. doi: 10.1006/viro.1998.9169. [DOI] [PubMed] [Google Scholar]
  38. Zobel A., Neumann G., Hobom G. RNA polymerase I catalysed transcription of insert viral cDNA. Nucleic Acids Res. 1993 Aug 11;21(16):3607–3614. doi: 10.1093/nar/21.16.3607. [DOI] [PMC free article] [PubMed] [Google Scholar]

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