Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1987 Nov;84(22):8140–8144. doi: 10.1073/pnas.84.22.8140

Genomic RNAs of influenza viruses are held in a circular conformation in virions and in infected cells by a terminal panhandle.

M T Hsu 1, J D Parvin 1, S Gupta 1, M Krystal 1, P Palese 1
PMCID: PMC299494  PMID: 2446318

Abstract

The viral RNA segments in influenza virions were shown to be circular in conformation by using psoralen crosslinking methods. Electron microscopy of purified RNA following treatment of virus with the psoralen reagent 4'-aminomethyltrioxsalen (AMT) revealed circles with lengths corresponding to the individual segments. RNA blot analysis using polyacrylamide gels demonstrated that RNA from AMT-treated virus had a slowed migration, consistent with it being a single-stranded circle. Furthermore, nuclease S1 protection assays indicated that the termini of the RNA segments form an approximately 15-base-pair-long panhandle. This structure is consistent with the partial sequence complementarity that has been observed for the termini of all influenza virus RNAs. By RNA blot analysis, circular structures of viral sense RNA were also found in influenza virus-infected cells at early and late time points. The circular RNA was the predominant species at the time when the major transcription product is message RNA. This finding and the observation that the termination signal for mRNA synthesis directly abuts the panhandle suggest that a panhandle in the template viral RNA is a cis regulatory signal promoting the synthesis of mRNA instead of plus-sense template. Also, since the panhandle is present in high concentration in virions, we suggest that it is required for packaging and that the input RNA after infection is in the proper conformation for synthesis of primary transcripts.

Full text

PDF

Images in this article

8141Image
on p.8141
8142Image
on p.8142
8142Image
on p.8142
8142Image
on p.8142
8142Image
on p.8142
8142Image
on p.8142
8143Image
on p.8143

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Auperin D. D., Romanowski V., Galinski M., Bishop D. H. Sequencing studies of pichinde arenavirus S RNA indicate a novel coding strategy, an ambisense viral S RNA. J Virol. 1984 Dec;52(3):897–904. doi: 10.1128/jvi.52.3.897-904.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Braam J., Ulmanen I., Krug R. M. Molecular model of a eucaryotic transcription complex: functions and movements of influenza P proteins during capped RNA-primed transcription. Cell. 1983 Sep;34(2):609–618. doi: 10.1016/0092-8674(83)90393-8. [DOI] [PubMed] [Google Scholar]
  3. Buonagurio D. A., Nakada S., Parvin J. D., Krystal M., Palese P., Fitch W. M. Evolution of human influenza A viruses over 50 years: rapid, uniform rate of change in NS gene. Science. 1986 May 23;232(4753):980–982. doi: 10.1126/science.2939560. [DOI] [PubMed] [Google Scholar]
  4. Cabradilla C. D., Jr, Holloway B. P., Obijeski J. F. Molecular cloning and sequencing of the La Crosse virus S RNA. Virology. 1983 Jul 30;128(2):463–468. doi: 10.1016/0042-6822(83)90271-4. [DOI] [PubMed] [Google Scholar]
  5. Compans R. W., Content J., Duesberg P. H. Structure of the ribonucleoprotein of influenza virus. J Virol. 1972 Oct;10(4):795–800. doi: 10.1128/jvi.10.4.795-800.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Desselberger U., Racaniello V. R., Zazra J. J., Palese P. The 3' and 5'-terminal sequences of influenza A, B and C virus RNA segments are highly conserved and show partial inverted complementarity. Gene. 1980 Feb;8(3):315–328. doi: 10.1016/0378-1119(80)90007-4. [DOI] [PubMed] [Google Scholar]
  7. Duesberg P. H. Distinct subunits of the ribonucleoprotein of influenza virus. J Mol Biol. 1969 Jun 28;42(3):485–499. doi: 10.1016/0022-2836(69)90237-x. [DOI] [PubMed] [Google Scholar]
  8. Hay A. J., Skehel J. J. Characterization of influenza virus RNA transcripts synthesized in vitro. J Gen Virol. 1979 Sep;44(3):599–608. doi: 10.1099/0022-1317-44-3-599. [DOI] [PubMed] [Google Scholar]
  9. Hsu M. T. Electron microscopic evidence for the cruciform structure in intracellular SV40 DNA. Virology. 1985 Jun;143(2):617–621. doi: 10.1016/0042-6822(85)90400-3. [DOI] [PubMed] [Google Scholar]
  10. Jennings P. A., Finch J. T., Winter G., Robertson J. S. Does the higher order structure of the influenza virus ribonucleoprotein guide sequence rearrangements in influenza viral RNA? Cell. 1983 Sep;34(2):619–627. doi: 10.1016/0092-8674(83)90394-x. [DOI] [PubMed] [Google Scholar]
  11. Keene J. D., Schubert M., Lazzarini R. A. Terminal sequences of vesicular stomatitis virus RNA are both complementary and conserved. J Virol. 1979 Oct;32(1):167–174. doi: 10.1128/jvi.32.1.167-174.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kos A., Dijkema R., Arnberg A. C., van der Meide P. H., Schellekens H. The hepatitis delta (delta) virus possesses a circular RNA. Nature. 1986 Oct 9;323(6088):558–560. doi: 10.1038/323558a0. [DOI] [PubMed] [Google Scholar]
  13. Melton D. A., Krieg P. A., Rebagliati M. R., Maniatis T., Zinn K., Green M. R. Efficient in vitro synthesis of biologically active RNA and RNA hybridization probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res. 1984 Sep 25;12(18):7035–7056. doi: 10.1093/nar/12.18.7035. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Nayak D. P., Sivasubramanian N., Davis A. R., Cortini R., Sung J. Complete sequence analyses show that two defective interfering influenza viral RNAs contain a single internal deletion of a polymerase gene. Proc Natl Acad Sci U S A. 1982 Apr;79(7):2216–2220. doi: 10.1073/pnas.79.7.2216. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Obijeski J. F., McCauley J., Skehel J. J. Nucleotide sequences at the terminal of La Crosse virus RNAs. Nucleic Acids Res. 1980 Jun 11;8(11):2431–2438. doi: 10.1093/nar/8.11.2431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Palese P., Schulman J. L. Differences in RNA patterns of influenza A viruses. J Virol. 1976 Mar;17(3):876–884. doi: 10.1128/jvi.17.3.876-884.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Palmer E. L., Obijeski J. F., Webb P. A., Johnson K. M. The circular, segmented nucleocapsid of an arenavirus-Tacaribe virus. J Gen Virol. 1977 Sep;36(3):541–545. doi: 10.1099/0022-1317-36-3-541. [DOI] [PubMed] [Google Scholar]
  18. Parvin J. D., Moscona A., Pan W. T., Leider J. M., Palese P. Measurement of the mutation rates of animal viruses: influenza A virus and poliovirus type 1. J Virol. 1986 Aug;59(2):377–383. doi: 10.1128/jvi.59.2.377-383.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Pettersson R. F., von Bonsdorff C. H. Ribonucleoproteins of Uukuniemi virus are circular. J Virol. 1975 Feb;15(2):386–392. doi: 10.1128/jvi.15.2.386-392.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Pons M. W., Rochovansky O. M. Ultraviolet inactivation of influenza virus RNA in vitro and vivo. Virology. 1979 Aug;97(1):183–189. doi: 10.1016/0042-6822(79)90385-4. [DOI] [PubMed] [Google Scholar]
  21. Robertson J. S. 5' and 3' terminal nucleotide sequences of the RNA genome segments of influenza virus. Nucleic Acids Res. 1979 Aug 24;6(12):3745–3757. doi: 10.1093/nar/6.12.3745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Robertson J. S., Schubert M., Lazzarini R. A. Polyadenylation sites for influenza virus mRNA. J Virol. 1981 Apr;38(1):157–163. doi: 10.1128/jvi.38.1.157-163.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Schenborn E. T., Mierendorf R. C., Jr A novel transcription property of SP6 and T7 RNA polymerases: dependence on template structure. Nucleic Acids Res. 1985 Sep 11;13(17):6223–6236. doi: 10.1093/nar/13.17.6223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Skehel J. J., Hay A. J. Nucleotide sequences at the 5' termini of influenza virus RNAs and their transcripts. Nucleic Acids Res. 1978 Apr;5(4):1207–1219. doi: 10.1093/nar/5.4.1207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Smith F. I., Parvin J. D., Palese P. Detection of single base substitutions in influenza virus RNA molecules by denaturing gradient gel electrophoresis of RNA-RNA or DNA-RNA heteroduplexes. Virology. 1986 Apr 15;150(1):55–64. doi: 10.1016/0042-6822(86)90265-5. [DOI] [PubMed] [Google Scholar]
  26. Stoeckle M. Y., Shaw M. W., Choppin P. W. Segment-specific and common nucleotide sequences in the noncoding regions of influenza B virus genome RNAs. Proc Natl Acad Sci U S A. 1987 May;84(9):2703–2707. doi: 10.1073/pnas.84.9.2703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Vezza A. C., Clewley J. P., Gard G. P., Abraham N. Z., Compans R. W., Bishop D. H. Virion RNA species of the arenaviruses Pichinde, Tacaribe, and Tamiami. J Virol. 1978 May;26(2):485–497. doi: 10.1128/jvi.26.2.485-497.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES