1887

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Volume 69, Issue 8

The Intracellular Distribution of Influenza Virus Matrix Protein and Nucleoprotein in Infected Cells and Their Relationship to Haemagglutinin in the Plasma Membrane Free

Abstract

Summary

Pre- and post-embedding immune electron microscopy techniques employing ferritin and large and small gold markers to detect cell surface and intracellular antigens respectively, have been combined in a study of influenza virus-infected cells. This has permitted, for the first time, the simultaneous detection of intracellular virus matrix protein (M), nucleoprotein (NP) and membrane haemagglutinin (HA). The technique facilitated an investigation of the possible physical interrelationship between these three proteins both in the infected cell, and on the infected cell membrane. Electron-dense bodies uniformly labelled by antibody to M protein were observed in the nucleus and cytoplasm. Similarly, NP was detected in both the nucleus and cytoplasm. Approximately 50% of the nuclear NP was located in close proximity to the M protein-containing dense bodies but mainly on the perimeter of the structures. A similar relationship of NP to the M-containing dense bodies was observed in the cytoplasm. M protein and NP were readily detected in sections of budding virions. Labelling of these proteins was also observed on the cytoplasmic face of the plasma membrane but the density of labelling only occasionally approached that of newly formed virions. These findings suggest that budding occurs very quickly after the internal proteins arrive at the plasma membrane. Double labelling experiments on the cell surface indicate that NP and HA behave as independent molecules and do not form tight complexes with each other.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): haemagglutinin , influenza virus and localization
© Society for General Microbiology 1988
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1988-08-01
2024-04-26
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References

  1. BÄCHI T., GERHARD W., LINDENMANN J., MÜHLETHALER K. 1969; Morphogenesis of influenza A virus in Ehrlich ascites tumor cells as revealed by thin-sectioning and freeze-etching. Journal of Virology 4:769–776
     [Google Scholar]
  2. BEESLEY J. E., CAMPBELL D. A. 1984; The use of ultrathin cryosections for localisation of influenza virus antigens in infected vero cell cultures. Histochemistry 80:497–501
     [Google Scholar]
  3. BENDAYAN M. 1984; Protein A-gold electron microscopic immunocytochemistry: methods, applications and limitations. Journal of Electron Microscopy Techniques 1:243–270
     [Google Scholar]
  4. BENDAYAN M., ORSTAVIK T. B. 1982; Immunocytochemical localization of kallikrein in rat exocrine pancreas. Journal of Histochemistry and Cytochemistry 30:58–66
     [Google Scholar]
  5. BINDER M., TOURMENTE S., ROTH J., RENAUD M., GEHRING W. J. 1986; In situ hybridization at the electron microscope level: localization of transcripts on ultrathin sections of Lowicryl K4M-embedded tissue using biotinylated probes and protein A-gold complexes. Journal of Cell Biology 102:1646–1653
     [Google Scholar]
  6. CHOPPIN P. W., KLENK H.-D., COMPANS R. W., CALIGUIRI L. A. 1971 The parainfluenza virus, SV5 and its relationship to the cell membrane. Perspectives in Virology 7127–158 Edited by Pollard M. New York & London: Academic Press;
     [Google Scholar]
  7. COMPANS R. W., DIMMOCK N. J. 1969; An electron microscopic study of single-cycle infection of chick embryo fibroblasts by influenza virus. Virology 39:449–515
     [Google Scholar]
  8. DAVEY J., COLMAN A., DIMMOCK N. J. 1985a; Location of influenza virus M, NP and NSI proteins in microinjected cells. Journal of General Virology 66:2319–2334
     [Google Scholar]
  9. DAVEY J., DIMMOCK N. J., COLMAN A. 1985b; Identification of the sequences responsible for the nuclear accumulation of the influenza virus nucleoprotein in Xenopus oocytes. Cell 40:667–675
     [Google Scholar]
  10. DE PÉTRIS S., RAFF M. C. 1972; Distribution of immunoglobulin on the surface of mouse lymphoid cells as determined by immunoferritin electron microscopy. Antibody-induced, temperature-dependent redistribution and its implications for membrane structure. European Journal of Immunology 2:523–535
     [Google Scholar]
  11. DOWDLE W. R., SCHILD G. C. 1975 Laboratory propagation of human influenza viruses, experimental host range and isolation from clinical material. The Influenza Viruses and Influenza243–268 Edited by Kilbourne E. D. New York & London: Academic Press;
     [Google Scholar]
  12. FLAWITH J. W. F., DIMMOCK N. J. 1979; Newly synthesized influenza virus proteins are transported from the nucleus. Journal of General Virology 45:527–531
     [Google Scholar]
  13. FRENS G. 1973; Controlled nucleation for the regulation of particle size in monodisperse gold solutions. Nature Physical Sciences 241:20–22
     [Google Scholar]
  14. GREGORIADES A. 1973; The membrane protein of influenza virus: extraction from virus and infected cells with acidic chloroform-methanol. Virology 54:369–383
     [Google Scholar]
  15. GRIFFITHS G., HOPPELER H. 1986; Quantitation of imunocytochemistry: correlation of immunogold labelling to absolute number of membrane antigens. Journal of Histochemistry and Cytochemistry 34:1389–1398
     [Google Scholar]
  16. HAY A. J. 1974; Studies on the formation of influenza virus envelope. Virology 60:394–418
     [Google Scholar]
  17. HAY A. J., SKEHEL J. J. 1975 Studies on the synthesis of influenza virus proteins. Negative Strand Viruses 2635–655 Edited by Mahy B. W. J., Barry R. D. New York & London: Academic Press;
     [Google Scholar]
  18. HORRISBERGER M., ROSSET J. 1977; Colloidal gold, a useful marker for transmission and scanning electron microscopy. Journal of Histochemistry and Cytochemistry 25:295–305
     [Google Scholar]
  19. HUDDLESTON J. A., BROWNLEE G. G. 1982; The sequence of the nucleoprotein gene of human influenza A virus, strain A/NT/60/68. Nucleic Acids Research 10:1029–1038
     [Google Scholar]
  20. KELLENBERGER E., DURRENBERGER M., WILLIGER W., CARLEMALM E., WURTZ M. 1987; The efficiency of immunolabel on Lowicryl sections compared to theoretical predictions. Journal of Histochemistry and Cytochemistry 35:959–969
     [Google Scholar]
  21. KRUG R. M., ETKIND P. R. 1973; Cytoplasmic and nuclear virus-specified proteins of influenza-infected MDCK cells. Virology 56:334–387
     [Google Scholar]
  22. LAZAROWITZ S. G., COMPANS R. W., CHOPPIN P. W. 1971; Influenza virus structural proteins and non-structural proteins in infected cells and their plasma membranes. Virology 46:830–843
     [Google Scholar]
  23. LENARD J., COMPANS R. W. 1974; The membrane of lipid-containing viruses. Biochimica et biophysica acta 344:51–94
     [Google Scholar]
  24. LOHMEYER J., TALENS L. T., KLENK H.-D. 1979; Biosynthesis of the influenza virus envelope in abortive infection. Journal of General Virology 42:73–88
     [Google Scholar]
  25. LUI C. 1955; Studies of influenza infection in ferrets by means of fluorescein-conjugated antibody. II. The role of “soluble antigen” in nuclear fluorescence and cross reactions. Journal of Experimental Medicine 101:665–676
     [Google Scholar]
  26. MULPFORDT H. 1982; The preparation of colloidal gold particles using tannic acid as an additional reducing agent. Experiential1127–1128
     [Google Scholar]
  27. OXFORD J. S., SCHILD G. C. 1975 Immunological studies with influenza matrix protein. Negative Strand Viruses 2611–620 Edited by Mahy B.W.J., Barry R. D. New York & London: Academic Press;
     [Google Scholar]
  28. PATTERSON S., OXFORD J. S., DOURMASHKIN R. R. 1979; Studies on the mechanism of influenza virus entry into cells. Journal of General Virology 43:223–229
     [Google Scholar]
  29. REES P. J., DIMMOCK N. J. 1981; Electrophoretic separation of influenza virus ribonucleoproteins. Journal of General Virology 53:125–132
     [Google Scholar]
  30. SCHILD G. C, OXFORD J. S., NEWMAN R. W. 1979; Evidence for antigenic variation in influenza A nucleoprotein. Virology 93:569–573
     [Google Scholar]
  31. SIMONS K., GAROFF H. 1980; The budding mechanisms of enveloped animal viruses. Journal of General Virology 50:1–21
     [Google Scholar]
  32. SLOT J., GEUZE H. J. 1981; Sizing of protein A-colloidal gold for immunoelectron microscopy. Journal of Cell Biology 90:533–536
     [Google Scholar]
  33. SMITH G. L., LEVIN J. Z., PALESE P., MOSS B. 1987; Synthesis and cellular location often influenza polypeptides individually expressed by recombinant vaccinia viruses. Virology 160:336–345
     [Google Scholar]
  34. TAYLOR J. M., HAMPSON A. W., WHITE D. O. 1969; The polypeptides of influenza virus. I. Cytoplasmic synthesis and nuclear accumulation. Virology 39:419–425
     [Google Scholar]
  35. TOWNSEND A. R. M., SKEHEL J. J., TAYLOR P. M., PALESE P. 1984a; Recognition of influenza A virus nucleoprotein by an H-2-restricted cytotoxic T-cell clone. Virology 133:456–459
     [Google Scholar]
  36. TOWNSEND A. R. M., MCMICHAEL A. J., CARTER N. P., HUDDLESTON J. A., BROWNLEE G. G. 1984b; Cytotoxic T cell recognition of influenza nucleoprotein and hemagglutinin expressed at the surface of transfected mouse L cells. Cell 39:13–25
     [Google Scholar]
  37. VIRELIZIER J. L., ALLISON A. C, OXFORD J. S., SCHILD G. C. 1977; Early presence of ribonucleoprotein antigen on the surface of influenza virus infected cells. Nature, London 266:52–54
     [Google Scholar]
  38. WATSON B. K., COONS A. H. 1954; Studies of influenza virus infection in chick embryos using fluorescent antibodies. Journal of Experimental Medicine 99:419–428
     [Google Scholar]
  39. YEWDELL J. W., FRANK E., GERHARD W. 1981; Expression of influenza A virus internal antigens on the surface of infected P815 cells. Journal of Immunology 126:1814–1819
     [Google Scholar]
  40. YEWDELL J. W., BENNINK J. R., SMITH G. L., MOSS B. 1985; Influenza A virus nucleoprotein is a major target antigen for cross reactive anti-influenza A virus cytotoxic T lymphocytes. Proceedings of the National Academy of Sciences, U.S.A 82:1785–1789
     [Google Scholar]
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The Intracellular Distribution of Influenza Virus Matrix Protein and Nucleoprotein in Infected Cells and Their Relationship to Haemagglutinin in the Plasma Membrane
J. Gen. Virol. 69, 1859 (1988); https://doi.org/10.1099/0022-1317-69-8-1859
/content/journal/jgv/10.1099/0022-1317-69-8-1859
/content/journal/jgv/10.1099/0022-1317-69-8-1859
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