Summary
The sites of replication and of accumulation of viral macromolecules were examined using fluorescent antibodies to viral products and to cell organelles. Synthesis of envelope protein and its accumulation in a narrow rim around the nucleus were detected at 4 hours post infection; concurrently, a progressive change was observed in the rough endoplasmic reticulum from a fine to a coarse network emanating from around the nucleus. This change in the network was visible by light microscopy. The immunofluorescent sites of viral RNA synthesis, located by use of anti-double stranded RNA, extended from the perinuclear region in another fine network which included many small foci or vesicles; these sites were also visible by light microscopy late in infection. None of these changes were associated with any visible redistribution of actin, intermediate filaments or microtubules, and no nuclear involvement was detected. However, when microtubules were disrupted by vinblastine treatment of cells, the distribution of the immunofluorescent sites of viral RNA synthesis was modified and the virus yield was reduced by at least 10-fold. These results confirmed our biochemical studies showing separation of viral sites of RNA synthesis and translation, and the accumulation of envelope protein in nuclear-associated membranes. The relevance of these observations is discussed in relation to the reports of specific membrane structures induced in flavivirus-infected cells.
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References
Boulton, R. W., Westaway, E. G.: Replication of the flavivirus Kunjin: Proteins, glycoproteins, and maturation associated with cell membranes. Virology69, 416–430 (1976).
Boulton, R. W., Westaway, E. G.: Togavirus RNA: Reversible effect of urea on genomes and absence of subgenomic viral RNA in Kunjin virus-infected cells. Arch. Virol.55, 201–208 (1977).
Bretherton, L., Toh, B. H., Jack, I.: IgM autoantibody to intermediate filaments inMycoplasma pneumoniae infections. Clin. Immunol. Immunpathol.18, 425–430 (1981).
Calberg-Bacq, C.-M., Rentier-Delrue, F., Osterrieth, P. M., Duchesne, P. Y.: Electron microscopy studies on Banzi virus particle and its development in the suckling mice brains. J. Ultrastruct. Res.53, 193–203 (1975).
Cardiff, R. D., Russ, S. B., Brandt, W. E., Russell, P. K.: Cytological localization of dengue-2 antigens: An immunological study with ultrastructural correlation. Infect. Immun.7, 809–816 (1973).
Demsey A., Steere, R. L., Brandt, W. E., Veltri, B. J.: Morphology and development of dengue-2 virus employing the freeze-fracture and thinsection technique. J. Ultrastruct. Res.46, 103–116 (1974).
Filshie, B. K., Rehacek, J.: Studies of the morphology of Murray Valley encephalitis and Japanese encephalitis viruses growing in cultured mosquito cells. Virology34, 435–443 (1968).
Green, J., Griffiths, G., Louvard, D., Quinn, P., Warren, G.: Passage of viral membrane proteins through the Golgi complex. J. Mol. Biol.152, 663–698 (1981).
Kos, K. A., Shapiro, D., Vaitzus, Z., Russell, P. K.: Viral polypeptide composition of Japanese encephalitis virus-infected cell membranes. Arch. Virol.47, 217–224 (1975a).
Kos, K. A., Osborne, B. A., Goldsby, R. A.: Inhibition of group B arbovirus antigen production and replication in cells enucleated with cytochalasin B. J. Virology15, 913–917 (1975b).
Leary, K., Blair, C. D.: Sequential events in the morphogenesis of Japanese encephalitis virus. J. Ultrastruct. Res.72, 123–129 (1980).
Lewin, B.: Cell structure and genetics. In: Gene Expression2: Eucaryotic Chromosomes, 2nd ed., 11–70. New York: John Wiley 1980.
Lubiniecki, A. S., Henry, C. J.: Autoradiographic localization of RNA synthesis directed by arboviruses in the cytoplasm of infected BHK-21 cells. Proc. Soc. Exp. Biol. Med.145, 1165–1169 (1974).
Mathy, J. P., Baum, R., Toh, B. H.: Autoantibodies to ribosomes and Systemic lupus erythematosis. Clin. Exp. Immunol.41, 73–80 (1980).
Matsumura, T., Stollar, V., Schlesinger, R. W.: Studies on the nature of dengue viruses. V. Structure and development of dengue virus in Vero cells. Virology46, 344–355 (1971).
Murphy, F. A.: Togavirus morphology and morphogenesis. In:Schlesinger, R. W. (eds.), The Togaviruses: Biology, Structure, Replication, 241–316. New York: Academic Press 1980.
Plagemann, P. G.: Vinblastine sulphate: Metaphase arrest, inhibition of RNA synthesis, and cytotoxicity in Novikoff rat hepatoma cells. J. Nat. Cancer Inst.45, 589–595 (1970).
Shapiro, D., Kos, K., Brandt, W. E., Russell, P. K.: Membrane-bound proteins of Japanese encephalitis in virus-infected chick embryo cells. Virology48, 360–372 (1972).
Sriurairatna, S., Bhamapravati, N., Diwan, A. R., Halstead, S. B.: Ultrastructural studies on dengue virus infection of human lymphoblasts. Infect. Immun.20, 173–179 (1978).
Stohlman, S. A., Wisseman, C. L., Jr., Eylar, O. R., Silverman, D. J.: Dengue virus-induced modifications of host cell membranes. J. Virology16, 1017–1026 (1975).
Stollar, B. D., Stollar, V.: Immunofluorescent demonstration of double-stranded RNA in the cytoplasm of Sindbis virus-infected cells. Virology42, 276–280 (1970).
Takeda, H., Oya, A., Hashimoto, K., Yasuda, T., Yamada, M.-A.: Association of virus specific replicative ribonucleic acid with nuclear membrane in chick embryo cells infected with Japanese encephalitis virus. J. gen. Virol.38, 281–291 (1978).
Toh, B. H., Gallichio, H. A., Jeffrey, P. L., Livett, B. G., Muller, H. K., Cauchi, M. N., Clarke, F. M.: Anti-actin stains synapses. Nature (London)264, 648–650 (1976).
Watt, F. M., Harris, H.: Microtubule-organizing centres in mammalian cells in culture. J. Cell Science44, 103–121 (1980).
Westaway, E. G.: Proteins specified by group B togaviruses in mammalian cells during productive infections. Virology51, 454–465 (1973).
Westaway, E. G.: Replication of Flaviviruses. In:Schlesinger, R. W. (ed.), The Togaviruses: Biology, Structure, Replication, 531–581. New York: Academic Press 1980.
Westaway, E. G., Ng, M. L.: Replication of flaviviruses: Separation of translation sites of Kunjin virus proteins and of cell proteins. Virology106, 107–122 (1980).
Westaway, E. G., Shew, M., Della-Porta, A. J.: Reactions of purified hemagglutinating antigens of flaviviruses with 19 S and 7 S antibodies. Infect. Immun.11, 630–634 (1975).
Wright, P. J., Warr, H. M., Westaway, E. G.: Synthesis of glycoproteins in cells infected by the flavivirus Kunjin. Virology109, 418–427 (1981).
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Lee Ng, M., Pedersen, J.S., Toh, B.H. et al. Immunofluorescent sites in vero cells infected with the flavivirus Kunjin. Archives of Virology 78, 177–190 (1983). https://doi.org/10.1007/BF01311313
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DOI: https://doi.org/10.1007/BF01311313