1887

Abstract

The objective of this work was to identify novel viral ‘evasion’ genes without homology in the database through functional assays. Using this approach, the ‘unassigned’, conserved murine gammaherpesvirus ORF20 gene was shown to localize in the nucleus and to induce cell-cycle arrest followed by apoptosis in both mouse and human cells. Such growth-arrested cells did not express phospho-histone H3, demonstrating that the virus protein caused arrest at the G stage of the cell cycle. To characterize the mechanism further, Western blots of ORF20-recombinant lentivirus-infected cells were developed with antibodies to cyclin B1, Cdc2 and phospho-Tyr-15-Cdc2. This analysis revealed a relative increase in cyclin B and phospho-Tyr-15-Cdc2, from 24 to 72 h after infection with recombinant lentivirus. The demonstration that Cdc2 is in its inactive phosphorylated form and the clearly increased levels of cyclin B indicated that the virus gene blocks the progression of cells into mitosis by acting at the level of the Cdc2–cyclin B complex. To confirm this result, the Cdc2–cyclin B complex in ORF20-expressing cells was shown to be essentially without kinase activity. As the ORF20 gene is conserved in all herpesvirus, it may be presumed to have evolved to fulfil an important, as yet undefined, biological role in host-cell modification.

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2007-05-01
2019-11-13
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References

  1. Abraham, R. T. ( 2001; ). Cell cycle checkpoint signaling through the ATM and ATR kinases. Genes Dev 15, 2177–2196.[CrossRef]
    [Google Scholar]
  2. Alba, M. M., Lee, D., Pearl, F. M., Shepherd, A. J., Martin, N., Orengo, C. A. & Kellam, P. ( 2001; ). VIDA: a virus database system for the organization of animal virus genome open reading frames. Nucleic Acids Res 29, 133–136.[CrossRef]
    [Google Scholar]
  3. Boldogh, I., AbuBakar, S. & Albrecht, T. ( 1990; ). Activation of proto-oncogenes: an immediate early event in human cytomegalovirus infection. Science 247, 561–564.[CrossRef]
    [Google Scholar]
  4. Bortz, E., Whitelegge, J. P., Jia, Q., Zhou, Z. H., Stewart, J. P., Wu, T. T. & Sun, R. ( 2003; ). Identification of proteins associated with murine gammaherpesvirus 68 virions. J Virol 77, 13425–13432.[CrossRef]
    [Google Scholar]
  5. Castedo, M., Perfettini, J. L., Roumier, T. & Kroemer, G. ( 2002; ). Cyclin-dependent kinase-1: linking apoptosis to cell cycle and mitotic catastrophe. Cell Death Differ 9, 1287–1293.[CrossRef]
    [Google Scholar]
  6. Cayrol, C. & Flemington, E. ( 1996; ). G0/G1 growth arrest mediated by a region encompassing the basic leucine zipper (bZIP) domain of the Epstein–Barr virus transactivator Zta. J Biol Chem 271, 31799–31802.[CrossRef]
    [Google Scholar]
  7. Ebrahimi, B., Dutia, B. M., Roberts, K. L., Garcia-Ramirez, J. J., Dickinson, P., Stewart, J. P., Ghazal, P., Roy, D. J. & Nash, A. A. ( 2003; ). Transcriptome profile of murine gammaherpesvirus-68 lytic infection. J Gen Virol 84, 99–109.[CrossRef]
    [Google Scholar]
  8. Flano, E., Husain, S. M., Sample, J. T., Woodland, D. L. & Blackman, M. A. ( 2000; ). Latent murine gamma-herpesvirus infection is established in activated B cells, dendritic cells, and macrophages. J Immunol 165, 1074–1081.[CrossRef]
    [Google Scholar]
  9. Fournier, N., Raj, K., Saudan, P., Utzig, S., Sahli, R., Simanis, V. & Beard, P. ( 1999; ). Expression of human papillomavirus 16 E2 protein in Schizosaccharomyces pombe delays the initiation of mitosis. Oncogene 18, 4015–4021.[CrossRef]
    [Google Scholar]
  10. Friborg, J., Jr, Kong, W., Hottiger, M. O. & Nabel, G. J. ( 1999; ). p53 inhibition by the LANA protein of KSHV protects against cell death. Nature 402, 889–894.
    [Google Scholar]
  11. Goh, W. C., Rogel, M. E., Kinsey, C. M., Michael, S. F., Fultz, P. N., Nowak, M. A., Hahn, B. H. & Emerman, M. ( 1998; ). HIV-1 Vpr increases viral expression by manipulation of the cell cycle: a mechanism for selection of Vpr in vivo. Nat Med 4, 65–71.[CrossRef]
    [Google Scholar]
  12. Groschel, B. & Bushman, F. ( 2005; ). Cell cycle arrest in G2/M promotes early steps of infection by human immunodeficiency virus. J Virol 79, 5695–5704.[CrossRef]
    [Google Scholar]
  13. He, J., Choe, S., Walker, R., Di Marzio, P., Morgan, D. O. & Landau, N. R. ( 1995; ). Human immunodeficiency virus type 1 viral protein R (Vpr) arrests cells in the G2 phase of the cell cycle by inhibiting p34cdc2 activity. J Virol 69, 6705–6711.
    [Google Scholar]
  14. Hobbs, W. E., II & DeLuca, N. A. ( 1999; ). Perturbation of cell cycle progression and cellular gene expression as a function of herpes simplex virus ICP0. J Virol 73, 8245–8255.
    [Google Scholar]
  15. Hoge, A. T., Hendrickson, S. B. & Burns, W. H. ( 2000; ). Murine gammaherpesvirus 68 cyclin D homologue is required for efficient reactivation from latency. J Virol 74, 7016–7023.[CrossRef]
    [Google Scholar]
  16. Ikeda, Y., Collins, M. K., Radcliffe, P. A., Mitrophanous, K. A. & Takeuchi, Y. ( 2002; ). Gene transduction efficiency in cells of different species by HIV and EIAV vectors. Gene Ther 9, 932–938.[CrossRef]
    [Google Scholar]
  17. Izumiya, Y., Lin, S. F., Ellison, T. J., Levy, A. M., Mayeur, G. L., Izumiya, C. & Kung, H. J. ( 2003; ). Cell cycle regulation by Kaposi’s sarcoma-associated herpesvirus K-bZIP: direct interaction with cyclin-CDK2 and induction of G1 growth arrest. J Virol 77, 9652–9661.[CrossRef]
    [Google Scholar]
  18. Jacobson, J. G., Chen, S. H., Cook, W. J., Kramer, M. F. & Coen, D. M. ( 1998; ). Importance of the herpes simplex virus UL24 gene for productive ganglionic infection in mice. Virology 242, 161–169.[CrossRef]
    [Google Scholar]
  19. Jean, S., LeVan, K. M., Song, B., Levine, M. & Knipe, D. M. ( 2001; ). Herpes simplex virus 1 ICP27 is required for transcription of two viral late (γ2) genes in infected cells. Virology 283, 273–284.[CrossRef]
    [Google Scholar]
  20. Kastan, M. B. & Bartek, J. ( 2004; ). Cell-cycle checkpoints and cancer. Nature 432, 316–323.[CrossRef]
    [Google Scholar]
  21. Lu, M. & Shenk, T. ( 1999; ). Human cytomegalovirus UL69 protein induces cells to accumulate in G1 phase of the cell cycle. J Virol 73, 676–683.
    [Google Scholar]
  22. Lukas, J. & Bartek, J. ( 2004; ). Cell division: the heart of the cycle. Nature 432, 564–567.[CrossRef]
    [Google Scholar]
  23. Nash, A. A., Dutia, B. M., Stewart, J. P. & Davison, A. J. ( 2001; ). Natural history of murine gamma-herpesvirus infection. Philos Trans R Soc Lond B Biol Sci 356, 569–579.[CrossRef]
    [Google Scholar]
  24. Park, J., Seo, T., Hwang, S., Lee, D., Gwack, Y. & Choe, J. ( 2000; ). The K-bZIP protein from Kaposi’s sarcoma-associated herpesvirus interacts with p53 and represses its transcriptional activity. J Virol 74, 11977–11982.[CrossRef]
    [Google Scholar]
  25. Pearson, A. & Coen, D. M. ( 2002; ). Identification, localization, and regulation of expression of the UL24 protein of herpes simplex virus type 1. J Virol 76, 10821–10828.[CrossRef]
    [Google Scholar]
  26. Simas, J. P. & Efstathiou, S. ( 1998; ). Murine gammaherpesvirus 68: a model for the study of gammaherpesvirus pathogenesis. Trends Microbiol 6, 276–282.[CrossRef]
    [Google Scholar]
  27. Song, B., Liu, J. J., Yeh, K. C. & Knipe, D. M. ( 2000; ). Herpes simplex virus infection blocks events in the G1 phase of the cell cycle. Virology 267, 326–334.[CrossRef]
    [Google Scholar]
  28. Sunil-Chandra, N. P., Efstathiou, S., Arno, J. & Nash, A. A. ( 1992a; ). Virological and pathological features of mice infected with murine gamma-herpesvirus 68. J Gen Virol 73, 2347–2356.[CrossRef]
    [Google Scholar]
  29. Sunil-Chandra, N. P., Efstathiou, S. & Nash, A. A. ( 1992b; ). Murine gammaherpesvirus 68 establishes a latent infection in mouse B lymphocytes in vivo. J Gen Virol 73, 3275–3279.[CrossRef]
    [Google Scholar]
  30. Upton, J. W., van Dyk, L. F. & Speck, S. H. ( 2005; ). Characterization of murine gammaherpesvirus 68 v-cyclin interactions with cellular cdks. Virology 341, 271–283.[CrossRef]
    [Google Scholar]
  31. Virgin, H. W., IV, Latreille, P., Wamsley, P., Hallsworth, K., Weck, K. E., Dal Canto, A. J. & Speck, S. H. ( 1997; ). Complete sequence and genomic analysis of murine gammaherpesvirus 68. J Virol 71, 5894–5904.
    [Google Scholar]
  32. Weck, K. E., Kim, S. S., Virgin, H. I. & Speck, S. H. ( 1999; ). Macrophages are the major reservoir of latent murine gammaherpesvirus 68 in peritoneal cells. J Virol 73, 3273–3283.
    [Google Scholar]
  33. Zhu, Y., Gelbard, H. A., Roshal, M., Pursell, S., Jamieson, B. D. & Planelles, V. ( 2001; ). Comparison of cell cycle arrest, transactivation, and apoptosis induced by the simian immunodeficiency virus SIVagm and human immunodeficiency virus type 1 vpr genes. J Virol 75, 3791–3801.[CrossRef]
    [Google Scholar]
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