1887

Abstract

A mutant library of 249 mutants with mutations that span the entire Epstein–Barr virus (EBV) genome was generated by transposition with EZ : : TN and insertion with an apramycin resistance gene by a PCR-targeting method. This study also demonstrates the feasibility of generating deletions and site-specific mutations in the promoter on the EBV genome to determine the regions in the promoter that are crucial to transcription. Analysing and mutants by microarray analysis revealed that these two genes regulate the transcription of EBV lytic genes differently. A mutation affects global expression of EBV lytic genes; almost no lytic gene is expressed by the mutant after lytic induction. However, although a mutant still transcribes most lytic genes, the expression of these lytic genes is inefficient. Furthermore, this study shows that the proximal Zta-response element in the promoter is crucial to transcription from the EBV genome, despite the fact that another work demonstrated that this site was unimportant in transient transfection analysis. Furthermore, mutants with a mutation in and cannot assemble viral capsids. Results of this study demonstrate the usefulness of a comprehensive mutant library in genetic analyses of EBV.

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2007-09-01
2019-10-20
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References

  1. Ahsan, N., Kanda, T., Nagashima, K. & Takada, K. ( 2005; ). Epstein-Barr virus transforming protein LMP1 plays a critical role in virus production. J Virol 79, 4415–4424.[CrossRef]
    [Google Scholar]
  2. Altmann, M., Pich, D., Ruiss, R., Wang, J., Sugden, B. & Hammerschmidt, W. ( 2006; ). Transcriptional activation by EBV nuclear antigen 1 is essential for the expression of EBV's transforming genes. Proc Natl Acad Sci U S A 103, 14188–14193.[CrossRef]
    [Google Scholar]
  3. Asai, R., Kato, A., Kato, K., Kanamori-Koyama, M., Sugimoto, K., Sairenji, T., Nishiyama, Y. & Kawaguchi, Y. ( 2006; ). Epstein-Barr virus protein kinase BGLF4 is a virion tegument protein that dissociates from virions in a phosphorylation-dependent process and phosphorylates the viral immediate-early protein BZLF1. J Virol 80, 5125–5134.[CrossRef]
    [Google Scholar]
  4. Bhende, P. M., Seaman, W. T., Delecluse, H. J. & Kenney, S. C. ( 2004; ). The EBV lytic switch protein, Z, preferentially binds to and activates the methylated viral genome. Nat Genet 36, 1099–1104.[CrossRef]
    [Google Scholar]
  5. Bhende, P. M., Seaman, W. T., Delecluse, H. J. & Kenney, S. C. ( 2005; ). BZLF1 activation of the methylated form of the BRLF1 immediate-early promoter is regulated by BZLF1 residue 186. J Virol 79, 7338–7348.[CrossRef]
    [Google Scholar]
  6. Bloss, T. A. & Sugden, B. ( 1994; ). Optimal lengths for DNAs encapsidated by Epstein-Barr virus. J Virol 68, 8217–8222.
    [Google Scholar]
  7. Chang, L. K. & Liu, S. T. ( 2000; ). Activation of the BRLF1 promoter and lytic cycle of Epstein–Barr virus by histone acetylation. Nucleic Acids Res 28, 3918–3925.[CrossRef]
    [Google Scholar]
  8. Chang, L. K., Wei, T. T., Chiu, Y. F., Tung, C. P., Chuang, J. Y., Hung, S. K., Li, C. & Liu, S. T. ( 2003; ). Inhibition of Epstein–Barr virus lytic cycle by (−)-epigallocatechin gallate. Biochem Biophys Res Commun 301, 1062–1068.[CrossRef]
    [Google Scholar]
  9. Chang, L. K., Chung, J. Y., Hong, Y. R., Ichimura, T., Nakao, M. & Liu, S. T. ( 2005; ). Activation of Sp1-mediated transcription by Rta of Epstein–Barr virus via an interaction with MCAF1. Nucleic Acids Res 33, 6528–6539.[CrossRef]
    [Google Scholar]
  10. Chau, C. M., Zhang, X. Y., McMahon, S. B. & Lieberman, P. M. ( 2006; ). Regulation of Epstein-Barr virus latency type by the chromatin boundary factor CTCF. J Virol 80, 5723–5732.[CrossRef]
    [Google Scholar]
  11. Chen, A., Divisconte, M., Jiang, X., Quink, C. & Wang, F. ( 2005; ). Epstein-Barr virus with the latent infection nuclear antigen 3B completely deleted is still competent for B-cell growth transformation in vitro. J Virol 79, 4506–4509.[CrossRef]
    [Google Scholar]
  12. Collins, C. M., Medveczky, M. M., Lund, T. & Medveczky, P. G. ( 2002; ). The terminal repeats and latency-associated nuclear antigen of herpesvirus saimiri are essential for episomal persistence of the viral genome. J Gen Virol 83, 2269–2278.
    [Google Scholar]
  13. Datsenko, K. A. & Wanner, B. L. ( 2000; ). One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 97, 6640–6645.[CrossRef]
    [Google Scholar]
  14. Decaussin, G., Leclerc, V. & Ooka, T. ( 1995; ). The lytic cycle of Epstein-Barr virus in the nonproducer Raji line can be rescued by the expression of a 135-kilodalton protein encoded by the BALF2 open reading frame. J Virol 69, 7309–7314.
    [Google Scholar]
  15. Delecluse, H. J., Hilsendegen, T., Pich, D., Zeidler, R. & Hammerschmidt, W. ( 1998; ). Propagation and recovery of intact, infectious Epstein-Barr virus from prokaryotic to human cells. Proc Natl Acad Sci U S A 95, 8245–8250.[CrossRef]
    [Google Scholar]
  16. Delecluse, H. J., Pich, D., Hilsendegen, T., Baum, C. & Hammerschmidt, W. ( 1999; ). A first-generation packaging cell line for Epstein-Barr virus-derived vectors. Proc Natl Acad Sci U S A 96, 5188–5193.[CrossRef]
    [Google Scholar]
  17. Dirmeier, U., Neuhierl, B., Kilger, E., Reisbach, G., Sandberg, M. L. & Hammerschmidt, W. ( 2003; ). Latent membrane protein 1 is critical for efficient growth transformation of human B cells by Epstein-Barr virus. Cancer Res 63, 2982–2989.
    [Google Scholar]
  18. Farina, A., Feederle, R., Raffa, S., Gonnella, R., Santarelli, R., Frati, L., Angeloni, A., Torrisi, M. R., Faggioni, A. & Delecluse, H. J. ( 2005; ). BFRF1 of Epstein-Barr virus is essential for efficient primary viral envelopment and egress. J Virol 79, 3703–3712.[CrossRef]
    [Google Scholar]
  19. Feederle, R. & Delecluse, H. J. ( 2004; ). Low level of lytic replication in a recombinant Epstein-Barr virus carrying an origin of replication devoid of BZLF1-binding sites. J Virol 78, 12082–12084.[CrossRef]
    [Google Scholar]
  20. Feederle, R., Kost, M., Baumann, M., Janz, A., Drouet, E., Hammerschmidt, W. & Delecluse, H. J. ( 2000; ). The Epstein–Barr virus lytic program is controlled by the co-operative functions of two transactivators. EMBO J 19, 3080–3089.[CrossRef]
    [Google Scholar]
  21. Feederle, R., Shannon-Lowe, C., Baldwin, G. & Delecluse, H. J. ( 2005; ). Defective infectious particles and rare packaged genomes produced by cells carrying terminal-repeat-negative Epstein-Barr virus. J Virol 79, 7641–7647.[CrossRef]
    [Google Scholar]
  22. Feederle, R., Neuhierl, B., Baldwin, G., Bannert, H., Hub, B., Mautner, J., Behrends, U. & Delecluse, H. J. ( 2006; ). Epstein-Barr virus BNRF1 protein allows efficient transfer from the endosomal compartment to the nucleus of primary B lymphocytes. J Virol 80, 9435–9443.[CrossRef]
    [Google Scholar]
  23. Grabusic, K., Maier, S., Hartmann, A., Mantik, A., Hammerschmidt, W. & Kempkes, B. ( 2006; ). The CR4 region of EBNA2 confers viability of Epstein–Barr virus-transformed B cells by CBF1-independent signalling. J Gen Virol 87, 3169–3176.[CrossRef]
    [Google Scholar]
  24. Griffin, B. E., Bjorck, E., Bjursell, G. & Lindahl, T. ( 1981; ). Sequence complexity of circular Epstein-Bar virus DNA in transformed cells. J Virol 40, 11–19.
    [Google Scholar]
  25. Gust, B., Challis, G. L., Fowler, K., Kieser, T. & Chater, K. F. ( 2003; ). PCR-targeted Streptomyces gene replacement identifies a protein domain needed for biosynthesis of the sesquiterpene soil odor geosmin. Proc Natl Acad Sci U S A 100, 1541–1546.[CrossRef]
    [Google Scholar]
  26. Hatfull, G., Bankier, A. T., Barrell, B. G. & Farrell, P. J. ( 1988; ). Sequence analysis of Raji Epstein-Barr virus DNA. Virology 164, 334–340.[CrossRef]
    [Google Scholar]
  27. Ho, S. N., Hunt, H. D., Horton, R. M., Pullen, J. K. & Pease, L. R. ( 1989; ). Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene 77, 51–59.[CrossRef]
    [Google Scholar]
  28. Hong, G. K., Delecluse, H. J., Gruffat, H., Morrison, T. E., Feng, W. H., Sergeant, A. & Kenney, S. C. ( 2004; ). The BRRF1 early gene of Epstein-Barr virus encodes a transcription factor that enhances induction of lytic infection by BRLF1. J Virol 78, 4983–4992.[CrossRef]
    [Google Scholar]
  29. Humme, S., Reisbach, G., Feederle, R., Delecluse, H. J., Bousset, K., Hammerschmidt, W. & Schepers, A. ( 2003; ). The EBV nuclear antigen 1 (EBNA1) enhances B cell immortalization several thousandfold. Proc Natl Acad Sci U S A 100, 10989–10994.[CrossRef]
    [Google Scholar]
  30. Hung, C. H. & Liu, S. T. ( 1999; ). Characterization of the Epstein–Barr virus BALF2 promoter. J Gen Virol 80, 2747–2750.
    [Google Scholar]
  31. Hutchings, I. A., Tierney, R. J., Kelly, G. L., Stylianou, J., Rickinson, A. B. & Bell, A. I. ( 2006; ). Methylation status of the Epstein-Barr virus (EBV) BamHI W latent cycle promoter and promoter activity: analysis with novel EBV-positive Burkitt and lymphoblastoid cell lines. J Virol 80, 10700–10711.[CrossRef]
    [Google Scholar]
  32. Janz, A., Oezel, M., Kurzeder, C., Mautner, J., Pich, D., Kost, M., Hammerschmidt, W. & Delecluse, H. J. ( 2000; ). Infectious Epstein-Barr virus lacking major glycoprotein BLLF1 (gp350/220) demonstrates the existence of additional viral ligands. J Virol 74, 10142–10152.[CrossRef]
    [Google Scholar]
  33. Kado, C. I. & Liu, S. T. ( 1981; ). Rapid procedure for detection and isolation of large and small plasmids. J Bacteriol 145, 1365–1373.
    [Google Scholar]
  34. Kanda, T., Yajima, M., Ahsan, N., Tanaka, M. & Takada, K. ( 2004; ). Production of high-titer Epstein-Barr virus recombinants derived from Akata cells by using a bacterial artificial chromosome system. J Virol 78, 7004–7015.[CrossRef]
    [Google Scholar]
  35. Kelly, G. L., Milner, A. E., Tierney, R. J., Croom-Carter, D. S., Altmann, M., Hammerschmidt, W., Bell, A. I. & Rickinson, A. B. ( 2005; ). Epstein-Barr virus nuclear antigen 2 (EBNA2) gene deletion is consistently linked with EBNA3A, -3B, and -3C expression in Burkitt's lymphoma cells and with increased resistance to apoptosis. J Virol 79, 10709–10717.[CrossRef]
    [Google Scholar]
  36. Li, C., Chen, R. S., Hung, S. K., Lee, Y. T., Yen, C. Y., Lai, Y. W., Teng, R. H., Huang, J. Y., Tang, Y. C. & other authors ( 2006; ). Detection of Epstein–Barr virus infection and gene expression in human tumors by microarray analysis. J Virol Methods 133, 158–166.[CrossRef]
    [Google Scholar]
  37. Lu, C. C., Jeng, Y. Y., Tsai, C. H., Liu, M. Y., Yeh, S. W., Hsu, T. Y. & Chen, M. R. ( 2006; ). Genome-wide transcription program and expression of the Rta responsive gene of Epstein-Barr virus. Virology 345, 358–372.[CrossRef]
    [Google Scholar]
  38. Murphy, K. C. ( 1998; ). Use of bacteriophage lambda recombination functions to promote gene replacement in Escherichia coli. J Bacteriol 180, 2063–2071.
    [Google Scholar]
  39. Neuhierl, B. & Delecluse, H. J. ( 2006; ). The Epstein-Barr virus BMRF1 gene is essential for lytic virus replication. J Virol 80, 5078–5081.[CrossRef]
    [Google Scholar]
  40. Ryan, J. L., Fan, H., Glaser, S. L., Schichman, S. A., Raab-Traub, N. & Gulley, M. L. ( 2004; ). Epstein-Barr virus quantitation by real-time PCR targeting multiple gene segments: a novel approach to screen for the virus in paraffin-embedded tissue and plasma. J Mol Diagn 6, 378–385.[CrossRef]
    [Google Scholar]
  41. Sambrook, J., Fritsch, E. F. & Maniatis, T. ( 1989; ). Molecular Cloning: A Laboratory Manual. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  42. Shannon-Lowe, C. D., Neuhierl, B., Baldwin, G., Rickinson, A. B. & Delecluse, H. J. ( 2006; ). Resting B cells as a transfer vehicle for Epstein-Barr virus infection of epithelial cells. Proc Natl Acad Sci U S A 103, 7065–7070.[CrossRef]
    [Google Scholar]
  43. Sharma, R. C. & Schimke, R. T. ( 1996; ). Preparation of electrocompetent E. coli using salt-free growth medium. Biotechniques 20, 42–44.
    [Google Scholar]
  44. Sinclair, A. J., Brimmell, M., Shanahan, F. & Farrell, P. J. ( 1991; ). Pathways of activation of the Epstein-Barr virus productive cycle. J Virol 65, 2237–2244.
    [Google Scholar]
  45. Skare, J., Farley, J., Strominger, J. L., Fresen, K. O., Cho, M. S. & zur Hausen, H. ( 1985; ). Transformation by Epstein-Barr virus requires DNA sequences in the region of BamHI fragments Y and H. J Virol 55, 286–297.
    [Google Scholar]
  46. Song, M. J., Hwang, S., Wong, W. H., Wu, T. T., Lee, S., Liao, H. I. & Sun, R. ( 2005; ). Identification of viral genes essential for replication of murine gamma-herpesvirus 68 using signature-tagged mutagenesis. Proc Natl Acad Sci U S A 102, 3805–3810.[CrossRef]
    [Google Scholar]
  47. Wang, X. & Hutt-Fletcher, L. M. ( 1998; ). Epstein-Barr virus lacking glycoprotein gp42 can bind to B cells but is not able to infect. J Virol 72, 158–163.
    [Google Scholar]
  48. Wang, J. T., Yang, P. W., Lee, C. P., Han, C. H., Tsai, C. H. & Chen, M. R. ( 2005; ). Detection of Epstein–Barr virus BGLF4 protein kinase in virus replication compartments and virus particles. J Gen Virol 86, 3215–3225.[CrossRef]
    [Google Scholar]
  49. Yuan, J., Cahir-McFarland, E., Zhao, B. & Kieff, E. ( 2006; ). Virus and cell RNAs expressed during Epstein-Barr virus replication. J Virol 80, 2548–2565.[CrossRef]
    [Google Scholar]
  50. Zalani, S., Holley-Guthrie, E. A., Gutsch, D. E. & Kenney, S. C. ( 1992; ). The Epstein-Barr virus immediate-early promoter BRLF1 can be activated by the cellular Sp1 transcription factor. J Virol 66, 7282–7292.
    [Google Scholar]
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