Journal of Biomedical Science

, Volume 4, Issue 4, pp 139–145 | Cite as

Inhibition of the synthesis of proteins needed for Epstein-Barr virus replication by antisense RNA against the zta gene

  • Mei-Ying Liu
  • Jen-Yang Chen
  • Ching-Hwa Ann Tsai
  • Tsuey-Ying Hsu
  • Czau-Siung Yang
Original Paper


Antisense RNA complementary to the Epstein-Barr virus (EBV) Zta gene, an immediate-early gene encoding a transactivator, was applied to inhibit EBV protein synthesis during its lytic cycle. A DNA fragment containing the Zta gene sequence was inserted into an expression vector, pMAMneo, in a sense and antisense direction under a dexamethasone-inducible murine mammary tumor virus LTR promoter, resulting in the construction of plasmids pZ(+) and pZ(−), respectively. Synthesis of Zta protein was reduced in pZ(−)-transfected cells upon dexamethasone induction. Because D-form early antigen and DNA polymerase are essential for viral DNA replication, the contents of these two viral proteins were examined. Amounts of the two lytic proteins were observed to be significantly repressed in pZ(−)-transfected cells. In contrast, both proteins were normally expressed in the sense plasmid pZ(+) or cells transfected with vector alone. Above results demonstrate that Zta antisense RNA can reduce the production of Zta protein and the other lytic proteins, possibly resulting in the inhibition of EBV replication.

Key Words

Epstein-Barr virus Antisense RNA Zta 


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  1. 1.
    Chappuis BB, Muller H, Stutte J, Hey MM, Hubner K, Muller-Hermelink HK. Identification of EBV-DNA in lymph nodes and lymphomas in AIDS. Virchows Arch B Cell Pathol 58:199–205;1990.Google Scholar
  2. 2.
    Colby BM, Shaw JE, Elion GB, Pagano JS. Effects of acyclovir [9-(2-hydroxyethoxymethyl]guanine on Epstein-Barr virus DNA replication. J Virol 34:560–568;1980.Google Scholar
  3. 3.
    de-The G. The epidemiology of Burkitt's lymphoma: Evidence for a causal relationship with Epstein-Barr virus. Am J Epidemiol 1:32–54;1979.Google Scholar
  4. 4.
    Furnari FB, Adams MD, Pagano JS. Regulation of the Epstein-Barr virus DNA polymerase gene. J Virol 66:2837–2845;1992.Google Scholar
  5. 5.
    Green P, Pines O, Inouye M. The role of antisense RNA in gene regulation. Annu Rev Biochem 55:569–597;1986.Google Scholar
  6. 6.
    Henle G, Henle W, Diehl V. Relation of Burkitt's tumor-associated herpes-type virus to infectious mononucleosis. Proc Natl Acad Sci USA 59:94–101;1968.Google Scholar
  7. 7.
    Hinuma Y, Grace JT. Cloning of immunoglobulin producing human leukemic and lymphoma cells in long term culture. Proc Soc Exp Biol Med 124:107–111;1967.Google Scholar
  8. 8.
    Holley-Guthrie EA, Quinlivan EB, Mar E-C, Kenney S. The Epstein-Barr virus (EBV) BMRF1 promoter for early antigen (EA-D) is regulated by the EBV transactivators, BRLF1 and BZLF1, in a cell-specific manner. J Virol 64:3753–3759;1990.Google Scholar
  9. 9.
    Imai S, Koizumi S, Sugiura M, Tokunaga M, Uemura Y, Yamamoto N, Tanaka S, Sato E, Osato T. Gastric carcinoma: Monoclonal epithelial malignant cells expressing Epstein-Barr virus latent infection protein. Proc Natl Acad Sci USA 91:9131–9135;1994.Google Scholar
  10. 10.
    Izant JG, Weintraub H. Inhibition of thymidine kinase gene expression by anti-sense RNA: A molecular approach to genetic analysis. Cell 36:1007–1015;1984.Google Scholar
  11. 11.
    Izant JG, Weintraub H. Constitutive and conditional suppression of exogenous and endogenous genes by anti-sense RNA. Science 229:345–352;1985.Google Scholar
  12. 12.
    Johnson DA, Gautsch JW, Sportsman JR, Elder JH. Improved technique utilizing non-fat dry milk for analysis of proteins and nucleic acids transferred to nitrocellulose. Gene Anal Tech 1:3–8;1984.Google Scholar
  13. 13.
    Kieff E, Liebowitz D. Epstein-Barr virus and its replication. In: Fields BN, Knipe DM, Chanock R, Hirsch MS, Melnick JL, Monath TP, Roizman B, eds. Virology, ed 2. New York, Raven Press, 1889–1920;1990.Google Scholar
  14. 14.
    Kupfer SR, Summers WC. Identification of a glucocorticoid-responsive element in Epstein-Barr virus. J Virol 64:1984–1990;1990.Google Scholar
  15. 15.
    Lin JC, De Clercq E, Pagano JS. Novel acyclic adenosine analogs inhibit Epstein-Barr virus replication. Antimicrob Agents Chemother 33:1431–1433;1987.Google Scholar
  16. 16.
    Lin JC, Smith MC, Pagano JS. Prolonged inhibitory effect of 9-(1,3-dihydroxy-2-propoxymethyl)guanine against replication of Epstein-Barr virus. J Virol 50:50–55;1984.Google Scholar
  17. 17.
    Lin JC, Smith MC, Pagano JS. Comparative efficacy and selectivity of some nucleoside analogs against Epstein-Barr virus. Antimicrob Agents Chemother 27:971–973;1985.Google Scholar
  18. 18.
    Liu MY, Chou WH, Nutter L, Hsu MM, Chen JY, Yang CS. Antibody against Epstein-Barr virus DNA polymerase activity in sera of patients with nasopharyngeal carcinoma. J Med Virol 28:101–105;1989.Google Scholar
  19. 19.
    Miller G, Rabson M, Heston L. Epstein-Barr virus with heterogeneous DNA disrupts latency. J Virol 50:174–182;1984.Google Scholar
  20. 20.
    Miller RL, Glaser R, Rapp F. Studies of an Epstein-Barr virus-induced DNA polymerase. Virology 76:494–502;1977.Google Scholar
  21. 21.
    Old LJ, Boyse EA, Oettgen HF, de Harven E, Geering G, Williamson B, Clifford P. Precipitating antibody in human serum to an antigen present in cultured Burkitt's lymphoma cells. Proc Natl Acad Sci USA 56:1699–1704;1966.Google Scholar
  22. 22.
    Pallesen G, Hamilton-Dutoit SJ, Rowe M, Young LS. Expression of Epstein-Barr virus latent gene products in tumour cells of Hodgkin's disease. Lancet 337:320–322;1991.Google Scholar
  23. 23.
    Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: A laboratory manual, ed 2. Cold Spring Harbor, Cold Spring Harbor Press, 1989.Google Scholar
  24. 24.
    Shapiro RS, McClain K, Frizzerr G, Gajl-Peczalska KJ, Kersey JH, Blazar BR, Arther DC, Patton DF, Greenberg JS, Burke B, Ramsay NKC, McGlave P, Filipovich AH. Epstein-Barr virus-associated B-cell lymphoproliferative disorders following bone marrow transplantation. Blood 71:1234–1243;1988.Google Scholar
  25. 25.
    Simons R, Kleckner N. Biological regulation by antisense RNA in prokaryotes. Annu Rev Genet 22:567–600;1988.Google Scholar
  26. 26.
    Su IJ, Hsieh HC. Clinicopathological spectrum of Epstein-Barr virus-associated T cell malignancies. Leuk Lymphoma 7:47–53;1992.Google Scholar
  27. 27.
    Summers W, Klein G. Inhibition of Epstein-Barr virus DNA synthesis and late gene expression by phosphonoacetic acid. J Virol 18:151–155;1976.Google Scholar
  28. 28.
    Takada K, Ono Y. Synchronous and sequential activation of latently infected Epstein-Barr virus genomes. J Virol 63:445–449;1989.Google Scholar
  29. 29.
    Takada K, Shimizu N, Sakuma S, Ono Y. Transactivation of the latent Epstein-Barr virus (EBV) genome after transfection of the EBV DNA fragment. J Virol 57:1016–1022;1986.Google Scholar
  30. 30.
    Towbin H, Staehelin T, Gorden J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354;1979.Google Scholar
  31. 31.
    Tsai CH A, Marshall VW, Ronald G. Characterization of two monoclonal antibodies to Epstein-Barr virus diffuse early antigen which react to two different epitopes and have different biological function. J Virol Methods 33:47–52;1991.Google Scholar
  32. 32.
    Tsai CH, Liu MT, Chen MR, Lu J, Yang HL, Chen JY, Yang CS. Characterization of monoclonal antibodies to the Zta and DNase proteins of Epstein-Barr virus. J Biomed Sci, in press.Google Scholar

Copyright information

© National Science Council 1997

Authors and Affiliations

  • Mei-Ying Liu
    • 1
  • Jen-Yang Chen
    • 1
  • Ching-Hwa Ann Tsai
    • 1
  • Tsuey-Ying Hsu
    • 1
  • Czau-Siung Yang
    • 1
  1. 1.Graduate Institute of Microbiology College of MedicineNational Taiwan University, Number 1, Section 1TaipeiTaiwan (ROC)

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