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. 1983 Jul;47(1):193–201. doi: 10.1128/jvi.47.1.193-201.1983

Identification of polypeptide components of the Epstein-Barr virus early antigen complex with monoclonal antibodies.

G R Pearson, B Vroman, B Chase, T Sculley, M Hummel, E Kieff
PMCID: PMC255226  PMID: 6306272

Abstract

Three monoclonal antibodies were produced against the Epstein-Barr virus-induced early antigen complex. These antibodies were shown to be specific for the early antigen complex by the fact that they only reacted with cells supporting a permissive or abortive Epstein-Barr virus infection and their synthesis was not affected by inhibitors of viral DNA synthesis. One monoclonal antibody, designated R3, was directed against a diffuse component of the early antigen complex since it reacted by immunofluorescence with cells fixed in acetone or methanol. The other two monoclonal antibodies, designated K8 and K9, reacted with a methanol-sensitive restricted component of this complex. The appearance of the R3 antigen in P3HR-1 superinfected Raji cells occurred approximately 4 h earlier than the antigen detected by K8. By both sodium dodecyl sulfate-polyacrylamide gel electrophoresis and radioimmunoelectrophoresis, it was determined that the R3 monoclonal antibody recognized two major polypeptides with molecular weights of approximately 50,000 to 52,000, whereas K8 and K9 precipitated a protein of approximately 85,000. The R3 monoclonal antibody also immunoprecipitated an in vitro primary translation product. It was, therefore, possible to map this product to the Epstein-Barr virus DNA BamH1 M fragment. These in vitro products were slightly smaller than the in vivo proteins, suggesting that these proteins probably undergo posttranslational modification during the virus replication cycle.

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Selected References

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  1. Bayliss G. J., Nonoyama M. Mechanisms of infection with Epstein-Barr virus. III. The synthesis of proteins in superinfected Raji cells. Virology. 1978 Jun 1;87(1):204–207. doi: 10.1016/0042-6822(78)90173-3. [DOI] [PubMed] [Google Scholar]
  2. Bayliss G. J., Wolf H. The regulated expression of Epstein-Barr virus. III. Proteins specified by EBV during the lytic cycle. J Gen Virol. 1981 Sep;56(Pt 1):105–118. doi: 10.1099/0022-1317-56-1-105. [DOI] [PubMed] [Google Scholar]
  3. Bodemer W. W., Summers W. C., Niederman J. C. Detection of virus-specific antigens in EB-(P3HR1) virus-superinfected Raji cells by immunoprecipitation. Virology. 1980 Jun;103(2):340–349. doi: 10.1016/0042-6822(80)90192-0. [DOI] [PubMed] [Google Scholar]
  4. Bos J. L., Polder L. J., Bernards R., Schrier P. I., van den Elsen P. J., van der Eb A. J., van Ormondt H. The 2.2 kb E1b mRNA of human Ad12 and Ad5 codes for two tumor antigens starting at different AUG triplets. Cell. 1981 Nov;27(1 Pt 2):121–131. doi: 10.1016/0092-8674(81)90366-4. [DOI] [PubMed] [Google Scholar]
  5. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  6. Derge J. G., Martos L. M., Tagamets M. A., Chang S. Y., Chakrabarty M. Identification of a critical period during the S phase for activation of the Epstein-Barr virus by 5-iododeoxyuridine. Nat New Biol. 1973 Aug 15;244(137):214–217. doi: 10.1038/newbio244214a0. [DOI] [PubMed] [Google Scholar]
  7. Feighny R. J., Farrell M. P., Pagano J. S. Polypeptide synthesis and phosphorylation in Epstein-Barr virus-infected cells. J Virol. 1980 May;34(2):455–463. doi: 10.1128/jvi.34.2.455-463.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Feighny R. J., Henry B. E., 2nd, Pagano J. S. Epstein-Barr virus polypeptides: effect of inhibition of viral DNA replication on their synthesis. J Virol. 1981 Jan;37(1):61–71. doi: 10.1128/jvi.37.1.61-71.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Feighny R. J., Henry B. E., 2nd, Pagano J. S. Epstein-Barr virus polypeptides: identification of early proteins and their synthesis and glycosylation. J Virol. 1981 Aug;39(2):651–655. doi: 10.1128/jvi.39.2.651-655.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Frink R. J., Anderson K. P., Wagner E. K. Herpes simplex virus type 1 HindIII fragment L encodes spliced and complementary mRNA species. J Virol. 1981 Aug;39(2):559–572. doi: 10.1128/jvi.39.2.559-572.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gerber P. Activation of Epstein-Barr virus by 5-bromodeoxyuridine in "virus-free" human cells (complement-fixing antigen-immunofluorescence-leukocytes). Proc Natl Acad Sci U S A. 1972 Jan;69(1):83–85. doi: 10.1073/pnas.69.1.83. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gergely L., Klein G., Ernberg I. Appearance of Epstein-Barr virus-associated antigens in infected Raji cells. Virology. 1971 Jul;45(1):10–21. doi: 10.1016/0042-6822(71)90107-3. [DOI] [PubMed] [Google Scholar]
  13. Henle G., Henle W., Klein G. Demonstration of two distinct components in the early antigen complex of Epstein-Barr virus-infected cells. Int J Cancer. 1971 Sep 15;8(2):272–282. doi: 10.1002/ijc.2910080212. [DOI] [PubMed] [Google Scholar]
  14. Henle W., Henle G., Zajac B. A., Pearson G., Waubke R., Scriba M. Differential reactivity of human serums with early antigens induced by Epstein-Barr virus. Science. 1970 Jul 10;169(3941):188–190. doi: 10.1126/science.169.3941.188. [DOI] [PubMed] [Google Scholar]
  15. Hummel M., Kieff E. Epstein-Barr virus RNA. VIII. Viral RNA in permissively infected B95-8 cells. J Virol. 1982 Jul;43(1):262–272. doi: 10.1128/jvi.43.1.262-272.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hummel M., Kieff E. Mapping of polypeptides encoded by the Epstein-Barr virus genome in productive infection. Proc Natl Acad Sci U S A. 1982 Sep;79(18):5698–5702. doi: 10.1073/pnas.79.18.5698. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kallin B., Luka J., Klein G. Immunochemical characterization of Epstein-Barr virus-associated early and late antigens in n-butyrate-treated P3HR-1 cells. J Virol. 1979 Dec;32(3):710–716. doi: 10.1128/jvi.32.3.710-716.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kawanishi M., Ito Y. Similarity of Epstein-Barr virus early polypeptides induced by various tumor promoters. Cancer Lett. 1982 May-Jun;16(1):19–23. doi: 10.1016/0304-3835(82)90086-6. [DOI] [PubMed] [Google Scholar]
  19. Kawanishi M., Sugawara K., Ito Y. Epstein-Barr virus-induced polypeptides: a comparative study with superinfected Raji, IUdR-Treated, and N-butyrate-treated P3HR-1 cells. Virology. 1981 Feb;109(1):72–81. doi: 10.1016/0042-6822(81)90472-4. [DOI] [PubMed] [Google Scholar]
  20. Michelson S., Horodniceanu F., Kress M., Tardy-Panit M. Human cytomegalovirus-induced immediate early antigens: analysis in sodium dodecyl sulfate-polyacrylamide gel electrophoresis after immunoprecipitation. J Virol. 1979 Oct;32(1):259–267. doi: 10.1128/jvi.32.1.259-267.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Mueller-Lantzsch N., Georg B., Yamamoto N., zur Hausen H. Epstein-Barr virus-induced proteins. III. Analysis of polypeptides from P3HR-1-EBV-superinfected NC37 cells by immunoprecipitation. Virology. 1980 Apr 15;102(1):231–233. doi: 10.1016/0042-6822(80)90087-2. [DOI] [PubMed] [Google Scholar]
  22. Mueller-Lantzsch N., Yamamoto N., zur Hausen H. Analysis of early and late Epstein-Barr virus associated polypeptides by immunoprecipitation. Virology. 1979 Sep;97(2):378–387. doi: 10.1016/0042-6822(79)90348-9. [DOI] [PubMed] [Google Scholar]
  23. Pearson G. R., Henle G., Henle W. Production of antigens associated with Epstein-Barr virus in experimentally infected lymphoblastoid cell lines. J Natl Cancer Inst. 1971 Jun;46(6):1243–1250. [PubMed] [Google Scholar]
  24. Preston C. M., McGeoch D. J. Identification and mapping of two polypeptides encoded within the herpes simplex virus type 1 thymidine kinase gene sequences. J Virol. 1981 May;38(2):593–605. doi: 10.1128/jvi.38.2.593-605.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Qualtiere L. F., Chase R., Vroman B., Pearson G. R. Identification of Epstein-Barr virus strain differences with monoclonal antibody to a membrane glycoprotein. Proc Natl Acad Sci U S A. 1982 Jan;79(2):616–620. doi: 10.1073/pnas.79.2.616. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Qualtiere L. F., Pearson G. R. Radioimmune precipitation study comparing the Epstein-Barr virus membrane antigens expressed on P3HR-1 virus-superinfected Raji cells to those expressed on cells in a B-95 virus-transformed producer culture activated with tumor-promoting agent (TPA). Virology. 1980 Apr 30;102(2):360–369. doi: 10.1016/0042-6822(80)90103-8. [DOI] [PubMed] [Google Scholar]
  27. Reedman B. M., Klein G. Cellular localization of an Epstein-Barr virus (EBV)-associated complement-fixing antigen in producer and non-producer lymphoblastoid cell lines. Int J Cancer. 1973 May;11(3):499–520. doi: 10.1002/ijc.2910110302. [DOI] [PubMed] [Google Scholar]
  28. Roubal J., Kallin B., Luka J., Klein G. Early DNA-binding polypeptides of Epstein-Barr virus. Virology. 1981 Aug;113(1):285–292. doi: 10.1016/0042-6822(81)90155-0. [DOI] [PubMed] [Google Scholar]
  29. Stark G. R., Williams J. G. Quantitative analysis of specific labelled RNA'S using DNA covalently linked to diazobenzyloxymethyl-paper. Nucleic Acids Res. 1979 Jan;6(1):195–203. doi: 10.1093/nar/6.1.195. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sugawara K., Kawanishi M., Ito Y. Epstein-barr virus-related DNA-binding proteins induced by n-butyrate in P3HR-1 cells. Virology. 1982 Jan 15;116(1):354–358. doi: 10.1016/0042-6822(82)90427-5. [DOI] [PubMed] [Google Scholar]
  31. Sugawara K., Osato T. Two distinct antigenic components in an Epstein-Barr virus-related early product induced by halogenated pyrimidines in non-producing human lymphoblastoid cells. Nat New Biol. 1973 Jun 13;243(128):209–210. doi: 10.1038/newbio243209a0. [DOI] [PubMed] [Google Scholar]
  32. Summers W. C., Klein G. Inhibition of Epstein-Barr virus DNA synthesis and late gene expression by phosphonoacetic acid. J Virol. 1976 Apr;18(1):151–155. doi: 10.1128/jvi.18.1.151-155.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. zur Hausen H., O'Neill F. J., Freese U. K., Hecker E. Persisting oncogenic herpesvirus induced by the tumour promotor TPA. Nature. 1978 Mar 23;272(5651):373–375. doi: 10.1038/272373a0. [DOI] [PubMed] [Google Scholar]

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