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. 1990 Jun;64(6):2759–2767. doi: 10.1128/jvi.64.6.2759-2767.1990

Identification of Epstein-Barr virus terminal protein 1 (TP1) in extracts of four lymphoid cell lines, expression in insect cells, and detection of antibodies in human sera.

B Frech 1, U Zimber-Strobl 1, K O Suentzenich 1, O Pavlish 1, G M Lenoir 1, G W Bornkamm 1, N Mueller-Lantzsch 1
PMCID: PMC249456  PMID: 2159542

Abstract

The terminal proteins TP1 and TP2 are putative products of Epstein-Barr virus (EBV) genes expressed during the latent cycle of the virus. They are predicted to code for 53- and 40-kilodalton integral membrane proteins. We used the baculovirus Autographa californica nuclear polyhedrosis virus as an expression vector to produce TP1 in large amounts in insect cells. The DNA sequences used to express TP1 originated from a TP1 cDNA derived from an M-ABA/CBL1 cDNA library. Rabbit antisera raised against procaryotic TP1 fusion proteins recognized a monomer and a dimer of the recombinant TP1 protein in the infected insect cells. Immunofluorescence studies of living insect cells showed that the recombinant protein is located in the plasma membrane. The insect cells infected with the recombinant baculovirus producing TP1 provided a test system to screen human antisera for TP1 antibodies. A total of 168 human EBV-positive and EBV-negative antisera were studied. TP1 antibodies were detected only in sera from nasopharyngeal carcinoma patients (16 out of 42). Rabbit antiserum raised against the recombinant TP1 protein expressed in the baculovirus system specifically recognized a protein of about 54 kilodaltons in the lymphoblastoid cell lines M-ABA and M-ABA/CBL1 and in the Burkitt's lymphoma cell lines BL18 and BL72. This protein could be located in the total membrane fraction of M-ABA cells and is upregulated by treating the cells with 12-O-tetradecanoylphorbol-13-acetate.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Albritton L. M., Tseng L., Scadden D., Cunningham J. M. A putative murine ecotropic retrovirus receptor gene encodes a multiple membrane-spanning protein and confers susceptibility to virus infection. Cell. 1989 May 19;57(4):659–666. doi: 10.1016/0092-8674(89)90134-7. [DOI] [PubMed] [Google Scholar]
  2. Baer R., Bankier A. T., Biggin M. D., Deininger P. L., Farrell P. J., Gibson T. J., Hatfull G., Hudson G. S., Satchwell S. C., Séguin C. DNA sequence and expression of the B95-8 Epstein-Barr virus genome. Nature. 1984 Jul 19;310(5974):207–211. doi: 10.1038/310207a0. [DOI] [PubMed] [Google Scholar]
  3. Bernheim A., Berger R., Lenoir G. Cytogenetic studies on African Burkitt's lymphoma cell lines: t(8;14), t(2;8) and t(8;22) translocations. Cancer Genet Cytogenet. 1981 Jun;3(4):307–315. doi: 10.1016/0165-4608(81)90039-x. [DOI] [PubMed] [Google Scholar]
  4. Boos H., Berger R., Kuklik-Roos C., Iftner T., Mueller-Lantzsch N. Enhancement of Epstein-Barr virus membrane protein (LMP) expression by serum, TPA, or n-butyrate in latently infected Raji cells. Virology. 1987 Jul;159(1):161–165. doi: 10.1016/0042-6822(87)90360-6. [DOI] [PubMed] [Google Scholar]
  5. Crawford D. H., Epstein M. A., Bornkamm G. W., Achong B. G., Finerty S., Thompson J. L. Biological and biochemical observations on isolates of EB virus from the malignant epithelial cells of two nasopharyngeal carcinomas. Int J Cancer. 1979 Sep 15;24(3):294–302. doi: 10.1002/ijc.2910240305. [DOI] [PubMed] [Google Scholar]
  6. Fennewald S., van Santen V., Kieff E. Nucleotide sequence of an mRNA transcribed in latent growth-transforming virus infection indicates that it may encode a membrane protein. J Virol. 1984 Aug;51(2):411–419. doi: 10.1128/jvi.51.2.411-419.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Helenius A., Simons K. Solubilization of membranes by detergents. Biochim Biophys Acta. 1975 Mar 25;415(1):29–79. doi: 10.1016/0304-4157(75)90016-7. [DOI] [PubMed] [Google Scholar]
  8. Henderson A., Ripley S., Heller M., Kieff E. Chromosome site for Epstein-Barr virus DNA in a Burkitt tumor cell line and in lymphocytes growth-transformed in vitro. Proc Natl Acad Sci U S A. 1983 Apr;80(7):1987–1991. doi: 10.1073/pnas.80.7.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Henle W., Diehl V., Kohn G., Zur Hausen H., Henle G. Herpes-type virus and chromosome marker in normal leukocytes after growth with irradiated Burkitt cells. Science. 1967 Sep 1;157(3792):1064–1065. doi: 10.1126/science.157.3792.1064. [DOI] [PubMed] [Google Scholar]
  10. Hennessy K., Fennewald S., Hummel M., Cole T., Kieff E. A membrane protein encoded by Epstein-Barr virus in latent growth-transforming infection. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7207–7211. doi: 10.1073/pnas.81.22.7207. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hinuma Y., Konn M., Yamaguchi J., Wudarski D. J., Blakeslee J. R., Jr, Grace J. T., Jr Immunofluorescence and herpes-type virus particles in the P3HR-1 Burkitt lymphoma cell line. J Virol. 1967 Oct;1(5):1045–1051. doi: 10.1128/jvi.1.5.1045-1051.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hudson G. S., Bankier A. T., Satchwell S. C., Barrell B. G. The short unique region of the B95-8 Epstein-Barr virus genome. Virology. 1985 Nov;147(1):81–98. doi: 10.1016/0042-6822(85)90229-6. [DOI] [PubMed] [Google Scholar]
  13. Klein G., Lindahl T., Jondal M., Leibold W., Menézes J., Nilsson K., Sundström C. Continuous lymphoid cell lines with characteristics of B cells (bone-marrow-derived), lacking the Epstein-Barr virus genome and derived from three human lymphomas. Proc Natl Acad Sci U S A. 1974 Aug;71(8):3283–3286. doi: 10.1073/pnas.71.8.3283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lanford R. E., Luckow V., Kennedy R. C., Dreesman G. R., Notvall L., Summers M. D. Expression and characterization of hepatitis B virus surface antigen polypeptides in insect cells with a baculovirus expression system. J Virol. 1989 Apr;63(4):1549–1557. doi: 10.1128/jvi.63.4.1549-1557.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Laux G., Perricaudet M., Farrell P. J. A spliced Epstein-Barr virus gene expressed in immortalized lymphocytes is created by circularization of the linear viral genome. EMBO J. 1988 Mar;7(3):769–774. doi: 10.1002/j.1460-2075.1988.tb02874.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lenoir G. M., Vuillaume M., Bonnardel C. The use of lymphomatous and lymphoblastoid cell lines in the study of Burkitt's lymphoma. IARC Sci Publ. 1985;(60):309–318. [PubMed] [Google Scholar]
  17. Lindahl T., Adams A., Bjursell G., Bornkamm G. W., Kaschka-Dierich C., Jehn U. Covalently closed circular duplex DNA of Epstein-Barr virus in a human lymphoid cell line. J Mol Biol. 1976 Apr 15;102(3):511–530. doi: 10.1016/0022-2836(76)90331-4. [DOI] [PubMed] [Google Scholar]
  18. Mann K. P., Staunton D., Thorley-Lawson D. A. Epstein-Barr virus-encoded protein found in plasma membranes of transformed cells. J Virol. 1985 Sep;55(3):710–720. doi: 10.1128/jvi.55.3.710-720.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Matsuo T., Heller M., Petti L., O'Shiro E., Kieff E. Persistence of the entire Epstein-Barr virus genome integrated into human lymphocyte DNA. Science. 1984 Dec 14;226(4680):1322–1325. doi: 10.1126/science.6095452. [DOI] [PubMed] [Google Scholar]
  20. Miller G., Lipman M. Release of infectious Epstein-Barr virus by transformed marmoset leukocytes. Proc Natl Acad Sci U S A. 1973 Jan;70(1):190–194. doi: 10.1073/pnas.70.1.190. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Mueller-Lantzsch N., Lenoir G. M., Sauter M., Takaki K., Béchet J. M., Kuklik-Roos C., Wunderlich D., Bornkamm G. W. Identification of the coding region for a second Epstein-Barr virus nuclear antigen (EBNA 2) by transfection of cloned DNA fragments. EMBO J. 1985 Jul;4(7):1805–1811. doi: 10.1002/j.1460-2075.1985.tb03854.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Polack A., Hartl G., Zimber U., Freese U. K., Laux G., Takaki K., Hohn B., Gissmann L., Bornkamm G. W. A complete set of overlapping cosmid clones of M-ABA virus derived from nasopharyngeal carcinoma and its similarity to other Epstein-Barr virus isolates. Gene. 1984 Mar;27(3):279–288. doi: 10.1016/0378-1119(84)90072-6. [DOI] [PubMed] [Google Scholar]
  23. Ricksten A., Kallin B., Alexander H., Dillner J., Fåhraeus R., Klein G., Lerner R., Rymo L. BamHI E region of the Epstein-Barr virus genome encodes three transformation-associated nuclear proteins. Proc Natl Acad Sci U S A. 1988 Feb;85(4):995–999. doi: 10.1073/pnas.85.4.995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Sample J., Liebowitz D., Kieff E. Two related Epstein-Barr virus membrane proteins are encoded by separate genes. J Virol. 1989 Feb;63(2):933–937. doi: 10.1128/jvi.63.2.933-937.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Sauter M., Mueller-Lantzsch N. Characterization of an Epstein-Barr virus nuclear antigen 2 variant (EBNA 2B) by specific sera. Virus Res. 1987 Aug;8(2):141–152. doi: 10.1016/0168-1702(87)90025-6. [DOI] [PubMed] [Google Scholar]
  26. Smith G. E., Summers M. D., Fraser M. J. Production of human beta interferon in insect cells infected with a baculovirus expression vector. Mol Cell Biol. 1983 Dec;3(12):2156–2165. doi: 10.1128/mcb.3.12.2156. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Speck S. H., Pfitzner A., Strominger J. L. An Epstein-Barr virus transcript from a latently infected, growth-transformed B-cell line encodes a highly repetitive polypeptide. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9298–9302. doi: 10.1073/pnas.83.24.9298. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Spindler K. R., Rosser D. S., Berk A. J. Analysis of adenovirus transforming proteins from early regions 1A and 1B with antisera to inducible fusion antigens produced in Escherichia coli. J Virol. 1984 Jan;49(1):132–141. doi: 10.1128/jvi.49.1.132-141.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Strnad B. C., Schuster T. C., Hopkins R. F., 3rd, Neubauer R. H., Rabin H. Identification of an Epstein-Barr virus nuclear antigen by fluoroimmunoelectrophoresis and radioimmunoelectrophoresis. J Virol. 1981 Jun;38(3):996–1004. doi: 10.1128/jvi.38.3.996-1004.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sugden B. An intricate route to immortality. Cell. 1989 Apr 7;57(1):5–7. doi: 10.1016/0092-8674(89)90165-7. [DOI] [PubMed] [Google Scholar]
  31. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Walter G., Scheidtmann K. H., Carbone A., Laudano A. P., Doolittle R. F. Antibodies specific for the carboxy- and amino-terminal regions of simian virus 40 large tumor antigen. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5197–5200. doi: 10.1073/pnas.77.9.5197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. White R. A., Wilcox M. Protein products of the bithorax complex in Drosophila. Cell. 1984 Nov;39(1):163–171. doi: 10.1016/0092-8674(84)90202-2. [DOI] [PubMed] [Google Scholar]

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