Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1983 Mar;80(6):1726–1729. doi: 10.1073/pnas.80.6.1726

Rescue of transforming Epstein-Barr virus (EBV) from EBV-genome-positive epithelial hybrid cells transfected with subgenomic fragments of EBV DNA

Jay Stoerker 1, Ronald Glaser 1,*
PMCID: PMC393676  PMID: 6300871

Abstract

Transfection experiments using subgenomic fragments of the B95-8 strain of Epstein-Barr virus (EBV) DNA and EBV genome (HR-1)-positive epithelial/Burkitt hybrid cells (D98/HR-1) were carried out to determine whether an interaction between the transfecting virus fragment(s) and the endogenous HR-1 EBV genome could take place. Expression of EBV-specific antigens, including early antigen and virus capsid antigen, was examined in transfected cells by immunofluorescence. Attempts were also made to recover biologically active EBV from the D98/HR-1 cells after transfection with cloned fragments of B95-8 DNA. We found that D98/HR-1 cells transfected with the BamHI H or H, F, and X fragments were positive for early antigen 3 days after transfection. Spent media from transfected D98/HR-1 cells maintained for 20-30 days in culture were pooled, filtered, concentrated, and used as a potential source of virus to inoculate human umbilical cord blood lymphocytes. No evidence of transformation was observed with such preparations. However, if spent medium from D98/HR-1 cell cultures was first treated with iododeoxyuridine (to induce EBV DNA synthesis and replicative cycle) and then transfected with the BamHI H, F, and X fragments of B95-8 DNA and used to infect cord blood lymphocytes, transformation was obtained. A lymphoblastoid cell line derived in this manner, designated HI-HFX, is an EBV nuclear antigen-positive nonproducer cell line. Similar results were obtained with preparations from iododeoxyuridine-treated D98/HR-1 cells transfected with the EB 26-36 fragment of B95-8 DNA cloned in a Charon 4A vector. The EB 26-36 fragment contains the BamHI H, F, and X regions.

Keywords: transfection, transformation

Full text

PDF
1726

Images in this article

1727Image
on p.1727
1727Image
on p.1727

Selected References

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

  1. Blattner F. R., Williams B. G., Blechl A. E., Denniston-Thompson K., Faber H. E., Furlong L., Grunwald D. J., Kiefer D. O., Moore D. D., Schumm J. W. Charon phages: safer derivatives of bacteriophage lambda for DNA cloning. Science. 1977 Apr 8;196(4286):161–169. doi: 10.1126/science.847462. [DOI] [PubMed] [Google Scholar]
  2. Bornkamm G. W., Delius H., Zimber U., Hudewentz J., Epstein M. A. Comparison of Epstein-Barr virus strains of different origin by analysis of the viral DNAs. J Virol. 1980 Sep;35(3):603–618. doi: 10.1128/jvi.35.3.603-618.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cho M. S., Fresen K. O., zur Hausen H. Multiplicity-dependent biological and biochemical properties of Epstein-Barr virus (EBV) rescued from non-producer lines after superinfection with P3HR-1 EBV. Int J Cancer. 1980 Sep 15;26(3):357–363. doi: 10.1002/ijc.2910260316. [DOI] [PubMed] [Google Scholar]
  4. Durnford J. M., Scanlon P. A., Glaser R. Time of synthesis and cellular location of Epstein-Barr virus-specified polypeptides radioimmunoprecipitated from iododeoxyuridine-induced Burkitt lymphoma/somatic cell hybrids. J Gen Virol. 1982 May;60(Pt 1):39–48. doi: 10.1099/0022-1317-60-1-39. [DOI] [PubMed] [Google Scholar]
  5. EPSTEIN M. A., ACHONG B. G., BARR Y. M. VIRUS PARTICLES IN CULTURED LYMPHOBLASTS FROM BURKITT'S LYMPHOMA. Lancet. 1964 Mar 28;1(7335):702–703. doi: 10.1016/s0140-6736(64)91524-7. [DOI] [PubMed] [Google Scholar]
  6. Fresen K. O., Cho M. S., Gissmann L., zur Hausen H. NC37-R1 Epstein-Barr virus (EBV): a possible recombinant between intracellular NC37 viral DNA and superinfecting P3HR-1 EBV. Intervirology. 1980;12(6):303–310. doi: 10.1159/000149089. [DOI] [PubMed] [Google Scholar]
  7. Glaser R., Farrugia R., Brown N. Effect of the host cell on the maintenance and replication of Epstein-Barr virus. Virology. 1976 Jan;69(1):132–142. doi: 10.1016/0042-6822(76)90200-2. [DOI] [PubMed] [Google Scholar]
  8. Glaser R., Nonoyama M., Decker B., Rapp F. Synthesis of Epstein-Barr virus antigens and DNA in activated Burkitt somatic cell hybrids. Virology. 1973 Sep;55(1):62–69. doi: 10.1016/s0042-6822(73)81008-6. [DOI] [PubMed] [Google Scholar]
  9. Glaser R., Nonoyama M. Epstein-Barr virus: detection of genome in somatic cell hybrids of Burkitt lymphoblastoid cells. Science. 1973 Feb 2;179(4072):492–493. doi: 10.1126/science.179.4072.492. [DOI] [PubMed] [Google Scholar]
  10. Glaser R., Nonoyama M. Host cell regulation of induction of Epstein-Barr virus. J Virol. 1974 Jul;14(1):174–176. doi: 10.1128/jvi.14.1.174-176.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Glaser R., O'Neill F. J. Hybridization of Burkitt lymphoblastoid cells. Science. 1972 Jun 16;176(4040):1245–1247. doi: 10.1126/science.176.4040.1245. [DOI] [PubMed] [Google Scholar]
  12. Glaser R., Rapp F. Rescue of Epstein-Barr virus from somatic cell hybrids of Burkitt lymphoblastoid cells. J Virol. 1972 Aug;10(2):288–296. doi: 10.1128/jvi.10.2.288-296.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Graham F. L., van der Eb A. J. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. doi: 10.1016/0042-6822(73)90341-3. [DOI] [PubMed] [Google Scholar]
  14. Grogan E., Miller G., Henle W., Rabson M., Shedd D., Niederman J. C. Expression of Epstein-Barr viral early antigen in monolayer tissue cultures after transfection with viral DNA and DNA fragments. J Virol. 1981 Dec;40(3):861–869. doi: 10.1128/jvi.40.3.861-869.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Heller M., Dambaugh T., Kieff E. Epstein-Barr virus DNA. IX. Variation among viral DNAs from producer and nonproducer infected cells. J Virol. 1981 May;38(2):632–648. doi: 10.1128/jvi.38.2.632-648.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Henle G., Henle W. Immunofluorescence in cells derived from Burkitt's lymphoma. J Bacteriol. 1966 Mar;91(3):1248–1256. doi: 10.1128/jb.91.3.1248-1256.1966. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Henle W., Henle G., Ho H. C., Burtin P., Cachin Y., Clifford P., de Schryver A., de-Thé G., Diehl V., Klein G. Antibodies to Epstein-Barr virus in nasopharyngeal carcinoma, other head and neck neoplasms, and control groups. J Natl Cancer Inst. 1970 Jan;44(1):225–231. [PubMed] [Google Scholar]
  18. 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]
  19. Miller G., Grogan E., Heston L., Robinson J., Smith D. Epstein-Barr viral DNA: infectivity for human placental cells. Science. 1981 Apr 24;212(4493):452–455. doi: 10.1126/science.6259735. [DOI] [PubMed] [Google Scholar]
  20. Old L. J., Boyse E. A., Oettgen H. F., Harven E. D., 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 U S A. 1966 Dec;56(6):1699–1704. doi: 10.1073/pnas.56.6.1699. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Raab-Traub N., Pritchett R., Kieff E. DNA of Epstein-Barr virus. III. Identification of restriction enzyme fragments that contain DNA sequences which differ among strains of Epstein-Barr virus. J Virol. 1978 Aug;27(2):388–398. doi: 10.1128/jvi.27.2.388-398.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. 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]
  23. Stoerker J., Parris D., Yajima Y., Glaser R. Pleiotropic expression of Epstein--Barr virus DNA in human epithelial cells. Proc Natl Acad Sci U S A. 1981 Sep;78(9):5852–5855. doi: 10.1073/pnas.78.9.5852. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Stoerker J., Yajima Y., Glaser R. The interaction of non-transforming Epstein-Barr virus (EBV) with cell-associated EBV DNA in superinfected lymphoblastoid cell lines. Int J Cancer. 1982 Oct 15;30(4):385–392. doi: 10.1002/ijc.2910300402. [DOI] [PubMed] [Google Scholar]
  25. Volsky D. J., Klein G., Volsky B., Shapiro I. M. Production of infectious Epstein--Barr virus in mouse lymphocytes. Nature. 1981 Oct 1;293(5831):399–401. doi: 10.1038/293399a0. [DOI] [PubMed] [Google Scholar]
  26. Yajima Y., Marczynska B., Nonoyama M. Transforming activity of Epstein-Barr virus obtained by superinfection of Raji cells. Proc Natl Acad Sci U S A. 1978 Apr;75(4):2008–2010. doi: 10.1073/pnas.75.4.2008. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES