Skip to main content
NCBI home page
Journal of Virology logoLink to Journal of Virology
. 1984 Nov;52(2):549–556. doi: 10.1128/jvi.52.2.549-556.1984

Electron microscopic evidence for replication of circular Epstein-Barr virus genomes in latently infected Raji cells.

E Gussander, A Adams
PMCID: PMC254557  PMID: 6092676

Abstract

Raji cells, collected at various times from a synchronized culture, were gently lysed, and the high-molecular-weight DNA was enriched ca. 10-fold for latent Epstein-Barr virus (EBV) genomes by equilibrium density gradient centrifugation in neutral CsCl. The heavy-density DNA pool, which included more than 90% of the total intracellular EBV DNA sequences, was further fractionated by velocity sedimentation on neutral glycerol gradients, and material from fractions containing potential EBV DNA replicative forms was examined in the electron microscope. Early in the cellular S phase, when the EBV DNA content was found to be doubling in parallel with host chromosome replication, half of the 50- to 55-micron circular EBV genomes were observed to have two or more DNA branch points or forks. Most molecules were in a relaxed theta configuration, indicative of the Cairns mode of DNA replication. In the supercoiled state, the two daughter strands of the partially replicated molecules were seen to be wrapped around each other. Two theta structures had more than two DNA forks, indicating that DNA replication can initiate more than once on the same DNA molecule. Late in the S phase, the EBV DNA sedimenting at positions where theta structures were found with early S phase samples was composed of catenated dimers rather than partially replicated genomes. It is concluded that the circular EBV genomes, which are the major intracellular form in latently infected cells, are maintained as independent replicons and are not synthesized from an integrated template.

Full text

PDF
549

Images in this article

552Image
on p.552
553Image
on p.553
554Image
on p.554
555Image
on p.555

Selected References

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

  1. Adams A., Bjursell G., Kaschka-Dierich C., Lindahl T. Circular Epstein-Barr virus genomes of reduced size in a human lymphoid cell line of infectious mononucleosis origin. J Virol. 1977 May;22(2):373–380. doi: 10.1128/jvi.22.2.373-380.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Adams A., Lindahl T. Epstein-Barr virus genomes with properties of circular DNA molecules in carrier cells. Proc Natl Acad Sci U S A. 1975 Apr;72(4):1477–1481. doi: 10.1073/pnas.72.4.1477. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Adams A., Lindahl T. Intracellular forms of Epstein-Barr virus DNA in Raji cells. IARC Sci Publ. 1975;(11 Pt 1):125–132. [PubMed] [Google Scholar]
  4. Adams A., Lindahl T., Klein G. Linear association between cellular DNA and Epstein-Barr virus DNA in a human lymphoblastoid cell line. Proc Natl Acad Sci U S A. 1973 Oct;70(10):2888–2892. doi: 10.1073/pnas.70.10.2888. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Andersson M., Lindahl T. Epstein-Barr virus DNA in human lymphoid cell lines: in vitro conversion. Virology. 1976 Aug;73(1):96–105. doi: 10.1016/0042-6822(76)90064-7. [DOI] [PubMed] [Google Scholar]
  6. Benbow R. M., Eisenberg M., Sinsheimer R. L. Multiple length DNA molecules of bacteriophage phi-X174. Nat New Biol. 1972 May 31;237(74):141–144. doi: 10.1038/newbio237141a0. [DOI] [PubMed] [Google Scholar]
  7. 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]
  8. Freifelder D. Molecular weights of coliphages and coliphage DNA. IV. Molecular weights of DNA from bacteriophages T4, T5 and T7 and the general problem of determination of M. J Mol Biol. 1970 Dec 28;54(3):567–577. doi: 10.1016/0022-2836(70)90127-0. [DOI] [PubMed] [Google Scholar]
  9. Griffin B. E., Björck E., Bjursell G., Lindahl T. Sequence complexity of circular Epstein-Bar virus DNA in transformed cells. J Virol. 1981 Oct;40(1):11–19. doi: 10.1128/jvi.40.1.11-19.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gussander E., Adams A. Intracellular state of Epstein-Barr virus DNA in producer cell lines. J Gen Virol. 1979 Nov;45(2):331–340. doi: 10.1099/0022-1317-45-2-331. [DOI] [PubMed] [Google Scholar]
  11. Hampar B., Tanaka A., Nonoyama M., Derge J. G. Replication of the resident repressed Epstein-Barr virus genome during the early S phase (S-1 period) of nonproducer Raji cells. Proc Natl Acad Sci U S A. 1974 Mar;71(3):631–633. doi: 10.1073/pnas.71.3.631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Henry B. E., 2nd, Raab-Traub N. J., Pagano J. S. Detection of autonomous replicating sequences (ars) in the genome of Epstein-Barr virus. Proc Natl Acad Sci U S A. 1983 Feb;80(4):1096–1100. doi: 10.1073/pnas.80.4.1096. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jaenisch R., Mayer A., Levine A. Replicating SV40 molecules containing closed circular template DNA strands. Nat New Biol. 1971 Sep 15;233(37):72–75. doi: 10.1038/newbio233072a0. [DOI] [PubMed] [Google Scholar]
  14. 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]
  15. Nonoyama M., Pagano J. S. Detection of Epstein-Barr viral genome in nonproductive cells. Nat New Biol. 1971 Sep 22;233(38):103–106. doi: 10.1038/newbio233103a0. [DOI] [PubMed] [Google Scholar]
  16. Nonoyama M., Pagano J. S. Homology between Epstein-Barr virus DNA and viral DNA from Burkitt's lymphoma and nasopharyngeal carcinoma determined by DNA-DNA reassociation kinetics. Nature. 1973 Mar 2;242(5392):44–47. doi: 10.1038/242044a0. [DOI] [PubMed] [Google Scholar]
  17. Nonoyama M., Pagano J. S. Separation of Epstein-Barr virus DNA from large chromosomal DNA in non-virus-producing cells. Nat New Biol. 1972 Aug 9;238(84):169–171. doi: 10.1038/newbio238169a0. [DOI] [PubMed] [Google Scholar]
  18. Oppenheim A., Shlomai Z., Ben-Bassat H. Initiation points for cellular deoxyribonucleic acid replication in human lymphoid cells converted by Epstein-Barr virus. Mol Cell Biol. 1981 Aug;1(8):753–762. doi: 10.1128/mcb.1.8.753. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Pritchett R. F., Hayward S. D., Kieff E. D. DNA of Epstein-Barr virus. I. Comparative studies of the DNA of Epstein-Barr virus from HR-1 and B95-8 cells: size, structure, and relatedness. J Virol. 1975 Mar;15(3):556–559. doi: 10.1128/jvi.15.3.556-559.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Pritchett R., Pendersen M., Kieff E. Complexity of EBV homologous DNA in continous lymphoblastoid cell lines. Virology. 1976 Oct 1;74(1):227–231. [PubMed] [Google Scholar]
  21. Sebring E. D., Kelly T. J., Jr, Thoren M. M., Salzman N. P. Structure of replicating simian virus 40 deoxyribonucleic acid molecules. J Virol. 1971 Oct;8(4):478–490. doi: 10.1128/jvi.8.4.478-490.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Sundin O., Varshavsky A. Terminal stages of SV40 DNA replication proceed via multiply intertwined catenated dimers. Cell. 1980 Aug;21(1):103–114. doi: 10.1016/0092-8674(80)90118-x. [DOI] [PubMed] [Google Scholar]
  23. Vogelstein B., Pardoll D. M., Coffey D. S. Supercoiled loops and eucaryotic DNA replicaton. Cell. 1980 Nov;22(1 Pt 1):79–85. doi: 10.1016/0092-8674(80)90156-7. [DOI] [PubMed] [Google Scholar]
  24. zur Hausen H. Oncogenic Herpes viruses. Biochim Biophys Acta. 1975 Mar 20;417(1):25–53. doi: 10.1016/0304-419x(75)90007-4. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Virology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES