Skip to main content
PMC 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
. 1995 Nov 7;92(23):10565–10569. doi: 10.1073/pnas.92.23.10565

Redefining the Epstein-Barr virus-encoded nuclear antigen EBNA-1 gene promoter and transcription initiation site in group I Burkitt lymphoma cell lines.

B C Schaefer 1, J L Strominger 1, S H Speck 1
PMCID: PMC40652  PMID: 7479841

Abstract

The Epstein-Barr virus-encoded nuclear antigen EBNA-1 gene promoter for the restricted Epstein-Barr virus (EBV) latency program operating in group I Burkitt lymphoma (BL) cell lines was previously identified incorrectly. Here we present evidence from RACE (rapid amplification of cDNA ends) cloning, reverse transcription-PCR, and S1 nuclease analyses, which demonstrates that the EBNA-1 gene promoter in group I BL cell lines is located in the viral BamHI Q fragment, immediately upstream of two low-affinity EBNA-1 binding sites. Transcripts initiated from this promoter, referred to as Qp, have the previously reported Q/U/K exon splicing pattern. Qp is active in group I BL cell lines but not in group III BL cell lines or in EBV immortalized B-lymphoblastoid cell lines. In addition, transient transfection of Qp-driven reporter constructs into both an EBV-negative BL cell line and a group I BL cell line gave rise to correctly initiated transcripts. Inspection of Qp revealed that it is a TATA-less promoter whose architecture is similar to the promoters of housekeeping genes, suggesting that Qp may be a default promoter which ensures EBNA-1 expression in cells that cannot run the full viral latency program. Elucidation of the genetic mechanism responsible for the EBNA-1-restricted program of EBV latency is an essential step in understanding control of viral latency in EBV-associated tumors.

Full text

PDF
10565

Images in this article

10566Fig. 1
on p.10566
10567Fig. 3
on p.10567
10568Fig. 4
on p.10568

Selected References

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

  1. Aviv H., Leder P. Purification of biologically active globin messenger RNA by chromatography on oligothymidylic acid-cellulose. Proc Natl Acad Sci U S A. 1972 Jun;69(6):1408–1412. doi: 10.1073/pnas.69.6.1408. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brandeis M., Frank D., Keshet I., Siegfried Z., Mendelsohn M., Nemes A., Temper V., Razin A., Cedar H. Sp1 elements protect a CpG island from de novo methylation. Nature. 1994 Sep 29;371(6496):435–438. doi: 10.1038/371435a0. [DOI] [PubMed] [Google Scholar]
  3. Brooks L., Yao Q. Y., Rickinson A. B., Young L. S. Epstein-Barr virus latent gene transcription in nasopharyngeal carcinoma cells: coexpression of EBNA1, LMP1, and LMP2 transcripts. J Virol. 1992 May;66(5):2689–2697. doi: 10.1128/jvi.66.5.2689-2697.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  5. Deacon E. M., Pallesen G., Niedobitek G., Crocker J., Brooks L., Rickinson A. B., Young L. S. Epstein-Barr virus and Hodgkin's disease: transcriptional analysis of virus latency in the malignant cells. J Exp Med. 1993 Feb 1;177(2):339–349. doi: 10.1084/jem.177.2.339. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Favaloro J., Treisman R., Kamen R. Transcription maps of polyoma virus-specific RNA: analysis by two-dimensional nuclease S1 gel mapping. Methods Enzymol. 1980;65(1):718–749. doi: 10.1016/s0076-6879(80)65070-8. [DOI] [PubMed] [Google Scholar]
  7. Flemington E., Speck S. H. Identification of phorbol ester response elements in the promoter of Epstein-Barr virus putative lytic switch gene BZLF1. J Virol. 1990 Mar;64(3):1217–1226. doi: 10.1128/jvi.64.3.1217-1226.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Frohman M. A., Dush M. K., Martin G. R. Rapid production of full-length cDNAs from rare transcripts: amplification using a single gene-specific oligonucleotide primer. Proc Natl Acad Sci U S A. 1988 Dec;85(23):8998–9002. doi: 10.1073/pnas.85.23.8998. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fåhraeus R., Fu H. L., Ernberg I., Finke J., Rowe M., Klein G., Falk K., Nilsson E., Yadav M., Busson P. Expression of Epstein-Barr virus-encoded proteins in nasopharyngeal carcinoma. Int J Cancer. 1988 Sep 15;42(3):329–338. doi: 10.1002/ijc.2910420305. [DOI] [PubMed] [Google Scholar]
  10. Gregory C. D., Rowe M., Rickinson A. B. Different Epstein-Barr virus-B cell interactions in phenotypically distinct clones of a Burkitt's lymphoma cell line. J Gen Virol. 1990 Jul;71(Pt 7):1481–1495. doi: 10.1099/0022-1317-71-7-1481. [DOI] [PubMed] [Google Scholar]
  11. Herbst H., Dallenbach F., Hummel M., Niedobitek G., Pileri S., Müller-Lantzsch N., Stein H. Epstein-Barr virus latent membrane protein expression in Hodgkin and Reed-Sternberg cells. Proc Natl Acad Sci U S A. 1991 Jun 1;88(11):4766–4770. doi: 10.1073/pnas.88.11.4766. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hitt M. M., Allday M. J., Hara T., Karran L., Jones M. D., Busson P., Tursz T., Ernberg I., Griffin B. E. EBV gene expression in an NPC-related tumour. EMBO J. 1989 Sep;8(9):2639–2651. doi: 10.1002/j.1460-2075.1989.tb08404.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Jansson A., Masucci M., Rymo L. Methylation of discrete sites within the enhancer region regulates the activity of the Epstein-Barr virus BamHI W promoter in Burkitt lymphoma lines. J Virol. 1992 Jan;66(1):62–69. doi: 10.1128/jvi.66.1.62-69.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Klein G., Dombos L., Gothoskar B. Sensitivity of Epstein-Barr virus (EBV) producer and non-producer human lymphoblastoid cell lines to superinfection with EB-virus. Int J Cancer. 1972 Jul 15;10(1):44–57. doi: 10.1002/ijc.2910100108. [DOI] [PubMed] [Google Scholar]
  15. Klein G. Epstein-Barr virus strategy in normal and neoplastic B cells. Cell. 1994 Jun 17;77(6):791–793. doi: 10.1016/0092-8674(94)90125-2. [DOI] [PubMed] [Google Scholar]
  16. Lear A. L., Rowe M., Kurilla M. G., Lee S., Henderson S., Kieff E., Rickinson A. B. The Epstein-Barr virus (EBV) nuclear antigen 1 BamHI F promoter is activated on entry of EBV-transformed B cells into the lytic cycle. J Virol. 1992 Dec;66(12):7461–7468. doi: 10.1128/jvi.66.12.7461-7468.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Loh E. Y., Elliott J. F., Cwirla S., Lanier L. L., Davis M. M. Polymerase chain reaction with single-sided specificity: analysis of T cell receptor delta chain. Science. 1989 Jan 13;243(4888):217–220. doi: 10.1126/science.2463672. [DOI] [PubMed] [Google Scholar]
  18. Macleod D., Charlton J., Mullins J., Bird A. P. Sp1 sites in the mouse aprt gene promoter are required to prevent methylation of the CpG island. Genes Dev. 1994 Oct 1;8(19):2282–2292. doi: 10.1101/gad.8.19.2282. [DOI] [PubMed] [Google Scholar]
  19. Masucci M. G., Contreras-Salazar B., Ragnar E., Falk K., Minarovits J., Ernberg I., Klein G. 5-Azacytidine up regulates the expression of Epstein-Barr virus nuclear antigen 2 (EBNA-2) through EBNA-6 and latent membrane protein in the Burkitt's lymphoma line rael. J Virol. 1989 Jul;63(7):3135–3141. doi: 10.1128/jvi.63.7.3135-3141.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Rowe D. T., Rowe M., Evan G. I., Wallace L. E., Farrell P. J., Rickinson A. B. Restricted expression of EBV latent genes and T-lymphocyte-detected membrane antigen in Burkitt's lymphoma cells. EMBO J. 1986 Oct;5(10):2599–2607. doi: 10.1002/j.1460-2075.1986.tb04540.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Rowe M., Lear A. L., Croom-Carter D., Davies A. H., Rickinson A. B. Three pathways of Epstein-Barr virus gene activation from EBNA1-positive latency in B lymphocytes. J Virol. 1992 Jan;66(1):122–131. doi: 10.1128/jvi.66.1.122-131.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Rowe M., Rowe D. T., Gregory C. D., Young L. S., Farrell P. J., Rupani H., Rickinson A. B. Differences in B cell growth phenotype reflect novel patterns of Epstein-Barr virus latent gene expression in Burkitt's lymphoma cells. EMBO J. 1987 Sep;6(9):2743–2751. doi: 10.1002/j.1460-2075.1987.tb02568.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Sample J., Brooks L., Sample C., Young L., Rowe M., Gregory C., Rickinson A., Kieff E. Restricted Epstein-Barr virus protein expression in Burkitt lymphoma is due to a different Epstein-Barr nuclear antigen 1 transcriptional initiation site. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6343–6347. doi: 10.1073/pnas.88.14.6343. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Schaefer B. C., Strominger J. L., Speck S. H. The Epstein-Barr virus BamHI F promoter is an early lytic promoter: lack of correlation with EBNA 1 gene transcription in group 1 Burkitt's lymphoma cell lines. J Virol. 1995 Aug;69(8):5039–5047. doi: 10.1128/jvi.69.8.5039-5047.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Schaefer B. C., Woisetschlaeger M., Strominger J. L., Speck S. H. Exclusive expression of Epstein-Barr virus nuclear antigen 1 in Burkitt lymphoma arises from a third promoter, distinct from the promoters used in latently infected lymphocytes. Proc Natl Acad Sci U S A. 1991 Aug 1;88(15):6550–6554. doi: 10.1073/pnas.88.15.6550. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Smith P. R., Griffin B. E. Transcription of the Epstein-Barr virus gene EBNA-1 from different promoters in nasopharyngeal carcinoma and B-lymphoblastoid cells. J Virol. 1992 Feb;66(2):706–714. doi: 10.1128/jvi.66.2.706-714.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Takada K., Horinouchi K., Ono Y., Aya T., Osato T., Takahashi M., Hayasaka S. An Epstein-Barr virus-producer line Akata: establishment of the cell line and analysis of viral DNA. Virus Genes. 1991 Apr;5(2):147–156. doi: 10.1007/BF00571929. [DOI] [PubMed] [Google Scholar]
  28. Tierney R. J., Steven N., Young L. S., Rickinson A. B. Epstein-Barr virus latency in blood mononuclear cells: analysis of viral gene transcription during primary infection and in the carrier state. J Virol. 1994 Nov;68(11):7374–7385. doi: 10.1128/jvi.68.11.7374-7385.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Weis L., Reinberg D. Transcription by RNA polymerase II: initiator-directed formation of transcription-competent complexes. FASEB J. 1992 Nov;6(14):3300–3309. doi: 10.1096/fasebj.6.14.1426767. [DOI] [PubMed] [Google Scholar]
  30. Woisetschlaeger M., Jin X. W., Yandava C. N., Furmanski L. A., Strominger J. L., Speck S. H. Role for the Epstein-Barr virus nuclear antigen 2 in viral promoter switching during initial stages of infection. Proc Natl Acad Sci U S A. 1991 May 1;88(9):3942–3946. doi: 10.1073/pnas.88.9.3942. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Woisetschlaeger M., Strominger J. L., Speck S. H. Mutually exclusive use of viral promoters in Epstein-Barr virus latently infected lymphocytes. Proc Natl Acad Sci U S A. 1989 Sep;86(17):6498–6502. doi: 10.1073/pnas.86.17.6498. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Woisetschlaeger M., Yandava C. N., Furmanski L. A., Strominger J. L., Speck S. H. Promoter switching in Epstein-Barr virus during the initial stages of infection of B lymphocytes. Proc Natl Acad Sci U S A. 1990 Mar;87(5):1725–1729. doi: 10.1073/pnas.87.5.1725. [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