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. 1993 Dec;67(12):7472–7481. doi: 10.1128/jvi.67.12.7472-7481.1993

Comparing regions of the Epstein-Barr virus ZEBRA protein which function as transcriptional activating sequences in Saccharomyces cerevisiae and in B cells.

G Miller 1, H Himmelfarb 1, L Heston 1, J Countryman 1, L Gradoville 1, R Baumann 1, T Chi 1, M Carey 1
PMCID: PMC238213  PMID: 8230468

Abstract

The ZEBRA protein activates expression of Epstein-Barr virus early-lytic-cycle genes in human B lymphocytes. Here it is shown that ZEBRA also behaves as a sequence-specific transcriptional activator in Saccharomyces cerevisiae. Deletional mutagenesis defined three regions of ZEBRA that participate in activation in S. cerevisiae. These regions are designated YI (amino acids [aa] 1 to 25), YII (aa 51 to 102), and YIII (aa 228 to 245). Two of the three regions of the native ZEBRA protein act together to mediate activation when assayed on ZEBRA binding sites. However, when fused to the DNA binding domain of GAL4 and assayed on GAL4 binding sites, regions YII and YIII were each sufficient to confer activation in S. cerevisiae. Regions of ZEBRA which affected activation in S. cerevisiae were also required in human B lymphocytes. The amino-terminal region of ZEBRA (aa 1 to 98) was required for activation both in S. cerevisiae and in human B cells; deletion of the carboxy-terminal 18 aa also significantly reduced activation in both cell types. Thus, the behavior of ZEBRA in human B cells and S. cerevisiae suggests that the protein contains universal activation motifs that interact with conserved components of the transcription machinery. However, certain deletion mutants of ZEBRA containing mutations in the N-terminal region exhibited discordant behaviors in S. cerevisiae and in B cells. For example, deletion of ZEBRA aa 26 to 51 impaired activation to a great extent in B cells but had little or no effect in S. cerevisiae. The discordant mutants may reflect interactions with a variable domain of a conserved component or unique interactions with specialized components of the basal transcription apparatus in different cells.

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Selected References

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

  1. Baumann R., Grogan E., Ptashne M., Miller G. Changing Epstein-Barr viral ZEBRA protein into a more powerful activator enhances its capacity to disrupt latency. Proc Natl Acad Sci U S A. 1993 May 15;90(10):4436–4440. doi: 10.1073/pnas.90.10.4436. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Buratowski S., Hahn S., Sharp P. A., Guarente L. Function of a yeast TATA element-binding protein in a mammalian transcription system. Nature. 1988 Jul 7;334(6177):37–42. doi: 10.1038/334037a0. [DOI] [PubMed] [Google Scholar]
  3. Carey M., Kolman J., Katz D. A., Gradoville L., Barberis L., Miller G. Transcriptional synergy by the Epstein-Barr virus transactivator ZEBRA. J Virol. 1992 Aug;66(8):4803–4813. doi: 10.1128/jvi.66.8.4803-4813.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chang Y. N., Dong D. L., Hayward G. S., Hayward S. D. The Epstein-Barr virus Zta transactivator: a member of the bZIP family with unique DNA-binding specificity and a dimerization domain that lacks the characteristic heptad leucine zipper motif. J Virol. 1990 Jul;64(7):3358–3369. doi: 10.1128/jvi.64.7.3358-3369.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chasman D. I., Leatherwood J., Carey M., Ptashne M., Kornberg R. D. Activation of yeast polymerase II transcription by herpesvirus VP16 and GAL4 derivatives in vitro. Mol Cell Biol. 1989 Nov;9(11):4746–4749. doi: 10.1128/mcb.9.11.4746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Countryman J., Jenson H., Seibl R., Wolf H., Miller G. Polymorphic proteins encoded within BZLF1 of defective and standard Epstein-Barr viruses disrupt latency. J Virol. 1987 Dec;61(12):3672–3679. doi: 10.1128/jvi.61.12.3672-3679.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Davis M. G., Huang E. S. Transfer and expression of plasmids containing human cytomegalovirus immediate-early gene 1 promoter-enhancer sequences in eukaryotic and prokaryotic cells. Biotechnol Appl Biochem. 1988 Feb;10(1):6–12. [PubMed] [Google Scholar]
  8. Farrell P. J., Rowe D. T., Rooney C. M., Kouzarides T. Epstein-Barr virus BZLF1 trans-activator specifically binds to a consensus AP-1 site and is related to c-fos. EMBO J. 1989 Jan;8(1):127–132. doi: 10.1002/j.1460-2075.1989.tb03356.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fields S., Jang S. K. Presence of a potent transcription activating sequence in the p53 protein. Science. 1990 Aug 31;249(4972):1046–1049. doi: 10.1126/science.2144363. [DOI] [PubMed] [Google Scholar]
  10. Flemington E. K., Borras A. M., Lytle J. P., Speck S. H. Characterization of the Epstein-Barr virus BZLF1 protein transactivation domain. J Virol. 1992 Feb;66(2):922–929. doi: 10.1128/jvi.66.2.922-929.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gill G., Sadowski I., Ptashne M. Mutations that increase the activity of a transcriptional activator in yeast and mammalian cells. Proc Natl Acad Sci U S A. 1990 Mar;87(6):2127–2131. doi: 10.1073/pnas.87.6.2127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gill G., Tjian R. A highly conserved domain of TFIID displays species specificity in vivo. Cell. 1991 Apr 19;65(2):333–340. doi: 10.1016/0092-8674(91)90166-v. [DOI] [PubMed] [Google Scholar]
  13. Giniger E., Ptashne M. Transcription in yeast activated by a putative amphipathic alpha helix linked to a DNA binding unit. Nature. 1987 Dec 17;330(6149):670–672. doi: 10.1038/330670a0. [DOI] [PubMed] [Google Scholar]
  14. Giot J. F., Mikaelian I., Buisson M., Manet E., Joab I., Nicolas J. C., Sergeant A. Transcriptional interference between the EBV transcription factors EB1 and R: both DNA-binding and activation domains of EB1 are required. Nucleic Acids Res. 1991 Mar 25;19(6):1251–1258. doi: 10.1093/nar/19.6.1251. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Greenblatt J. Transcription. Riding high on the TATA box. Nature. 1992 Nov 5;360(6399):16–17. doi: 10.1038/360016a0. [DOI] [PubMed] [Google Scholar]
  17. Himmelfarb H. J., Pearlberg J., Last D. H., Ptashne M. GAL11P: a yeast mutation that potentiates the effect of weak GAL4-derived activators. Cell. 1990 Dec 21;63(6):1299–1309. doi: 10.1016/0092-8674(90)90425-e. [DOI] [PubMed] [Google Scholar]
  18. Horikoshi M., Yamamoto T., Ohkuma Y., Weil P. A., Roeder R. G. Analysis of structure-function relationships of yeast TATA box binding factor TFIID. Cell. 1990 Jun 29;61(7):1171–1178. doi: 10.1016/0092-8674(90)90681-4. [DOI] [PubMed] [Google Scholar]
  19. Ingles C. J., Shales M., Cress W. D., Triezenberg S. J., Greenblatt J. Reduced binding of TFIID to transcriptionally compromised mutants of VP16. Nature. 1991 Jun 13;351(6327):588–590. doi: 10.1038/351588a0. [DOI] [PubMed] [Google Scholar]
  20. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Johnston M., Davis R. W. Sequences that regulate the divergent GAL1-GAL10 promoter in Saccharomyces cerevisiae. Mol Cell Biol. 1984 Aug;4(8):1440–1448. doi: 10.1128/mcb.4.8.1440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Kakidani H., Ptashne M. GAL4 activates gene expression in mammalian cells. Cell. 1988 Jan 29;52(2):161–167. doi: 10.1016/0092-8674(88)90504-1. [DOI] [PubMed] [Google Scholar]
  23. Lambert P. F., Dostatni N., McBride A. A., Yaniv M., Howley P. M., Arcangioli B. Functional analysis of the papilloma virus E2 trans-activator in Saccharomyces cerevisiae. Genes Dev. 1989 Jan;3(1):38–48. doi: 10.1101/gad.3.1.38. [DOI] [PubMed] [Google Scholar]
  24. Lech K., Anderson K., Brent R. DNA-bound Fos proteins activate transcription in yeast. Cell. 1988 Jan 29;52(2):179–184. doi: 10.1016/0092-8674(88)90506-5. [DOI] [PubMed] [Google Scholar]
  25. Lieberman P. M., Berk A. J. The Zta trans-activator protein stabilizes TFIID association with promoter DNA by direct protein-protein interaction. Genes Dev. 1991 Dec;5(12B):2441–2454. doi: 10.1101/gad.5.12b.2441. [DOI] [PubMed] [Google Scholar]
  26. Lieberman P. M., Hardwick J. M., Sample J., Hayward G. S., Hayward S. D. The zta transactivator involved in induction of lytic cycle gene expression in Epstein-Barr virus-infected lymphocytes binds to both AP-1 and ZRE sites in target promoter and enhancer regions. J Virol. 1990 Mar;64(3):1143–1155. doi: 10.1128/jvi.64.3.1143-1155.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lue N. F., Buchman A. R., Kornberg R. D. Activation of yeast RNA polymerase II transcription by a thymidine-rich upstream element in vitro. Proc Natl Acad Sci U S A. 1989 Jan;86(2):486–490. doi: 10.1073/pnas.86.2.486. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Ma J., Ptashne M. Deletion analysis of GAL4 defines two transcriptional activating segments. Cell. 1987 Mar 13;48(5):847–853. doi: 10.1016/0092-8674(87)90081-x. [DOI] [PubMed] [Google Scholar]
  29. Metzger D., White J. H., Chambon P. The human oestrogen receptor functions in yeast. Nature. 1988 Jul 7;334(6177):31–36. doi: 10.1038/334031a0. [DOI] [PubMed] [Google Scholar]
  30. Miller G. The switch between latency and replication of Epstein-Barr virus. J Infect Dis. 1990 May;161(5):833–844. doi: 10.1093/infdis/161.5.833. [DOI] [PubMed] [Google Scholar]
  31. Moore P. A., Ruben S. M., Rosen C. A. Conservation of transcriptional activation functions of the NF-kappa B p50 and p65 subunits in mammalian cells and Saccharomyces cerevisiae. Mol Cell Biol. 1993 Mar;13(3):1666–1674. doi: 10.1128/mcb.13.3.1666. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Morrissey L. C., Barsoum J., Androphy E. J. Trans activation by the bovine papillomavirus E2 protein in Saccharomyces cerevisiae. J Virol. 1989 Oct;63(10):4422–4425. doi: 10.1128/jvi.63.10.4422-4425.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Nikolov D. B., Hu S. H., Lin J., Gasch A., Hoffmann A., Horikoshi M., Chua N. H., Roeder R. G., Burley S. K. Crystal structure of TFIID TATA-box binding protein. Nature. 1992 Nov 5;360(6399):40–46. doi: 10.1038/360040a0. [DOI] [PubMed] [Google Scholar]
  34. Packham G., Economou A., Rooney C. M., Rowe D. T., Farrell P. J. Structure and function of the Epstein-Barr virus BZLF1 protein. J Virol. 1990 May;64(5):2110–2116. doi: 10.1128/jvi.64.5.2110-2116.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Ptashne M., Gann A. A. Activators and targets. Nature. 1990 Jul 26;346(6282):329–331. doi: 10.1038/346329a0. [DOI] [PubMed] [Google Scholar]
  36. Ptashne M. How eukaryotic transcriptional activators work. Nature. 1988 Oct 20;335(6192):683–689. doi: 10.1038/335683a0. [DOI] [PubMed] [Google Scholar]
  37. Pugh B. F., Tjian R. Diverse transcriptional functions of the multisubunit eukaryotic TFIID complex. J Biol Chem. 1992 Jan 15;267(2):679–682. [PubMed] [Google Scholar]
  38. Sadowski I., Ptashne M. A vector for expressing GAL4(1-147) fusions in mammalian cells. Nucleic Acids Res. 1989 Sep 25;17(18):7539–7539. doi: 10.1093/nar/17.18.7539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Schena M., Yamamoto K. R. Mammalian glucocorticoid receptor derivatives enhance transcription in yeast. Science. 1988 Aug 19;241(4868):965–967. doi: 10.1126/science.3043665. [DOI] [PubMed] [Google Scholar]
  40. Struhl K. The JUN oncoprotein, a vertebrate transcription factor, activates transcription in yeast. Nature. 1988 Apr 14;332(6165):649–650. doi: 10.1038/332649a0. [DOI] [PubMed] [Google Scholar]
  41. Swaffield J. C., Bromberg J. F., Johnston S. A. Alterations in a yeast protein resembling HIV Tat-binding protein relieve requirement for an acidic activation domain in GAL4. Nature. 1992 Jun 25;357(6380):698–700. doi: 10.1038/357698a0. [DOI] [PubMed] [Google Scholar]
  42. Taylor N., Flemington E., Kolman J. L., Baumann R. P., Speck S. H., Miller G. ZEBRA and a Fos-GCN4 chimeric protein differ in their DNA-binding specificities for sites in the Epstein-Barr virus BZLF1 promoter. J Virol. 1991 Aug;65(8):4033–4041. doi: 10.1128/jvi.65.8.4033-4041.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Triezenberg S. J., Kingsbury R. C., McKnight S. L. Functional dissection of VP16, the trans-activator of herpes simplex virus immediate early gene expression. Genes Dev. 1988 Jun;2(6):718–729. doi: 10.1101/gad.2.6.718. [DOI] [PubMed] [Google Scholar]
  44. Urier G., Buisson M., Chambard P., Sergeant A. The Epstein-Barr virus early protein EB1 activates transcription from different responsive elements including AP-1 binding sites. EMBO J. 1989 May;8(5):1447–1453. doi: 10.1002/j.1460-2075.1989.tb03527.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Wu L. C., Tan T. H., Shahied S. I. Expression and characterization of the trans-activating protein Tax of human T-cell leukemia virus type I in Saccharomyces cerevisiae. J Virol. 1992 Dec;66(12):7253–7261. doi: 10.1128/jvi.66.12.7253-7261.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Yocum R. R., Hanley S., West R., Jr, Ptashne M. Use of lacZ fusions to delimit regulatory elements of the inducible divergent GAL1-GAL10 promoter in Saccharomyces cerevisiae. Mol Cell Biol. 1984 Oct;4(10):1985–1998. doi: 10.1128/mcb.4.10.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]

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