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. 1993 Jul;13(7):4423–4431. doi: 10.1128/mcb.13.7.4423

Differential transcriptional activation by v-myb and c-myb in animal cells and Saccharomyces cerevisiae.

R H Chen 1, J S Lipsick 1
PMCID: PMC360011  PMID: 8321242

Abstract

The v-myb oncogene and its cellular homolog c-myb encode sequence-specific DNA-binding proteins which regulate transcription from promoters containing Myb-binding sites in animal cells. We have developed a Saccharomyces cerevisiae system to assay transcriptional activation by v-Myb and c-Myb. In yeast strains containing integrated reporter genes, activation was strictly dependent upon both the Myb DNA-binding domain and the Myb recognition element. BAS1, an endogenous Myb-related yeast protein, was not required for transactivation by animal Myb proteins and by itself had no detectable effect on a Myb reporter gene. Deletion analyses demonstrated that a domain of v-Myb C terminal to the previously mapped Myb transcriptional activation domain was required for transactivation in animal cells but not in S. cerevisiae. The same domain is also required for the efficient transformation of myeloid cells by v-Myb. In contrast to results in animal cells, in S. cerevisiae the full-length c-Myb was a much stronger transactivator than a protein bearing the oncogenic N- and C-terminal truncations of v-Myb. These results imply that negative regulation of c-Myb by its own termini requires an additional animal cell protein or small molecule that is not present in S. cerevisiae.

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Selected References

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

  1. Berger S. L., Piña B., Silverman N., Marcus G. A., Agapite J., Regier J. L., Triezenberg S. J., Guarente L. Genetic isolation of ADA2: a potential transcriptional adaptor required for function of certain acidic activation domains. Cell. 1992 Jul 24;70(2):251–265. doi: 10.1016/0092-8674(92)90100-q. [DOI] [PubMed] [Google Scholar]
  2. Bortner D. M., Ostrowski M. C. Analysis of the v-myb structural components important for transactivation of gene expression. Nucleic Acids Res. 1991 Apr 11;19(7):1533–1539. doi: 10.1093/nar/19.7.1533. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Boyle W. J., Lipsick J. S., Baluda M. A. Antibodies to the evolutionarily conserved amino-terminal region of the v-myb-encoded protein detect the c-myb protein in widely divergent metazoan species. Proc Natl Acad Sci U S A. 1986 Jul;83(13):4685–4689. doi: 10.1073/pnas.83.13.4685. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. 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]
  5. Cavallini B., Huet J., Plassat J. L., Sentenac A., Egly J. M., Chambon P. A yeast activity can substitute for the HeLa cell TATA box factor. Nature. 1988 Jul 7;334(6177):77–80. doi: 10.1038/334077a0. [DOI] [PubMed] [Google Scholar]
  6. Chen C., Okayama H. High-efficiency transformation of mammalian cells by plasmid DNA. Mol Cell Biol. 1987 Aug;7(8):2745–2752. doi: 10.1128/mcb.7.8.2745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chen J. H. Expression of endogenous avian myeloblastosis virus information in different chicken cells. J Virol. 1980 Oct;36(1):162–170. doi: 10.1128/jvi.36.1.162-170.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chien C. T., Bartel P. L., Sternglanz R., Fields S. The two-hybrid system: a method to identify and clone genes for proteins that interact with a protein of interest. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9578–9582. doi: 10.1073/pnas.88.21.9578. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Clarke M. F., Kukowska-Latallo J. F., Westin E., Smith M., Prochownik E. V. Constitutive expression of a c-myb cDNA blocks Friend murine erythroleukemia cell differentiation. Mol Cell Biol. 1988 Feb;8(2):884–892. doi: 10.1128/mcb.8.2.884. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Coll J., Saule S., Martin P., Raes M. B., Lagrou C., Graf T., Beug H., Simon I. E., Stehelin D. The cellular oncogenes c-myc, c-myb and c-erb are transcribed in defined types of avian hematopoietic cells. Exp Cell Res. 1983 Nov;149(1):151–162. doi: 10.1016/0014-4827(83)90388-9. [DOI] [PubMed] [Google Scholar]
  11. Dalton S., Treisman R. Characterization of SAP-1, a protein recruited by serum response factor to the c-fos serum response element. Cell. 1992 Feb 7;68(3):597–612. doi: 10.1016/0092-8674(92)90194-h. [DOI] [PubMed] [Google Scholar]
  12. Donahue T. F., Cigan A. M. Sequence and structural requirements for efficient translation in yeast. Methods Enzymol. 1990;185:366–372. doi: 10.1016/0076-6879(90)85032-j. [DOI] [PubMed] [Google Scholar]
  13. Dubendorff J. W., Whittaker L. J., Eltman J. T., Lipsick J. S. Carboxy-terminal elements of c-Myb negatively regulate transcriptional activation in cis and in trans. Genes Dev. 1992 Dec;6(12B):2524–2535. doi: 10.1101/gad.6.12b.2524. [DOI] [PubMed] [Google Scholar]
  14. Evan G. I., Lewis G. K., Bishop J. M. Isolation of monoclonal antibodies specific for products of avian oncogene myb. Mol Cell Biol. 1984 Dec;4(12):2843–2850. doi: 10.1128/mcb.4.12.2843. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fields S., Song O. A novel genetic system to detect protein-protein interactions. Nature. 1989 Jul 20;340(6230):245–246. doi: 10.1038/340245a0. [DOI] [PubMed] [Google Scholar]
  16. Frampton J., Gibson T. J., Ness S. A., Döderlein G., Graf T. Proposed structure for the DNA-binding domain of the Myb oncoprotein based on model building and mutational analysis. Protein Eng. 1991 Dec;4(8):891–901. doi: 10.1093/protein/4.8.891. [DOI] [PubMed] [Google Scholar]
  17. Gabrielsen O. S., Sentenac A., Fromageot P. Specific DNA binding by c-Myb: evidence for a double helix-turn-helix-related motif. Science. 1991 Sep 6;253(5024):1140–1143. doi: 10.1126/science.1887237. [DOI] [PubMed] [Google Scholar]
  18. Garcia A., LaMontagne K., Reavis D., Stober-Grässer U., Lipsick J. S. Determinants of sequence-specific DNA-binding by p48v-myb. Oncogene. 1991 Feb;6(2):265–273. [PubMed] [Google Scholar]
  19. Gewirtz A. M., Calabretta B. A c-myb antisense oligodeoxynucleotide inhibits normal human hematopoiesis in vitro. Science. 1988 Dec 2;242(4883):1303–1306. doi: 10.1126/science.2461588. [DOI] [PubMed] [Google Scholar]
  20. Gietz R. D., Sugino A. New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites. Gene. 1988 Dec 30;74(2):527–534. doi: 10.1016/0378-1119(88)90185-0. [DOI] [PubMed] [Google Scholar]
  21. Gonda T. J., Sheiness D. K., Bishop J. M. Transcripts from the cellular homologs of retroviral oncogenes: distribution among chicken tissues. Mol Cell Biol. 1982 Jun;2(6):617–624. doi: 10.1128/mcb.2.6.617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Graf T. Myb: a transcriptional activator linking proliferation and differentiation in hematopoietic cells. Curr Opin Genet Dev. 1992 Apr;2(2):249–255. doi: 10.1016/s0959-437x(05)80281-3. [DOI] [PubMed] [Google Scholar]
  23. Grässer F. A., Graf T., Lipsick J. S. Protein truncation is required for the activation of the c-myb proto-oncogene. Mol Cell Biol. 1991 Aug;11(8):3987–3996. doi: 10.1128/mcb.11.8.3987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Grässer F. A., LaMontagne K., Whittaker L., Stohr S., Lipsick J. S. A highly conserved cysteine in the v-Myb DNA-binding domain is essential for transformation and transcriptional trans-activation. Oncogene. 1992 May;7(5):1005–1009. [PubMed] [Google Scholar]
  25. Guehmann S., Vorbrueggen G., Kalkbrenner F., Moelling K. Reduction of a conserved Cys is essential for Myb DNA-binding. Nucleic Acids Res. 1992 May 11;20(9):2279–2286. doi: 10.1093/nar/20.9.2279. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Howe K. M., Reakes C. F., Watson R. J. Characterization of the sequence-specific interaction of mouse c-myb protein with DNA. EMBO J. 1990 Jan;9(1):161–169. doi: 10.1002/j.1460-2075.1990.tb08092.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Hu Y. L., Ramsay R. G., Kanei-Ishii C., Ishii S., Gonda T. J. Transformation by carboxyl-deleted Myb reflects increased transactivating capacity and disruption of a negative regulatory domain. Oncogene. 1991 Sep;6(9):1549–1553. [PubMed] [Google Scholar]
  28. Ibanez C. E., Lipsick J. S. Structural and functional domains of the myb oncogene: requirements for nuclear transport, myeloid transformation, and colony formation. J Virol. 1988 Jun;62(6):1981–1988. doi: 10.1128/jvi.62.6.1981-1988.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Ibanez C. E., Lipsick J. S. trans activation of gene expression by v-myb. Mol Cell Biol. 1990 May;10(5):2285–2293. doi: 10.1128/mcb.10.5.2285. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Introna M., Golay J., Frampton J., Nakano T., Ness S. A., Graf T. Mutations in v-myb alter the differentiation of myelomonocytic cells transformed by the oncogene. Cell. 1990 Dec 21;63(6):1289–1297. doi: 10.1016/0092-8674(90)90424-d. [DOI] [PubMed] [Google Scholar]
  31. 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]
  32. Johnson A. D., Herskowitz I. A repressor (MAT alpha 2 Product) and its operator control expression of a set of cell type specific genes in yeast. Cell. 1985 Aug;42(1):237–247. doi: 10.1016/s0092-8674(85)80119-7. [DOI] [PubMed] [Google Scholar]
  33. 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]
  34. Kalkbrenner F., Guehmann S., Moelling K. Transcriptional activation by human c-myb and v-myb genes. Oncogene. 1990 May;5(5):657–661. [PubMed] [Google Scholar]
  35. Katzen A. L., Kornberg T. B., Bishop J. M. Isolation of the proto-oncogene c-myb from D. melanogaster. Cell. 1985 Jun;41(2):449–456. doi: 10.1016/s0092-8674(85)80018-0. [DOI] [PubMed] [Google Scholar]
  36. Klempnauer K. H., Arnold H., Biedenkapp H. Activation of transcription by v-myb: evidence for two different mechanisms. Genes Dev. 1989 Oct;3(10):1582–1589. doi: 10.1101/gad.3.10.1582. [DOI] [PubMed] [Google Scholar]
  37. Klempnauer K. H., Sippel A. E. The highly conserved amino-terminal region of the protein encoded by the v-myb oncogene functions as a DNA-binding domain. EMBO J. 1987 Sep;6(9):2719–2725. doi: 10.1002/j.1460-2075.1987.tb02565.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Lane T., Ibanez C., Garcia A., Graf T., Lipsick J. Transformation by v-myb correlates with trans-activation of gene expression. Mol Cell Biol. 1990 Jun;10(6):2591–2598. doi: 10.1128/mcb.10.6.2591. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Leprince D., Gegonne A., Coll J., de Taisne C., Schneeberger A., Lagrou C., Stehelin D. A putative second cell-derived oncogene of the avian leukaemia retrovirus E26. Nature. 1983 Nov 24;306(5941):395–397. doi: 10.1038/306395a0. [DOI] [PubMed] [Google Scholar]
  40. Lillie J. W., Green M. R. Transcription activation by the adenovirus E1a protein. Nature. 1989 Mar 2;338(6210):39–44. doi: 10.1038/338039a0. [DOI] [PubMed] [Google Scholar]
  41. Lüscher B., Christenson E., Litchfield D. W., Krebs E. G., Eisenman R. N. Myb DNA binding inhibited by phosphorylation at a site deleted during oncogenic activation. Nature. 1990 Apr 5;344(6266):517–522. doi: 10.1038/344517a0. [DOI] [PubMed] [Google Scholar]
  42. Lüscher B., Eisenman R. N. New light on Myc and Myb. Part II. Myb. Genes Dev. 1990 Dec;4(12B):2235–2241. doi: 10.1101/gad.4.12b.2235. [DOI] [PubMed] [Google Scholar]
  43. Ma J., Ptashne M. A new class of yeast transcriptional activators. Cell. 1987 Oct 9;51(1):113–119. doi: 10.1016/0092-8674(87)90015-8. [DOI] [PubMed] [Google Scholar]
  44. 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]
  45. Moelling K., Pfaff E., Beug H., Beimling P., Bunte T., Schaller H. E., Graf T. DNA-binding activity is associated with purified myb proteins from AMV and E26 viruses and is temperature-sensitive for E26 ts mutants. Cell. 1985 Apr;40(4):983–990. doi: 10.1016/0092-8674(85)90358-7. [DOI] [PubMed] [Google Scholar]
  46. Mucenski M. L., McLain K., Kier A. B., Swerdlow S. H., Schreiner C. M., Miller T. A., Pietryga D. W., Scott W. J., Jr, Potter S. S. A functional c-myb gene is required for normal murine fetal hepatic hematopoiesis. Cell. 1991 May 17;65(4):677–689. doi: 10.1016/0092-8674(91)90099-k. [DOI] [PubMed] [Google Scholar]
  47. Ness S. A., Marknell A., Graf T. The v-myb oncogene product binds to and activates the promyelocyte-specific mim-1 gene. Cell. 1989 Dec 22;59(6):1115–1125. doi: 10.1016/0092-8674(89)90767-8. [DOI] [PubMed] [Google Scholar]
  48. Nishina Y., Nakagoshi H., Imamoto F., Gonda T. J., Ishii S. Trans-activation by the c-myb proto-oncogene. Nucleic Acids Res. 1989 Jan 11;17(1):107–117. doi: 10.1093/nar/17.1.107. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Nomura N., Takahashi M., Matsui M., Ishii S., Date T., Sasamoto S., Ishizaki R. Isolation of human cDNA clones of myb-related genes, A-myb and B-myb. Nucleic Acids Res. 1988 Dec 9;16(23):11075–11089. doi: 10.1093/nar/16.23.11075. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Nunn M. F., Seeburg P. H., Moscovici C., Duesberg P. H. Tripartite structure of the avian erythroblastosis virus E26 transforming gene. Nature. 1983 Nov 24;306(5941):391–395. doi: 10.1038/306391a0. [DOI] [PubMed] [Google Scholar]
  51. Ogata K., Hojo H., Aimoto S., Nakai T., Nakamura H., Sarai A., Ishii S., Nishimura Y. Solution structure of a DNA-binding unit of Myb: a helix-turn-helix-related motif with conserved tryptophans forming a hydrophobic core. Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6428–6432. doi: 10.1073/pnas.89.14.6428. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Paz-Ares J., Ghosal D., Wienand U., Peterson P. A., Saedler H. The regulatory c1 locus of Zea mays encodes a protein with homology to myb proto-oncogene products and with structural similarities to transcriptional activators. EMBO J. 1987 Dec 1;6(12):3553–3558. doi: 10.1002/j.1460-2075.1987.tb02684.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Privalsky M. L., Sharif M., Yamamoto K. R. The viral erbA oncogene protein, a constitutive repressor in animal cells, is a hormone-regulated activator in yeast. Cell. 1990 Dec 21;63(6):1277–1286. doi: 10.1016/0092-8674(90)90423-c. [DOI] [PubMed] [Google Scholar]
  54. Ptashne M. How eukaryotic transcriptional activators work. Nature. 1988 Oct 20;335(6192):683–689. doi: 10.1038/335683a0. [DOI] [PubMed] [Google Scholar]
  55. Punyammalee B., Crabeel M., de Lannoy C., Perbal B., Glansdorff N. Two c-myb proteins differing by their aminotermini exhibit different transcriptional transactivation activities (yeast/reporter-effector system). Oncogene. 1991 Jan;6(1):11–19. [PubMed] [Google Scholar]
  56. Roussel M., Saule S., Lagrou C., Rommens C., Beug H., Graf T., Stehelin D. Three new types of viral oncogene of cellular origin specific for haematopoietic cell transformation. Nature. 1979 Oct 11;281(5731):452–455. doi: 10.1038/281452a0. [DOI] [PubMed] [Google Scholar]
  57. Sakura H., Kanei-Ishii C., Nagase T., Nakagoshi H., Gonda T. J., Ishii S. Delineation of three functional domains of the transcriptional activator encoded by the c-myb protooncogene. Proc Natl Acad Sci U S A. 1989 Aug;86(15):5758–5762. doi: 10.1073/pnas.86.15.5758. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Schena M., Freedman L. P., Yamamoto K. R. Mutations in the glucocorticoid receptor zinc finger region that distinguish interdigitated DNA binding and transcriptional enhancement activities. Genes Dev. 1989 Oct;3(10):1590–1601. doi: 10.1101/gad.3.10.1590. [DOI] [PubMed] [Google Scholar]
  59. 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]
  60. Schiestl R. H., Gietz R. D. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier. Curr Genet. 1989 Dec;16(5-6):339–346. doi: 10.1007/BF00340712. [DOI] [PubMed] [Google Scholar]
  61. Seed B., Sheen J. Y. A simple phase-extraction assay for chloramphenicol acyltransferase activity. Gene. 1988 Jul 30;67(2):271–277. doi: 10.1016/0378-1119(88)90403-9. [DOI] [PubMed] [Google Scholar]
  62. Shen-Ong G. L. The myb oncogene. Biochim Biophys Acta. 1990 Jun 1;1032(1):39–52. doi: 10.1016/0304-419x(90)90011-o. [DOI] [PubMed] [Google Scholar]
  63. Stober-Grässer U., Brydolf B., Bin X., Grässer F., Firtel R. A., Lipsick J. S. The Myb DNA-binding domain is highly conserved in Dictyostelium discoideum. Oncogene. 1992 Mar;7(3):589–596. [PubMed] [Google Scholar]
  64. Thomas B. J., Rothstein R. Elevated recombination rates in transcriptionally active DNA. Cell. 1989 Feb 24;56(4):619–630. doi: 10.1016/0092-8674(89)90584-9. [DOI] [PubMed] [Google Scholar]
  65. Tice-Baldwin K., Fink G. R., Arndt K. T. BAS1 has a Myb motif and activates HIS4 transcription only in combination with BAS2. Science. 1989 Nov 17;246(4932):931–935. doi: 10.1126/science.2683089. [DOI] [PubMed] [Google Scholar]
  66. Todokoro K., Watson R. J., Higo H., Amanuma H., Kuramochi S., Yanagisawa H., Ikawa Y. Down-regulation of c-myb gene expression is a prerequisite for erythropoietin-induced erythroid differentiation. Proc Natl Acad Sci U S A. 1988 Dec;85(23):8900–8904. doi: 10.1073/pnas.85.23.8900. [DOI] [PMC free article] [PubMed] [Google Scholar]
  67. Westin E. H., Gallo R. C., Arya S. K., Eva A., Souza L. M., Baluda M. A., Aaronson S. A., Wong-Staal F. Differential expression of the amv gene in human hematopoietic cells. Proc Natl Acad Sci U S A. 1982 Apr;79(7):2194–2198. doi: 10.1073/pnas.79.7.2194. [DOI] [PMC free article] [PubMed] [Google Scholar]
  68. Weston K., Bishop J. M. Transcriptional activation by the v-myb oncogene and its cellular progenitor, c-myb. Cell. 1989 Jul 14;58(1):85–93. doi: 10.1016/0092-8674(89)90405-4. [DOI] [PubMed] [Google Scholar]

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