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. 1997 Apr 15;25(8):1493–1501. doi: 10.1093/nar/25.8.1493

c-Myc/Max heterodimers bind cooperatively to the E-box sequences located in the first intron of the rat ornithine decarboxylase (ODC) gene.

A J Walhout 1, J M Gubbels 1, R Bernards 1, P C van der Vliet 1, H T Timmers 1
PMCID: PMC146624  PMID: 9162900

Abstract

The oncoprotein c-Myc plays an important role in cell proliferation, transformation, inhibition of differentiation and apoptosis. These functions most likely result from the transcription factor activity of c-Myc. As a heterodimer with Max, the c-Myc protein binds to the E-box sequence (CACGTG), which is also recognized by USF dimers. In order to test differences in target gene recognition of c-Myc/Max, Max and USF dimers, we compared the DNA binding characteristics of these proteins in vitro using vaccinia viruses expressing full-length c-Myc and Max proteins. As expected, purified c-Myc/max binds specifically to a consensus E-box. The optimal conditions for DNA binding by either c-Myc/Max, Max or USF dimers differ with respect to ionic strength and Mg2+ ion concentration. Most interestingly, the c-Myc/Max complex binds with a high affinity to its natural target, the rat ODC gene, which contains two adjacent, consensus E-boxes. High affinity binding results from teh ability of c-Myc/Max dimers to bind cooperatively to these E-boxes. We propose that differential cooperative binding by E-box binding transcription factors could contribute to target gene specificity.

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Selected References

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

  1. Amati B., Brooks M. W., Levy N., Littlewood T. D., Evan G. I., Land H. Oncogenic activity of the c-Myc protein requires dimerization with Max. Cell. 1993 Jan 29;72(2):233–245. doi: 10.1016/0092-8674(93)90663-b. [DOI] [PubMed] [Google Scholar]
  2. Amati B., Dalton S., Brooks M. W., Littlewood T. D., Evan G. I., Land H. Transcriptional activation by the human c-Myc oncoprotein in yeast requires interaction with Max. Nature. 1992 Oct 1;359(6394):423–426. doi: 10.1038/359423a0. [DOI] [PubMed] [Google Scholar]
  3. Amati B., Littlewood T. D., Evan G. I., Land H. The c-Myc protein induces cell cycle progression and apoptosis through dimerization with Max. EMBO J. 1993 Dec 15;12(13):5083–5087. doi: 10.1002/j.1460-2075.1993.tb06202.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Amin C., Wagner A. J., Hay N. Sequence-specific transcriptional activation by Myc and repression by Max. Mol Cell Biol. 1993 Jan;13(1):383–390. doi: 10.1128/mcb.13.1.383. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ayer D. E., Eisenman R. N. A switch from Myc:Max to Mad:Max heterocomplexes accompanies monocyte/macrophage differentiation. Genes Dev. 1993 Nov;7(11):2110–2119. doi: 10.1101/gad.7.11.2110. [DOI] [PubMed] [Google Scholar]
  6. Ayer D. E., Kretzner L., Eisenman R. N. Mad: a heterodimeric partner for Max that antagonizes Myc transcriptional activity. Cell. 1993 Jan 29;72(2):211–222. doi: 10.1016/0092-8674(93)90661-9. [DOI] [PubMed] [Google Scholar]
  7. Ayer D. E., Lawrence Q. A., Eisenman R. N. Mad-Max transcriptional repression is mediated by ternary complex formation with mammalian homologs of yeast repressor Sin3. Cell. 1995 Mar 10;80(5):767–776. doi: 10.1016/0092-8674(95)90355-0. [DOI] [PubMed] [Google Scholar]
  8. Bello-Fernandez C., Packham G., Cleveland J. L. The ornithine decarboxylase gene is a transcriptional target of c-Myc. Proc Natl Acad Sci U S A. 1993 Aug 15;90(16):7804–7808. doi: 10.1073/pnas.90.16.7804. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Blackwell T. K., Kretzner L., Blackwood E. M., Eisenman R. N., Weintraub H. Sequence-specific DNA binding by the c-Myc protein. Science. 1990 Nov 23;250(4984):1149–1151. doi: 10.1126/science.2251503. [DOI] [PubMed] [Google Scholar]
  10. Blackwood E. M., Eisenman R. N. Max: a helix-loop-helix zipper protein that forms a sequence-specific DNA-binding complex with Myc. Science. 1991 Mar 8;251(4998):1211–1217. doi: 10.1126/science.2006410. [DOI] [PubMed] [Google Scholar]
  11. Carthew R. W., Chodosh L. A., Sharp P. A. An RNA polymerase II transcription factor binds to an upstream element in the adenovirus major late promoter. Cell. 1985 Dec;43(2 Pt 1):439–448. doi: 10.1016/0092-8674(85)90174-6. [DOI] [PubMed] [Google Scholar]
  12. Chin L., Schreiber-Agus N., Pellicer I., Chen K., Lee H. W., Dudast M., Cordon-Cardo C., DePinho R. A. Contrasting roles for Myc and Mad proteins in cellular growth and differentiation. Proc Natl Acad Sci U S A. 1995 Aug 29;92(18):8488–8492. doi: 10.1073/pnas.92.18.8488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Chodosh L. A., Carthew R. W., Sharp P. A. A single polypeptide possesses the binding and transcription activities of the adenovirus major late transcription factor. Mol Cell Biol. 1986 Dec;6(12):4723–4733. doi: 10.1128/mcb.6.12.4723. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Crouch D. H., Fisher F., Clark W., Jayaraman P. S., Goding C. R., Gillespie D. A. Gene-regulatory properties of Myc helix-loop-helix/leucine zipper mutants: Max-dependent DNA binding and transcriptional activation in yeast correlates with transforming capacity. Oncogene. 1993 Jul;8(7):1849–1855. [PubMed] [Google Scholar]
  15. Dang C. V., McGuire M., Buckmire M., Lee W. M. Involvement of the 'leucine zipper' region in the oligomerization and transforming activity of human c-myc protein. Nature. 1989 Feb 16;337(6208):664–666. doi: 10.1038/337664a0. [DOI] [PubMed] [Google Scholar]
  16. Desbarats L., Gaubatz S., Eilers M. Discrimination between different E-box-binding proteins at an endogenous target gene of c-myc. Genes Dev. 1996 Feb 15;10(4):447–460. doi: 10.1101/gad.10.4.447. [DOI] [PubMed] [Google Scholar]
  17. Eilers M., Schirm S., Bishop J. M. The MYC protein activates transcription of the alpha-prothymosin gene. EMBO J. 1991 Jan;10(1):133–141. doi: 10.1002/j.1460-2075.1991.tb07929.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Evan G. I., Hancock D. C. Studies on the interaction of the human c-myc protein with cell nuclei: p62c-myc as a member of a discrete subset of nuclear proteins. Cell. 1985 Nov;43(1):253–261. doi: 10.1016/0092-8674(85)90030-3. [DOI] [PubMed] [Google Scholar]
  19. Ferré-D'Amaré A. R., Pognonec P., Roeder R. G., Burley S. K. Structure and function of the b/HLH/Z domain of USF. EMBO J. 1994 Jan 1;13(1):180–189. doi: 10.1002/j.1460-2075.1994.tb06247.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Ferré-D'Amaré A. R., Prendergast G. C., Ziff E. B., Burley S. K. Recognition by Max of its cognate DNA through a dimeric b/HLH/Z domain. Nature. 1993 May 6;363(6424):38–45. doi: 10.1038/363038a0. [DOI] [PubMed] [Google Scholar]
  21. Galaktionov K., Chen X., Beach D. Cdc25 cell-cycle phosphatase as a target of c-myc. Nature. 1996 Aug 8;382(6591):511–517. doi: 10.1038/382511a0. [DOI] [PubMed] [Google Scholar]
  22. Gaubatz S., Imhof A., Dosch R., Werner O., Mitchell P., Buettner R., Eilers M. Transcriptional activation by Myc is under negative control by the transcription factor AP-2. EMBO J. 1995 Apr 3;14(7):1508–1519. doi: 10.1002/j.1460-2075.1995.tb07137.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Gaubatz S., Meichle A., Eilers M. An E-box element localized in the first intron mediates regulation of the prothymosin alpha gene by c-myc. Mol Cell Biol. 1994 Jun;14(6):3853–3862. doi: 10.1128/mcb.14.6.3853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Gregor P. D., Sawadogo M., Roeder R. G. The adenovirus major late transcription factor USF is a member of the helix-loop-helix group of regulatory proteins and binds to DNA as a dimer. Genes Dev. 1990 Oct;4(10):1730–1740. doi: 10.1101/gad.4.10.1730. [DOI] [PubMed] [Google Scholar]
  25. Gu W., Cechova K., Tassi V., Dalla-Favera R. Opposite regulation of gene transcription and cell proliferation by c-Myc and Max. Proc Natl Acad Sci U S A. 1993 Apr 1;90(7):2935–2939. doi: 10.1073/pnas.90.7.2935. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Halazonetis T. D., Kandil A. N. Determination of the c-MYC DNA-binding site. Proc Natl Acad Sci U S A. 1991 Jul 15;88(14):6162–6166. doi: 10.1073/pnas.88.14.6162. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Hateboer G., Timmers H. T., Rustgi A. K., Billaud M., van 't Veer L. J., Bernards R. TATA-binding protein and the retinoblastoma gene product bind to overlapping epitopes on c-Myc and adenovirus E1A protein. Proc Natl Acad Sci U S A. 1993 Sep 15;90(18):8489–8493. doi: 10.1073/pnas.90.18.8489. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Hurlin P. J., Quéva C., Koskinen P. J., Steingrímsson E., Ayer D. E., Copeland N. G., Jenkins N. A., Eisenman R. N. Mad3 and Mad4: novel Max-interacting transcriptional repressors that suppress c-myc dependent transformation and are expressed during neural and epidermal differentiation. EMBO J. 1995 Nov 15;14(22):5646–5659. doi: 10.1002/j.1460-2075.1995.tb00252.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Jones R. M., Branda J., Johnston K. A., Polymenis M., Gadd M., Rustgi A., Callanan L., Schmidt E. V. An essential E box in the promoter of the gene encoding the mRNA cap-binding protein (eukaryotic initiation factor 4E) is a target for activation by c-myc. Mol Cell Biol. 1996 Sep;16(9):4754–4764. doi: 10.1128/mcb.16.9.4754. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Kato G. J., Barrett J., Villa-Garcia M., Dang C. V. An amino-terminal c-myc domain required for neoplastic transformation activates transcription. Mol Cell Biol. 1990 Nov;10(11):5914–5920. doi: 10.1128/mcb.10.11.5914. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Kerkhoff E., Bister K., Klempnauer K. H. Sequence-specific DNA binding by Myc proteins. Proc Natl Acad Sci U S A. 1991 May 15;88(10):4323–4327. doi: 10.1073/pnas.88.10.4323. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Kretzner L., Blackwood E. M., Eisenman R. N. Myc and Max proteins possess distinct transcriptional activities. Nature. 1992 Oct 1;359(6394):426–429. doi: 10.1038/359426a0. [DOI] [PubMed] [Google Scholar]
  33. Lemaître J. M., Bocquet S., Thierry N., Buckle R., Méchali M. Production of a functional full-length Xenopus laevis c-Myc protein in insect cells. Gene. 1994 Dec 15;150(2):325–330. doi: 10.1016/0378-1119(94)90446-4. [DOI] [PubMed] [Google Scholar]
  34. Li L. H., Nerlov C., Prendergast G., MacGregor D., Ziff E. B. c-Myc represses transcription in vivo by a novel mechanism dependent on the initiator element and Myc box II. EMBO J. 1994 Sep 1;13(17):4070–4079. doi: 10.1002/j.1460-2075.1994.tb06724.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Littlewood T. D., Evan G. I. Transcription factors 2: helix-loop-helix. Protein Profile. 1994;1(6):635–709. [PubMed] [Google Scholar]
  36. Maheswaran S., Lee H., Sonenshein G. E. Intracellular association of the protein product of the c-myc oncogene with the TATA-binding protein. Mol Cell Biol. 1994 Feb;14(2):1147–1152. doi: 10.1128/mcb.14.2.1147. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Marcu K. B., Bossone S. A., Patel A. J. myc function and regulation. Annu Rev Biochem. 1992;61:809–860. doi: 10.1146/annurev.bi.61.070192.004113. [DOI] [PubMed] [Google Scholar]
  38. Meichle A., Philipp A., Eilers M. The functions of Myc proteins. Biochim Biophys Acta. 1992 Dec 16;1114(2-3):129–146. doi: 10.1016/0304-419x(92)90011-m. [DOI] [PubMed] [Google Scholar]
  39. Moss B., Elroy-Stein O., Mizukami T., Alexander W. A., Fuerst T. R. Product review. New mammalian expression vectors. Nature. 1990 Nov 1;348(6296):91–92. doi: 10.1038/348091a0. [DOI] [PubMed] [Google Scholar]
  40. Nakano R., Zhao L. J., Padmanabhan R. Overproduction of adenovirus DNA polymerase and preterminal protein in HeLa cells. Gene. 1991 Sep 15;105(2):173–178. doi: 10.1016/0378-1119(91)90148-5. [DOI] [PubMed] [Google Scholar]
  41. Packham G., Cleveland J. L. Ornithine decarboxylase is a mediator of c-Myc-induced apoptosis. Mol Cell Biol. 1994 Sep;14(9):5741–5747. doi: 10.1128/mcb.14.9.5741. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Packham G., Cleveland J. L. c-Myc and apoptosis. Biochim Biophys Acta. 1995 Jul 28;1242(1):11–28. doi: 10.1016/0304-419x(94)00015-t. [DOI] [PubMed] [Google Scholar]
  43. Papoulas O., Williams N. G., Kingston R. E. DNA binding activities of c-Myc purified from eukaryotic cells. J Biol Chem. 1992 May 25;267(15):10470–10480. [PubMed] [Google Scholar]
  44. Roy A. L., Carruthers C., Gutjahr T., Roeder R. G. Direct role for Myc in transcription initiation mediated by interactions with TFII-I. Nature. 1993 Sep 23;365(6444):359–361. doi: 10.1038/365359a0. [DOI] [PubMed] [Google Scholar]
  45. Roy A. L., Meisterernst M., Pognonec P., Roeder R. G. Cooperative interaction of an initiator-binding transcription initiation factor and the helix-loop-helix activator USF. Nature. 1991 Nov 21;354(6350):245–248. doi: 10.1038/354245a0. [DOI] [PubMed] [Google Scholar]
  46. Sawadogo M., Roeder R. G. Factors involved in specific transcription by human RNA polymerase II: analysis by a rapid and quantitative in vitro assay. Proc Natl Acad Sci U S A. 1985 Jul;82(13):4394–4398. doi: 10.1073/pnas.82.13.4394. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Sawadogo M., Van Dyke M. W., Gregor P. D., Roeder R. G. Multiple forms of the human gene-specific transcription factor USF. I. Complete purification and identification of USF from HeLa cell nuclei. J Biol Chem. 1988 Aug 25;263(24):11985–11993. [PubMed] [Google Scholar]
  48. Schreiber-Agus N., Chin L., Chen K., Torres R., Rao G., Guida P., Skoultchi A. I., DePinho R. A. An amino-terminal domain of Mxi1 mediates anti-Myc oncogenic activity and interacts with a homolog of the yeast transcriptional repressor SIN3. Cell. 1995 Mar 10;80(5):777–786. doi: 10.1016/0092-8674(95)90356-9. [DOI] [PubMed] [Google Scholar]
  49. Tobias K. E., Shor J., Kahana C. c-Myc and Max transregulate the mouse ornithine decarboxylase promoter through interaction with two downstream CACGTG motifs. Oncogene. 1995 Nov 2;11(9):1721–1727. [PubMed] [Google Scholar]
  50. Wu S., Peña A., Korcz A., Soprano D. R., Soprano K. J. Overexpression of Mxi1 inhibits the induction of the human ornithine decarboxylase gene by the Myc/Max protein complex. Oncogene. 1996 Feb 1;12(3):621–629. [PubMed] [Google Scholar]
  51. Wurm F. M., Gwinn K. A., Kingston R. E. Inducible overproduction of the mouse c-myc protein in mammalian cells. Proc Natl Acad Sci U S A. 1986 Aug;83(15):5414–5418. doi: 10.1073/pnas.83.15.5414. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Zervos A. S., Gyuris J., Brent R. Mxi1, a protein that specifically interacts with Max to bind Myc-Max recognition sites. Cell. 1993 Jan 29;72(2):223–232. doi: 10.1016/0092-8674(93)90662-a. [DOI] [PubMed] [Google Scholar]
  53. van Steeg H., van Oostrom C. T., Hodemaekers H. M., van Kreyl C. F. Cloning and functional analysis of the rat ornithine decarboxylase-encoding gene. Gene. 1990 Sep 14;93(2):249–256. doi: 10.1016/0378-1119(90)90232-g. [DOI] [PubMed] [Google Scholar]

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