Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 2003 Sep 15;374(Pt 3):723–729. doi: 10.1042/BJ20030032

Modulation of DNA topoisomerase II alpha promoter activity by members of the Sp (specificity protein) and NF-Y (nuclear factor Y) families of transcription factors.

Natisha Magan 1, Agnieszka P Szremska 1, Richard J Isaacs 1, Kathryn M Stowell 1
PMCID: PMC1223628  PMID: 12769819

Abstract

Topo IIalpha (topoisomerase IIalpha) is a major target of several commonly used anticancer drugs and is subject to down-regulation at the transcriptional level in some drug-resistant cell lines and tumours in response to chemotherapy. Clinical resistance to such drugs has been correlated with down-regulation of topo IIalpha at transcription in some drug-resistant cell lines and tumours. Putative binding sites for a variety of transcription factors, including Sp1 (specificity protein 1) and NF-Y (nuclear factor Y) have previously been identified in the topo IIalpha promoter, but their functional significance and interactions have not been described following exposure to anti-cancer drugs. The binding of these factors to specific putative regulatory elements in the topo IIalpha promoter was studied using electrophoretic-mobility-shift assays. Sp1 was found to bind strongly to both distal and proximal GC-rich elements and NF-Y to ICB1 (the first inverted CCAAT box). The functional significance of transcription-factor binding was studied using transient transfection of HeLa cells using a luciferase reporter driven by a 617-bp minimal promoter containing point mutations in putative regulatory elements. Sp1 and NF-Y were both found to be transcriptional modulators with activator or repressor functions depending on protein/DNA context. Moreover, a functional interaction between Sp1 and NF-Y bound at proximal elements was observed.

Full Text

The Full Text of this article is available as a PDF (215.2 KB).

Selected References

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

  1. Adachi N., Nomoto M., Kohno K., Koyama H. Cell-cycle regulation of the DNA topoisomerase IIalpha promoter is mediated by proximal CCAAT boxes: possible involvement of acetylation. Gene. 2000 Mar 7;245(1):49–57. doi: 10.1016/s0378-1119(00)00040-8. [DOI] [PubMed] [Google Scholar]
  2. Beck W. T., Danks M. K., Wolverton J. S., Kim R., Chen M. Drug resistance associated with altered DNA topoisomerase II. Adv Enzyme Regul. 1993;33:113–127. doi: 10.1016/0065-2571(93)90012-3. [DOI] [PubMed] [Google Scholar]
  3. Biggs J. R., Kudlow J. E., Kraft A. S. The role of the transcription factor Sp1 in regulating the expression of the WAF1/CIP1 gene in U937 leukemic cells. J Biol Chem. 1996 Jan 12;271(2):901–906. doi: 10.1074/jbc.271.2.901. [DOI] [PubMed] [Google Scholar]
  4. Falck J., Jensen P. B., Sehested M. Evidence for repressional role of an inverted CCAAT box in cell cycle-dependent transcription of the human DNA topoisomerase IIalpha gene. J Biol Chem. 1999 Jun 25;274(26):18753–18758. doi: 10.1074/jbc.274.26.18753. [DOI] [PubMed] [Google Scholar]
  5. Framson P., Bornstein P. A serum response element and a binding site for NF-Y mediate the serum response of the human thrombospondin 1 gene. J Biol Chem. 1993 Mar 5;268(7):4989–4996. [PubMed] [Google Scholar]
  6. Froelich-Ammon S. J., Osheroff N. Topoisomerase poisons: harnessing the dark side of enzyme mechanism. J Biol Chem. 1995 Sep 15;270(37):21429–21432. doi: 10.1074/jbc.270.37.21429. [DOI] [PubMed] [Google Scholar]
  7. Furukawa M., Uchiumi T., Nomoto M., Takano H., Morimoto R. I., Naito S., Kuwano M., Kohno K. The role of an inverted CCAAT element in transcriptional activation of the human DNA topoisomerase IIalpha gene by heat shock. J Biol Chem. 1998 Apr 24;273(17):10550–10555. doi: 10.1074/jbc.273.17.10550. [DOI] [PubMed] [Google Scholar]
  8. Hagen G., Müller S., Beato M., Suske G. Sp1-mediated transcriptional activation is repressed by Sp3. EMBO J. 1994 Aug 15;13(16):3843–3851. doi: 10.1002/j.1460-2075.1994.tb06695.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hasan S., Koda T., Kakinuma M. An upstream NF-Y-binding site is required for transcriptional activation from the hst promoter in F9 embryonal carcinoma cells. J Biol Chem. 1994 Oct 7;269(40):25042–25048. [PubMed] [Google Scholar]
  10. Herbomel P., Bourachot B., Yaniv M. Two distinct enhancers with different cell specificities coexist in the regulatory region of polyoma. Cell. 1984 Dec;39(3 Pt 2):653–662. doi: 10.1016/0092-8674(84)90472-0. [DOI] [PubMed] [Google Scholar]
  11. Ho S. N., Hunt H. D., Horton R. M., Pullen J. K., Pease L. R. Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene. 1989 Apr 15;77(1):51–59. doi: 10.1016/0378-1119(89)90358-2. [DOI] [PubMed] [Google Scholar]
  12. Hochhauser D., Stanway C. A., Harris A. L., Hickson I. D. Cloning and characterization of the 5'-flanking region of the human topoisomerase II alpha gene. J Biol Chem. 1992 Sep 15;267(26):18961–18965. [PubMed] [Google Scholar]
  13. Inoue T., Kamiyama J., Sakai T. Sp1 and NF-Y synergistically mediate the effect of vitamin D(3) in the p27(Kip1) gene promoter that lacks vitamin D response elements. J Biol Chem. 1999 Nov 5;274(45):32309–32317. doi: 10.1074/jbc.274.45.32309. [DOI] [PubMed] [Google Scholar]
  14. Isaacs R. J., Davies S. L., Sandri M. I., Redwood C., Wells N. J., Hickson I. D. Physiological regulation of eukaryotic topoisomerase II. Biochim Biophys Acta. 1998 Oct 1;1400(1-3):121–137. doi: 10.1016/s0167-4781(98)00131-6. [DOI] [PubMed] [Google Scholar]
  15. Isaacs R. J., Harris A. L., Hickson I. D. Regulation of the human topoisomerase IIalpha gene promoter in confluence-arrested cells. J Biol Chem. 1996 Jul 12;271(28):16741–16747. doi: 10.1074/jbc.271.28.16741. [DOI] [PubMed] [Google Scholar]
  16. Kubo T., Kohno K., Ohga T., Taniguchi K., Kawanami K., Wada M., Kuwano M. DNA topoisomerase II alpha gene expression under transcriptional control in etoposide/teniposide-resistant human cancer cells. Cancer Res. 1995 Sep 1;55(17):3860–3864. [PubMed] [Google Scholar]
  17. Ladias J. A., Hadzopoulou-Cladaras M., Kardassis D., Cardot P., Cheng J., Zannis V., Cladaras C. Transcriptional regulation of human apolipoprotein genes ApoB, ApoCIII, and ApoAII by members of the steroid hormone receptor superfamily HNF-4, ARP-1, EAR-2, and EAR-3. J Biol Chem. 1992 Aug 5;267(22):15849–15860. [PubMed] [Google Scholar]
  18. Li J. M., Nichols M. A., Chandrasekharan S., Xiong Y., Wang X. F. Transforming growth factor beta activates the promoter of cyclin-dependent kinase inhibitor p15INK4B through an Sp1 consensus site. J Biol Chem. 1995 Nov 10;270(45):26750–26753. doi: 10.1074/jbc.270.45.26750. [DOI] [PubMed] [Google Scholar]
  19. Mantovani R. The molecular biology of the CCAAT-binding factor NF-Y. Gene. 1999 Oct 18;239(1):15–27. doi: 10.1016/s0378-1119(99)00368-6. [DOI] [PubMed] [Google Scholar]
  20. Mastrangelo I. A., Courey A. J., Wall J. S., Jackson S. P., Hough P. V. DNA looping and Sp1 multimer links: a mechanism for transcriptional synergism and enhancement. Proc Natl Acad Sci U S A. 1991 Jul 1;88(13):5670–5674. doi: 10.1073/pnas.88.13.5670. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. McEwen D. G., Ornitz D. M. Regulation of the fibroblast growth factor receptor 3 promoter and intron I enhancer by Sp1 family transcription factors. J Biol Chem. 1998 Feb 27;273(9):5349–5357. doi: 10.1074/jbc.273.9.5349. [DOI] [PubMed] [Google Scholar]
  22. Mo Y. Y., Wang Q., Beck W. T. Down-regulation of topoisomerase IIalpha in CEM cells selected for merbarone resistance is associated with reduced expression of Sp3. Cancer Res. 1997 Nov 15;57(22):5004–5008. [PubMed] [Google Scholar]
  23. Ng S. W., Eder J. P., Schnipper L. E., Chan V. T. Molecular cloning and characterization of the promoter for the Chinese hamster DNA topoisomerase II alpha gene. J Biol Chem. 1995 Oct 27;270(43):25850–25858. doi: 10.1074/jbc.270.43.25850. [DOI] [PubMed] [Google Scholar]
  24. Pommier Y. DNA topoisomerase I and II in cancer chemotherapy: update and perspectives. Cancer Chemother Pharmacol. 1993;32(2):103–108. doi: 10.1007/BF00685611. [DOI] [PubMed] [Google Scholar]
  25. Reith W., Siegrist C. A., Durand B., Barras E., Mach B. Function of major histocompatibility complex class II promoters requires cooperative binding between factors RFX and NF-Y. Proc Natl Acad Sci U S A. 1994 Jan 18;91(2):554–558. doi: 10.1073/pnas.91.2.554. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Roder K. H., Wolf S. S., Schweizer M. Interaction of Sp1 and NF-Y in the diet-induced regulation of the rat fatty acid synthase (FAS) gene. Biochem Soc Trans. 1997 Feb;25(1):72S–72S. doi: 10.1042/bst025072s. [DOI] [PubMed] [Google Scholar]
  27. Roder K., Wolf S. S., Beck K. F., Schweizer M. Cooperative binding of NF-Y and Sp1 at the DNase I-hypersensitive site, fatty acid synthase insulin-responsive element 1, located at -500 in the rat fatty acid synthase promoter. J Biol Chem. 1997 Aug 22;272(34):21616–21624. doi: 10.1074/jbc.272.34.21616. [DOI] [PubMed] [Google Scholar]
  28. Ronchi A., Bellorini M., Mongelli N., Mantovani R. CCAAT-box binding protein NF-Y (CBF, CP1) recognizes the minor groove and distorts DNA. Nucleic Acids Res. 1995 Nov 25;23(22):4565–4572. doi: 10.1093/nar/23.22.4565. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sandri M. I., Hochhauser D., Ayton P., Camplejohn R. C., Whitehouse R., Turley H., Gatter K., Hickson I. D., Harris A. L. Differential expression of the topoisomerase II alpha and beta genes in human breast cancers. Br J Cancer. 1996 Jun;73(12):1518–1524. doi: 10.1038/bjc.1996.286. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Su W., Jackson S., Tjian R., Echols H. DNA looping between sites for transcriptional activation: self-association of DNA-bound Sp1. Genes Dev. 1991 May;5(5):820–826. doi: 10.1101/gad.5.5.820. [DOI] [PubMed] [Google Scholar]
  31. Wang J. C. DNA topoisomerases. Annu Rev Biochem. 1996;65:635–692. doi: 10.1146/annurev.bi.65.070196.003223. [DOI] [PubMed] [Google Scholar]
  32. Wang Q., Zambetti G. P., Suttle D. P. Inhibition of DNA topoisomerase II alpha gene expression by the p53 tumor suppressor. Mol Cell Biol. 1997 Jan;17(1):389–397. doi: 10.1128/mcb.17.1.389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Watt P. M., Hickson I. D. Structure and function of type II DNA topoisomerases. Biochem J. 1994 Nov 1;303(Pt 3):681–695. doi: 10.1042/bj3030681. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Yoon J. H., Kim J. K., Rha G. B., Oh M., Park S. H., Seong R. H., Hong S. H., Park S. D. Sp1 mediates cell proliferation-dependent regulation of rat DNA topoisomerase IIalpha gene promoter. Biochem J. 1999 Dec 1;344(Pt 2):367–374. [PMC free article] [PubMed] [Google Scholar]
  35. Zwicker J., Gross C., Lucibello F. C., Truss M., Ehlert F., Engeland K., Müller R. Cell cycle regulation of cdc25C transcription is mediated by the periodic repression of the glutamine-rich activators NF-Y and Sp1. Nucleic Acids Res. 1995 Oct 11;23(19):3822–3830. doi: 10.1093/nar/23.19.3822. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES