Skip to main content
SpringerLink
Log in

YeastSaccharomyces cerevisiae as a model system to study the cytotoxic activity of the antitumor drug camptothecin

  • Session 1: Topoisomerase Basic Science
  • Camptothecin, Drug Resistance, DNA Topoisomerase I
  • Published:
Cancer Chemotherapy and Pharmacology Aims and scope Submit manuscript

Abstract

Eukaryotic DNA topoisomerase I catalyzes the relaxation of positively and negatively supercoiled DNA and plays a critical role in processes involving DNA, such as DNA replication, transcription and recombination. The enzyme is encoded by theTOP1 gene and is highly conserved in its amino acid sequence and sensitivity to the anti-neoplatic agent camptothecin. This plant alkaloid specifically targets DNA topoisomerase I by reversibly stabilizing the covalent enzyme-DNA intermediate. Presumably, it is the interaction of these drug-stabilized adducts with other cellular components, such as replication forks, that actually produces the DNA lesions leading to cell death. A conservation of the mechanism(s) of camptothecin-induced cell killing is also implicit in studies of the yeastSaccharomyces cerevisiae, where the camptothecin sensitivity of ΔTOP1 yeast cells can be restored by plasmids expressing either yeast or humanTOP1 sequences. This genetically tractable system is currently being exploited to describe the specific molecular interactions required for the cytotoxic action of camptothecin. The results of mutational analyses of yeast and human DNA topoisomerase I are presented, as well as a genetic screen designed to identify genes, other thanTOP1, that are required for the cytotoxic activity of camptothecin.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Varshavsky A, Levinger L, Sundin O, Barsoum J, Ozkayanak E, Swerdlow P, Finley D (1983) Cellular and SV40 chromatin: replication, segregation, ubiquitination, nuclease-hypersensitive sites, HMG-containing nucleosomes, and heterochromatin-specific protein. Cold Spring Harbor Symp Quant Biol 47: 511–528

    Google Scholar 

  2. Wang JC (1985) DNA topoisomerases. Annu Rev Biochem 54:665–697

    Google Scholar 

  3. Vosberg P-H (1985) DNA topoisomerase enzymes that control DNA conformation. Curr Top Microbiol Immunol 114:19–102

    Google Scholar 

  4. Wang JC, Liu LF (1990) DNA replication: topological aspects and the roles of DNA topoisomerases: In: Cozzarelli NR, Wang JC (eds) DNA topology and its biological effects. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, p 321–340

    Google Scholar 

  5. Liu LF, Wang JC (1987) Supercoiling of the DNA template during transcription. Proc Natl Acad Sci USA 84: 7024–7027

    Google Scholar 

  6. Giaever GN, Wang JC (1989) Supercoiling of intracellular DNA can occur in eukaryotic cells. Cell 55: 849–856

    Google Scholar 

  7. Champoux JJ (1990) Mechanistic aspects of type-I topoisomerases. In: Cozzarelli NR, Wang JC (eds) DNA topology and its biological effects. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, pp 217–242

    Google Scholar 

  8. Hsieh T (1990) Mechanistic aspects of type-II topoisomerases. In: Cozzarelli NR, Wang JC (eds) DNA topology and its biological effects. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, pp 243–263

    Google Scholar 

  9. Tse T-C, Kirkegaard K, Wang JC (1980) Covalent bonds between protein and DNA. J Biol Chem 255: 5560–5565

    Google Scholar 

  10. Champoux JJ (1981) DNA is linked to the rat liver DNA nicking-closing enzyme by a phosphodiester bond to tyrosine. J Biol Chem 256: 4805–4809

    Google Scholar 

  11. Goto T, Wang JC (1985) Cloning of yeast TOP1, the gene encoding DNA topoisomerase I, and construction of mutants defective in both DNA topoisomerase I and DNA topoisomerase II. Proc Natl Acad Sci USA 82: 7178–7182

    Google Scholar 

  12. Thrash C, Bankier AT, Barrell BG, Sternglanz R (1985) Cloning, characterization, and sequence of the yeast DNA topoisomerase I gene. Proc Natl Acad Sci USA 82:4374–4378

    Google Scholar 

  13. Uemura T, Morino K, Uzawa S, Shiozaki K, Yanagida M (1987) Cloning and sequencing ofSchizosaccharomyces pombe DNA topoisomerase I gene, and effect of gene disruption. Nucleic Acids Res 15: 9727–9739

    Google Scholar 

  14. D'Arpa P, Machlin PS, Ratrie H, Rothfield NF, Cleveland DW, Earnshaw WC (1988) cDNA cloning of human DNA topoisomerase I: catalytic activity of 67.7-kDa carboxyl-terminal fragment. Proc Natl Acad Sci USA 85: 2543–2547

    Google Scholar 

  15. Hsieh T-S, Brown SD, Huang P, Fostel J (1992) Isolation and characterization of a gene encoding DNA topoisomerase I inDrosophila melanogaster Nucleic Acids Res 20: 6177–6182

    Google Scholar 

  16. Lynn RM, Bjornsti M-A, Caron P, Wang JC (1989) Peptide sequencing and site-directed mutagenesis identify tyrosine-727 as the active site tyrosine ofSaccharomyces cerevisiae DNA topoisomerase I. Proc Natl Acad Sci USA 86: 3559–3563

    Google Scholar 

  17. Eng W, Pandit SD, Sternglanz R (1989) Mapping of the active site tyrosine of eukaryotic DNA topoisomerase I. J Biol Chem 264: 13373–13376

    Google Scholar 

  18. Choder M (1991) A general topoisomerase I-dependent transcriptional repression in the stationary phase in yeast. Genes Dev 5: 2315–2326

    Google Scholar 

  19. Merino A, Madden KR, Lane WS, Champoux JJ, Reinberg D (1993) DNA topoisomerase I is involved in both repression and activation of transcription. Nature 365: 227–232

    Google Scholar 

  20. Brill SJ, Sternglanz R (1988) Transcription-dependent DNA supercoiling in yeast DNA topoisomerase mutants. Cell 54: 403–411

    Google Scholar 

  21. Christman MF, Dietrich FS, Fink G (1988) Mitotic recombination in the rDNA ofS. cerevisiae is suppressed by the combined action of DNA topoisomerases I and II. Cell 55: 413–425

    Google Scholar 

  22. Kim RA, Wang JC (1989) A subthreshold level of DNA topoisomerases leads to the excision of yeast rDNA as extrachromosomal rings. Cell 39: 377–386

    Google Scholar 

  23. Ross WE (1985) DNA topoisomerases as targets for cancer therapy. Biochem Pharmacol 34: 4191–4195

    Google Scholar 

  24. Wang JC (1987) DNA topoisomerases: from a laboratory curiosity to a subject in cancer chemotherapy. Natl Cancer Inst Monogr 4: 3–6

    Google Scholar 

  25. Liu LF (1989) DNA topoisomerase poisons as antitumor drugs. Annu Rev Biochem 58: 351–375

    Google Scholar 

  26. Liu LF, Wang JC (1991) Biochemistry of DNA topoisomerases and their poisons. In: Potmesil M, Kohn KW (eds) DNA topoisomerases in cancer. Oxford University Press, Oxford, pp 13–22

    Google Scholar 

  27. Bjornsti M-A (1991) DNA topoisomerases. Curr Opin Struct Biol 1: 99–103

    Google Scholar 

  28. Hsiang Y-H, Hertzberg R, Hecht S, Liu LF (1985) Camptothecin induces protein-linked DNA breaks in mammalian DNA topoisomerase I. J Biol Chem 260: 14873–14878

    Google Scholar 

  29. Hertzberg RP, Caranfa MJ, Hecht SM (1989) On the mechanism of topoisomerase I inhibition by camptothecin: evidence for binding to an enzyme-DNA complex. Biochemistry 28: 4629–4638

    Google Scholar 

  30. Covey JM, Jaxel C, Kohn K, Pommier Y (1989) Protein-linked DNA strand breaks induced in mammalian cells by camptothecin, an inhibitor of topoisomerase I. Cancer Res 49: 5016–5022

    Google Scholar 

  31. Porter SE, Champoux JJ (1989) The basis for camptothecin enhancement of DNA breakage by eukaryotic topoisomerase I. Nucleic Acids Res 17: 8521–8532

    Google Scholar 

  32. Hsiang YH, Lihou MG, Liu LF (1989) Arrest of replication forks by drug-stabilized topoisomerase I-DNA cleavable complexes as a mechanism of cell killing by camptothecin. Cancer Res 49: 5077–5082

    Google Scholar 

  33. Holm C, Covet JM, Kerrigan D, Pommier Y (1989) Differential requirement of DNA replication for the cytotoxicity of DNA topoisomerase I and II inhibitors in Chinese hamster DC3F cells. Cancer Res 49: 6365–6368

    Google Scholar 

  34. D'Arpa P, Beardmore C, Liu LF (1990) Involvement of nucleic acid synthesis in cell killing mechanisms of topoisomerase poisons. Cancer Res 50: 6919–6924

    Google Scholar 

  35. Gallo RC, Whang-Peng J, Adamson RH (1971) Studies on the antitumor activity, mechanism of action and cell cycle effects of camptothecin. J Natl Cancer Inst 46: 789–795

    Google Scholar 

  36. Wall ME, Wani MC (1991) Chemistry and antitimor activity of camptothecins, In: Potmesil M, Kohn KW (eds) DNA topoisomerases in cancer. Oxford University Press, Oxford, pp 93–102

    Google Scholar 

  37. Pommier Y, Jaxel C, Heise CR, Kerrigan D, Kohn KW (1991) Structure-activity relationship of topoisomerase I inhibition by camptothecin derivatives: evidence for the existence of a ternary complex. In: Potmesil M, Kohn KW (eds) DNA topoisomerases in cancer. Oxford University Press, Oxford, pp 121–132

    Google Scholar 

  38. Crow RT, Crothers DM (1992) Structural modifications of camptothecin and effects on topoisomerase I inhibition. J Med Chem 35: 4160–4164

    Google Scholar 

  39. Nitiss J, Wang JC (1988) DNA topoisomerase-targeting antitumor drugs can be studied in yeast. Proc Natl Acad Sci USA 85:7501–7505

    Google Scholar 

  40. Eng W-K, Faucette L, Johnson RK, Sternglanz R (1988) Evidence that DNA topoisomerase I is necessary for the cytotoxic effects of camptothecin. Mol Pharmacol 34: 755–760

    Google Scholar 

  41. Bjornsti M-A, Benedetti P, Viglianti GA, Wang JC (1989) Expression of human DNA topoisomerase I in yeast cells lacking DNA topoisomerase I: restoration of sensitivity of the cells to the antitumor drug camptothecin. Cancer Res 49: 6318–6323

    Google Scholar 

  42. Benedetti P, Tsai-Pflugfelder M, Wang JC (1992) The use of yeast and yeast strains expressing human DNA topoisomerases in the study of anticancer drugs: In: Tsuruo T, Ogawa M, Carter SK (eds) Drug resistance as a biochemical target in cancer chemotherapy. Academic Press, San Diego, p 129–145

    Google Scholar 

  43. Benedetti P, Fiorani P, Capuani L, Wang JC (1993) Camptothecin resistance from a single mutation changing glycine 363 of human DNA topoisomerase I to cysteine. Cancer Res 53: 4343–4348

    Google Scholar 

  44. Knab AM, Fertala J, Bjornsti M-A (1993) Mechanisms of camptothecin resistance in yeast DNA topoisomerase I mutants. J Biol Chem 268: 22322–22330

    Google Scholar 

  45. Sherman F (1991) Getting started with yeast. Methods Enzymol 194: 1–21

    Google Scholar 

  46. Lee MP, Brown SD, Chen A, Hsieh T-S (1993) DNA topoisomerase I is essential inDrosophila melanogaster. Proc Natl Acad Sci USA 90: 6656–6660

    Google Scholar 

  47. Tamura K, Kohchi C, Yamada R, Ikeda T, Koiwai O, Patterson E, Keene J, Okada E, Kjeldsen K, Andoh T (1991) Molecular cloning of a cDNA of a camptothecin-resistant human DNA topoisomerase I and identification of mutation sites. Nucleic Acids Res 19: 69–75

    Google Scholar 

  48. Kubota N, Kanzawa F, Nishio K, Takeda Y, Ohmori T, Fujiwara Y, Terashima Y, Saijo N (1992) Detection of topoisomerase I gene mutation in CPT-11 resistant lung cancer cell line. Biochem Biophys Res Commun 188: 571–577

    Google Scholar 

  49. Tanizawa A, Bertrand R, Kohlhagen G Tabuchi A, Jenkins J, Pommier Y (1993) Cloning of Chinese hamster DNA topoisomerase I cDNA and identification of a single point mutation responsible for camptothecin resistance. J Biol Chem 268: 25463–25468

    Google Scholar 

  50. Shuman S, Kane EM, Morham SG (1989) Mapping the active-site tyrosine of vaccinia virus DNA topoisomerase I. Proc Natl Acad Sci USA 86: 9793–9797

    Google Scholar 

  51. Shuman S, Golder M, Moss B (1988) Characterization of a vaccinia virus DNA topoisomerase I expressed inEscherichia coli. J Biol Chem 263: 16401–16407

    Google Scholar 

  52. Levin NA, Bjornsti M-A, Fink GR (1993) A novel mutation in DNA topoisomerase I of yeast causes DNA damage and RAD9-dependent cell cycle arrest. Genetics 133: 799–847

    Google Scholar 

  53. Caron PR, Wang JC (1993) DNA topoisomerases as targets of therapeutics: a structural overview. In: Andoh T, Ikeda H Oguro M (eds) Molecular biology of DNA topoisomerases and its application to chemotherapy. CRC, Boca Raton, Florida, p 1–18

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Thomas Jefferson University, 233 South 10th Street, 19107, Philadelphia, PA, USA

    Mary-Ann Bjornsti & Anne M. Knab

  2. Istituto di Biologia Cellulare C.N.R., Viale Marx 43, I-00137, Rome, Italy

    Piero Benedetti

Authors
  1. Mary-Ann Bjornsti
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Anne M. Knab
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Piero Benedetti
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Bjornsti, MA., Knab, A.M. & Benedetti, P. YeastSaccharomyces cerevisiae as a model system to study the cytotoxic activity of the antitumor drug camptothecin. Cancer Chemother. Pharmacol. 34 (Suppl 1), S1–S5 (1994). https://doi.org/10.1007/BF00684856

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00684856

Key words

Search

Navigation