Apoptosis

, Volume 12, Issue 12, pp 2135–2142 | Cite as

ROS production by adrenodoxin does not cause apoptosis in fission yeast

  • Evi Derouet-Hümbert
  • Călin-Aurel Drăgan
  • Tarek Hakki
  • Matthias Bureik
Original Paper
First Online:

Abstract

We previously showed that production of reactive oxygen species (ROS) caused by overexpression of the mitochondrial electron transfer protein adrenodoxin (Adx) induces apoptosis in mammalian cells. In the fission yeast Schizosaccharomyces pombe, ROS are also produced in cells that undergo an apoptotic-like cell death, but it is not yet clear whether they are actually causative for this phenomenon or whether they are merely produced as a by-product. Therefore, the purpose of this study was to trigger mitochondrial ROS production in fission yeast by overexpression of either wildtype Adx (Adx-WT) or of several activated Adx mutants and to investigate its consequences. It was found that strong expression of either Adx-WT or Adx-S112W did not produce any ROS, while Adx-D113Y caused a twofold and Adx1–108 a threefold increase in ROS formation as compared to basal levels. However, no typical apoptotic markers or decreased viability could be observed in these strains. Since we previously observed that an increase in mitochondrial ROS formation of about 60% above basal levels is sufficient to strongly induce apoptosis in mammalian cells, we conclude that S. pombe is either very robust to mitochondrial ROS production or does not undergo apoptotic cell death in response to mitochondrial ROS at all.

Keywords

Adrenodoxin Apoptosis Cytochrome P450 system Fission yeast Mitochondria Reactive oxygen species 
This is a preview of subscription content, log in to check access.

Notes

Acknowledgements

This work was in part supported by a grant of the Bundesministerium für Bildung und Forschung (BMBF) to M.B. (0312641). E. D.-H. is supported by a scholarship from the Marianne und Dr. Fritz Walter Fischer-Stiftung (T192/13359/2003/sm). We thank Katja Simon for technical help.

References

  1. 1.
    Fadeel B, Orrenius S (2005) Apoptosis: a basic biological phenomenon with wide-ranging implications in human disease. J Intern Med 258:479–517PubMedCrossRefGoogle Scholar
  2. 2.
    Frohlich KU, Fussi H, Ruckenstuhl C (2007) Yeast apoptosis – from genes to pathways. Semin Cancer Biol 17:112–121PubMedCrossRefGoogle Scholar
  3. 3.
    Cairns J (2002) A DNA damage checkpoint in Escherichia coli. DNA Repair (Amst) 1:699–701CrossRefGoogle Scholar
  4. 4.
    Ink B, Zornig M, Baum B et al (1997) Human Bak induces cell death in Schizosaccharomyces pombe with morphological changes similar to those with apoptosis in mammalian cells. Mol Cell Biol 17:2468–2474PubMedGoogle Scholar
  5. 5.
    Burhans WC, Weinberger M, Marchetti MA et al (2003) Apoptosis-like yeast cell death in response to DNA damage and replication defects. Mutat Res 532:227–243PubMedGoogle Scholar
  6. 6.
    Marchetti MA, Weinberger M, Murakami Y, Burhans WC, Huberman JA (2006) Production of reactive oxygen species in response to replication stress and inappropriate mitosis in fission yeast. J Cell Sci 119:124–131PubMedCrossRefGoogle Scholar
  7. 7.
    Poon B, Jowett JB, Stewart SA et al (1997) Human immunodeficiency virus type 1 vpr gene induces phenotypic effects similar to those of the DNA alkylating agent, nitrogen mustard. J Virol 71:3961–3971PubMedGoogle Scholar
  8. 8.
    Somasundaran M, Sharkey M, Brichacek B et al (2002) Evidence for a cytopathogenicity determinant in HIV-1 Vpr. Proc Natl Acad Sci USA 99:9503–9508PubMedCrossRefGoogle Scholar
  9. 9.
    Le Rouzic E, Benichou S (2005) The Vpr protein from HIV-1: distinct roles along the viral life cycle. Retrovirology 2:11PubMedCrossRefGoogle Scholar
  10. 10.
    Zhao Y, Cao J, O’Gorman MR, Yu M, Yogev R (1996) Effect of human immunodeficiency virus type 1 protein R (vpr) gene expression on basic cellular function of fission yeast Schizosaccharomyces pombe. J Virol 70:5821–5826PubMedGoogle Scholar
  11. 11.
    Zhao Y, Yu M, Chen M et al (1998) Pleiotropic effects of HIV-1 protein R (Vpr) on morphogenesis and cell survival in fission yeast and antagonism by pentoxifylline. Virology 246:266–276PubMedCrossRefGoogle Scholar
  12. 12.
    Benko Z, Liang D, Agbottah E et al (2004) Anti-Vpr activity of a yeast chaperone protein. J Virol 78:11016–11029PubMedCrossRefGoogle Scholar
  13. 13.
    Benko Z, Liang D, Agbottah E et al (2007) Antagonistic interaction of HIV-1 Vpr with Hsf-mediated cellular heat shock response and Hsp16 in fission yeast (Schizosaccharomyces pombe). Retrovirology 4:16PubMedCrossRefGoogle Scholar
  14. 14.
    Zelivianski S, Liang D, Chen M, Mirkin BL, Zhao RY (2006) Suppressive effect of elongation factor 2 on apoptosis induced by HIV-1 viral protein R. Apoptosis 11:377–388PubMedCrossRefGoogle Scholar
  15. 15.
    Rodriguez-Menocal L, D’Urso G (2004) Programmed cell death in fission yeast. FEMS Yeast Res 5:111–117PubMedCrossRefGoogle Scholar
  16. 16.
    Gould KL, Nurse P (1989) Tyrosine phosphorylation of the fission yeast cdc2+ protein kinase regulates entry into mitosis. Nature 342:39–45PubMedCrossRefGoogle Scholar
  17. 17.
    Castedo M, Perfettini JL, Roumier T, Kroemer G (2002) Cyclin-dependent kinase-1: linking apoptosis to cell cycle and mitotic catastrophe. Cell Death Differ 9:1287–1293PubMedCrossRefGoogle Scholar
  18. 18.
    Castedo M, Perfettini JL, Roumier T et al (2004) Cell death by mitotic catastrophe: a molecular definition. Oncogene 23:2825–2837PubMedCrossRefGoogle Scholar
  19. 19.
    Fisher A. (1988) Intracellular production of oxygen-derived free radicals. In: Halliwell B (ed) Oxygen radicals, tissue injury. FASEB Publishing, Bethesda, MD, pp 34–39Google Scholar
  20. 20.
    Derouet-Hümbert E, Römer K, Bureik M (2005) Adrenodoxin (Adx) and CYP11A1 (P450scc) induce apoptosis by the generation of reactive oxygen species in mitochondria. Biol Chem 386:453–461PubMedCrossRefGoogle Scholar
  21. 21.
    Eisenberg T, Buttner S, Kroemer G, Madeo F (2007) The mitochondrial pathway in yeast apoptosis. Apoptosis 12:1011–1023PubMedCrossRefGoogle Scholar
  22. 22.
    Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Press, Cold Spring Harbor, NYGoogle Scholar
  23. 23.
    Alfa C, Fantes P, Hyams J, McLeod M, Warbrick E (1993) Experiments with fission yeast. A laboratory course manual. Cold Spring Harbor Press, Cold Spring Harbor, NYGoogle Scholar
  24. 24.
    Moreno S, Klar A, Nurse P (1991) Molecular genetic analysis of fission yeast Schizosaccharomyces pombe. Meth Enzymol 194:795–823PubMedCrossRefGoogle Scholar
  25. 25.
    Bureik M, Schiffler B, Hiraoka Y, Vogel F, Bernhardt R (2002) Functional expression of human mitochondrial CYP11B2 in fission yeast and identification of a new internal electron transfer protein, etp1. Biochemistry 41:2311–2321PubMedCrossRefGoogle Scholar
  26. 26.
    Maundrell K (1990) nmt1 of fission yeast. A highly transcribed gene completely repressed by thiamine. J Biol Chem 265:10857–10864PubMedGoogle Scholar
  27. 27.
    Dragan C-A, Zearo S, Hannemann F, Bernhardt R, Bureik M (2005) Efficient conversion of 11-deoxycortisol to cortisol (hydrocortisone) by recombinant fission yeast Schizosaccharomyces pombe. FEMS Yeast Res 5:621–625PubMedCrossRefGoogle Scholar
  28. 28.
    Hakki T, Zearo S, Dragan CA, Bureik M, Bernhardt R (in press) Coexpression of redox partners increases the hydrocortisone (cortisol) production efficiency in CYP11B1 expressing fission yeast Schizosaccharomyces pombe. J BiotechnolGoogle Scholar
  29. 29.
    Hannemann F, Bichet A, Ewen KM, Bernhardt R (2006) Cytochrome P450 systems-biological variations of electron transport chains. Biochim Biophys Acta 1770:330–344PubMedGoogle Scholar
  30. 30.
    Uhlmann H, Iametti S, Vecchio G, Bonomi F, Bernhardt R (1997) Pro108 is important for folding and stabilization of adrenal ferredoxin, but does not influence the functional properties of the protein. Eur J Biochem 248:897–902PubMedCrossRefGoogle Scholar
  31. 31.
    Schiffler B, Kiefer M, Wilken A et al (2001) The interaction of bovine adrenodoxin with CYP11A1 (cytochrome P450scc) and CYP11B1 (cytochrome P45011β). Acceleration of reduction and substrate conversion by site-directed mutagenesis of adrenodoxin. J Biol Chem 276:36225–36232PubMedCrossRefGoogle Scholar
  32. 32.
    Bichet A, Hannemann F, Rekowski M, Bernhardt R (2007) A new application of the yeast two-hybrid system in protein engineering. Protein Eng Des Sel 20:117–123PubMedCrossRefGoogle Scholar
  33. 33.
    Dragan CA, Blank LM, Bureik M (2006) Increased TCA cycle activity and reduced oxygen consumption during cytochrome P450-dependent biotransformation in fission yeast. Yeast 23:779–794PubMedCrossRefGoogle Scholar
  34. 34.
    Ehmer PB, Bureik M, Bernhardt R, Müller U, Hartmann RW (2002) Development of a test system for inhibitors of human aldosterone synthase (CYP11B2): screening in fission yeast and evaluation of selectivity in V79 cells. J Steroid Biochem Mol Biol 81:173–179PubMedCrossRefGoogle Scholar
  35. 35.
    Bureik M, Hübel K, Dragan C-A et al (2004) Development of test systems for the discovery of selective human aldosterone synthase (CYP11B2) and 11β-hydroxylase (CYP11B1) inhibitors. Discovery of a new lead compound for the therapy of congestive heart failure, myocardial fibrosis and hypertension. Mol Cell Endocrinol 217:249–254PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Evi Derouet-Hümbert
    • 1
  • Călin-Aurel Drăgan
    • 2
  • Tarek Hakki
    • 1
  • Matthias Bureik
    • 1
  1. 1.Department of Biochemistry, Building A 2-4Saarland UniversitySaarbruckenGermany
  2. 2.PomBioTech GmbHStarterzentrum der Universität des SaarlandesSaarbruckenGermany

Personalised recommendations