Advertisement

Histochemistry and Cell Biology

, Volume 120, Issue 2, pp 129–141 | Cite as

Hydrogen peroxide induces caspase activation and programmed cell death in the amitochondrial Tritrichomonas foetus

  • Rafael M. Mariante
  • Cinthya A. Guimarães
  • Rafael Linden
  • Marlene BenchimolEmail author
Original Paper

Abstract

Tritrichomonas foetus is an amitochondrial parasite protist which lacks typical eukaryote organelles such as mitochondria and peroxisomes, but possesses the hydrogenosome, a double-membrane-bound organelle that produces ATP. The cell death of amitochondrial organisms is poorly studied. In the present work, the cytotoxic effects of hydrogen peroxide on T. foetus and its participation on cell death were analyzed. We took advantage of several microscopy techniques, including videomicroscopy, light microscopy immunocytochemistry for detection of caspase activation, and scanning and transmission electron microscopy. We report here that in T. foetus: (1) H2O2 leads to loss of motility and induces cell death, (2) the dying cells exhibit some characteristics similar to those found during the death of other organisms, and (3) a caspase-like protein seems to be activated during the death process. Thus, we propose that, although T. foetus does not present mitochondria nor any known pathways of cell death, it is likely that it bears mechanisms of cell demise. T. foetus exhibits morphological and physiological alterations in response to H2O2 treatment. The hydrogenosome, a unique organelle which is supposed to share a common ancestral origin with mitochondria and has an important role in oxidative responses in trichomonads, is a candidate for participating in this event.

Keywords

Tritrichomonas foetus Programmed cell death Hydrogen peroxide Caspase-3-like protein Hydrogenosomes 

Abbreviations

TUNEL

Terminal deoxyribonucleotide transferase (TdT)-mediated dUTP nick-end labeling

PARP

Poly (ADP-ribose) polymerase

DAPI

4′,6-Diamidino-2-phenylindole dihydrochloride

Notes

Acknowledgements

The authors thank Dr. Anu Srinivasan (Idun Pharmaceuticals, USA) for kindly providing the CM1 polyclonal antibody and Dr. Jose Morgado Diaz for helping with the western blot experiments. This work was supported by CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico), PRONEX (Programa de Núcleo de Excelência), FAPERJ (Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro), CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior), and AUSU (Associação Universitária Santa Úrsula). C.A. Guimarães is an EMBO Post Doctoral Fellow. R. Linden is a fellow of the John Simon Guggenheim Foundation.

References

  1. Ameisen JC, Idziorek T, Billaut-Mulot O, Loyens M, Tissier J-P, Potentier A, Quaissi A (1995) Apoptosis in a unicellular eukaryote (Trypanosoma cruzi): implications for the evolutionary origin and role of programmed cell death in the control of cell proliferation, differentiation and survival. Cell Death Differ 2:285–300Google Scholar
  2. Arnoult D, Tatischeff I, Estaquier J, Girard M, Sureau F, Tissier JP, Grodet A, Dellinger M, Traincard F, Kahn A, Ameisen JC, Petit PX (2001) On the evolutionary conservation of the cell death pathway: mitochondrial release of an apoptosis-inducing factor during Dictyostelium discoideum cell death. Mol Biol Cell 12:3016–3030Google Scholar
  3. Arnoult D, Akarid K, Grodet A, Petit PX, Estaquier J, Ameisen JC (2002) On the evolution of programmed cell death: apoptosis of the unicellular eukaryote Leishmania major involves cysteine proteinase activation and mitochondrion permeabilization. Cell Death Differ 9:65–81CrossRefPubMedGoogle Scholar
  4. Benchimol M (1999) Hydrogenosome autophagy: an ultrastructural and cytochemical study. Biol Cell 91:165–174PubMedGoogle Scholar
  5. Benchimol M (2001) Hydrogenosome morphological variation induced by fibronectin and other drugs in Trichomonas vaginalis and Tritrichomonas foetus. Parasitol Res 87:215–222CrossRefPubMedGoogle Scholar
  6. Benchimol M, De Souza W (1983) Fine structure and cytochemistry of the hydrogenosome of Tritrichomonas foetus. J Protozool 30:422–425PubMedGoogle Scholar
  7. Benchimol M, Almeida JCA, de Souza W (1996a) Further studies on the organization of the hydrogenosome in Tritrichomonas foetus. Tissue Cell 28:287–299PubMedGoogle Scholar
  8. Benchimol M, Johnson PJ, de Souza W (1996b) Morphogenesis of the hydrogenosome: an ultrastructural study. Biol Cell 87:197–205PubMedGoogle Scholar
  9. Benchimol M, Ribeiro KC, Mariante RM, Alderete JF (2001) Structure and division of the Golgi complex in Trichomonas vaginalis and Trichomonas foetus. Eur J Cell Biol 80:593–607PubMedGoogle Scholar
  10. Brenner C, Kroemer G (2000) Apoptosis. Mitochondria: the death signal integrators. Science 289:1150–1151CrossRefPubMedGoogle Scholar
  11. Bui ETN, Bradley PJ, Johnson PJ (1996) A common evolutionary origin for mitochondria and hydrogenosomes. Proc Natl Acad Sci U S A 93:9651–9656CrossRefPubMedGoogle Scholar
  12. Chang HY, Yang X (2000) Proteases for cell suicide: functions and regulation of caspases. Microbiol Mol Biol Rev 64:821–846CrossRefPubMedGoogle Scholar
  13. Chiu R, Novikov L, Mukherjee S, Shields D (2002) A caspase cleavage fragment of p115 induces fragmentation of the Golgi apparatus and apoptosis. J Cell Biol 159:637–648CrossRefPubMedGoogle Scholar
  14. Chose O, Noel C, Gerbod D, Brenner C, Viscogliosi E, Roseto A (2002) A form of cell death with some features resembling apoptosis in the amitochondrial unicellular organism Trichomonas vaginalis. Exp Cell Res 276:32–39CrossRefPubMedGoogle Scholar
  15. Christensen ST, Wheatley DN, Rasmussen MI, Rasmussen L (1995) Mechanisms controlling death, survival and proliferation in a model unicellular eukaryote Tetrahymena thermophila. Cell Death Differ 2:301–308Google Scholar
  16. Christensen ST, Chemnitz J, Straarup EM, Kristiansen K, Wheatley DN, Rasmussen L (1998) Staurosporine-induced cell death in Tetrahymena thermophila has mixed characteristics of both apoptotic and autophagic degeneration. Cell Biol Int 22:591–598CrossRefPubMedGoogle Scholar
  17. Clarke PG (1990) Developmental cell death: morphological diversity and multiple mechanisms. Anat Embryol 181:195–213PubMedGoogle Scholar
  18. Clemens DL, Johnson PJ (2000) Failure to detect DNA in hydrogenosomes of Trichomonas vaginalis by nick translation and immunomicroscopy. Mol Biochem Parasitol 106:307–313CrossRefPubMedGoogle Scholar
  19. Clement MV, Pervaiz S (1999) Reactive oxygen intermediates regulate cellular response to apoptotic stimuli: an hypothesis. Free Radic Res 30:247–252PubMedGoogle Scholar
  20. Cohen GM (1997) Caspases: the executioners of apoptosis. Biochem J 326:1–16PubMedGoogle Scholar
  21. Cornillon S, Foa C, Davoust J, Buonavista N, Gross JD, Golstein P (1994) Programmed cell death in Dictyostelium. J Cell Sci 107:2691–2704PubMedGoogle Scholar
  22. Das M, Mukherjee SB, Shaha C (2001), Hydrogen peroxide induces apoptosis-like death in Leishmania donovani promastigotes. J Cell Sci 114:2461–2469PubMedGoogle Scholar
  23. Davis SR, Lushbaugh WB (1993) Oxidative stress and Trichomonas vaginalis: the effect of hydrogen peroxide in vitro. Am J Trop Med Hyg 48:480–487PubMedGoogle Scholar
  24. Diamond LS (1957) The establishment of various trichomonads of animals and man in axenic cultures. J Parasitol 43:488–490Google Scholar
  25. Dyall SD, Johnson PJ (2000) Origins of hydrogenosomes and mitochondria: evolution and organelle biogenesis. Curr Opin Microbiol 3:404–411CrossRefPubMedGoogle Scholar
  26. Embley TM, Horner DA, Hirt RP (1997) Anaerobic eukaryote evolution: hydrogenosomes as biochemically modified mitochondria. TREE 12:433–441Google Scholar
  27. Fahimi HD, Baumgart E, Beier K, Pill J, Hartig F, Volkl A (1993) Ultrastructural and biochemical aspects of peroxisome proliferation and biogenesis in different mammalian species. In: Gibson G, Lake B (eds) Peroxisomes, biology and importance in toxicology and medicine. Taylor and Francis, London, pp 395–424Google Scholar
  28. Germot A, Philippe H, Guyader HL (1996) Presence of a mitochondrial-type 70-kDa heat shock protein in Trichomonas vaginalis suggests a very early mitochondrial endosymbiosis in eukaryotes. Proc Natl Acad Sci U S A 93:14614–14617CrossRefPubMedGoogle Scholar
  29. Glausmann H, Ericsson JLE, Marzella L (1981) Mechanisms of intralysosomal degradation with special reference to autophagocytosis and heterophagocytosis of cell organelles. Int Rev Cytol 73:149–182PubMedGoogle Scholar
  30. Granger BL, Warwood SJ, Benchimol M, de Souza W (2000) Transient invagination of flagella by Tritrichomonas foetus. Parasitol Res 86:699–709PubMedGoogle Scholar
  31. Green DR, Reed JC (1998) Mitochondria and apoptosis. Science 281:1309–1312PubMedGoogle Scholar
  32. Hockenbery DM, Oltvai ZN, Yin XM, Milliman CL, Korsmeyer SJ (1993) Bcl-2 functions in an antioxidant pathway to prevent apoptosis. Cell 75:241–251PubMedGoogle Scholar
  33. Jacobson MD, Weil M, Raff MC (1997) Programmed cell death in animal development. Cell 88:347–354PubMedGoogle Scholar
  34. Johnson PJ, Lahti CJ, Bradley PJ (1993) Biogenesis of the hydrogenosome in the anaerobic protist Trichomonas vaginalis. J Parasitol 79:664–670PubMedGoogle Scholar
  35. Kothakota S, Azuma T, Reinhard C, Klippel A, Tang J, Chu K, McGarry TJ, Kirschner MW, Koths K, Kwiatkowski DJ, Williams LT (1997) Caspase-3-generated fragment of gelsolin: effector of morphological change in apoptosis. Science 278:294–298PubMedGoogle Scholar
  36. Kroemer G, Reed JC (2000) Mitochondrial control of cell death. Nat Med 6:513–519PubMedGoogle Scholar
  37. Kurland CG, Andersson SGE (1999) Origins of mitochondria and hydrogenosomes. Curr Opin Microbiol 2:535–541CrossRefPubMedGoogle Scholar
  38. Lane JD, Lucocq J, Pryde J, Barr FA, Woodman PG, Allan VJ, Lowe M (2002) Caspase-mediated cleavage of the stacking protein GRASP65 is required for Golgi fragmentation during apoptosis. J Cell Biol 156:495–509CrossRefPubMedGoogle Scholar
  39. Lazebnik YA, Takahashi A, Moir RD, Goldman RD, Poirier GG, Kaufmann SH, Earnshaw WC (1995) Studies of the lamin proteinase reveal multiple parallel biochemical pathways during apoptotic execution. Proc Natl Acad Sci U S A 92:9042–9046PubMedGoogle Scholar
  40. Lee N, Bertholet S, Debrabant A, Muller J, Duncan R, Nakhasi HL (2002) Programmed cell death in the unicellular protozoan parasite Leishmania. Cell Death Differ 9:53–64CrossRefPubMedGoogle Scholar
  41. Li J, Huang CY, Zheng RL, Cui KR, Li JF (2000) Hydrogen peroxide induces apoptosis in human hepatoma cells and alters cell redox status. Cell Biol Int 24:9–23CrossRefPubMedGoogle Scholar
  42. Lindmark DG, Müller M (1973) Hydrogenosome, a cytoplasmic organelle of the anaerobic flagellate Tritrichomonas foetus, and its role in pyruvate metabolism. J Biol Chem 248:7724–7728PubMedGoogle Scholar
  43. Lindmark DG, Müller M (1974) Superoxide dismutase in the anaerobic flagellates, Tritrichomonas foetus and Monocercomonas sp. J Biol Chem 249:4634–4637PubMedGoogle Scholar
  44. Lloyd D, Harris JC, Maroulis S, Biagini GA, Wadley RB, Turner MP, Edwards MR (2000) The microaerophilic flagellate Giardia intestinalis: oxygen and its reaction products collapse membrane potential and cause cytotoxicity. Microbiology 146:3109–3118PubMedGoogle Scholar
  45. Loeffler M, Kroemer G (2000) The mitochondrion in cell death control: certainties and incognita. Exp Cell Res 256:19–26CrossRefPubMedGoogle Scholar
  46. Madeo F, Frohlich E, Ligr M, Grey M, Sigrist SJ, Wolf DH, Frohlich K (1999) Oxygen stress: a regulator of apoptosis in yeast. J Cell Biol 145:757–767PubMedGoogle Scholar
  47. Mancini M, Machamer CE, Roy S, Nicholson DW, Thornberry NA, Casciola-Rosen LA, Rosen A (2000) Caspase-2 is localized at the Golgi complex and cleaves golgin-160 during apoptosis. J Cell Biol 149:603–612CrossRefPubMedGoogle Scholar
  48. Mattos A, Solé-Cava A, de Carli G, Benchimol M (1997) Fine structure and isozymic characterization of trichomonadids protozoa. Parasitol Res 83:290–295CrossRefPubMedGoogle Scholar
  49. Mauel J, Schnyder J, Baggiolini M (1984) Intracellular parasite killing induced by electron carriers. II. Correlation between parasite killing and the induction of oxidative events in macrophages. Mol Biochem Parasitol 13:97–110CrossRefPubMedGoogle Scholar
  50. Meier P, Finch A, Evan G (2000) Apoptosis in development. Nature 407:796–801Google Scholar
  51. Moreira ME, Del Portillo HA, Milder RV, Balanco JM, Barcinski MA (1996) Heat shock induction of apoptosis in promastigotes of the unicellular organism Leishmania (Leishmania) amazonensis. J Cell Physiol 167:305–313CrossRefPubMedGoogle Scholar
  52. Müller M (1973) Biochemical cytology of trichomonad flagellates. I. Subcellular localization of hydrolases, dehydrogenases, and catalase in Tritrichomonas foetus. J Cell Biol 57:453–474PubMedGoogle Scholar
  53. Müller M (1990) Structure. In: Honigberg BM (ed) Trichomonads parasitic in humans. Springer, Berlin Heidelberg New York, pp 5–35Google Scholar
  54. Müller M (1993) The hydrogenosome. J Gen Microbiol 139:2879–2889PubMedGoogle Scholar
  55. Müller M (1997) Evolutionary origins of trichomonad hydrogenosomes. Parasitol Today 13:166–167CrossRefGoogle Scholar
  56. Nabi ZF, Rabinovitch M (1984) Inhibition by superoxide dismutase and catalase of the damage of isolated Leishmania mexicana amazonensis by phenazine methosulfate. Mol Biochem Parasitol 10:297–303CrossRefPubMedGoogle Scholar
  57. Namura S, Zhu J, Fink K, Endres M, Srinivasan A, Tomaselli KJ, Yuan J, Moskowitz MA (1998) Activation and cleavage of caspase-3 in apoptosis induced by experimental cerebral ischemia. J Neurosci 18:3659–3668PubMedGoogle Scholar
  58. Page-Sharp M, Behm CA, Smith GD (1996) Tritrichomonas foetus and Trichomonas vaginalis: the pattern of inactivation of hydrogenase activity by oxygen and activities of catalase and ascorbate peroxidase. Microbiology 142:207–211PubMedGoogle Scholar
  59. Raff MC (1992) Social controls on cell survival and cell death. Nature 356:397–400Google Scholar
  60. Ridgley EL, Xiong Z, Ruben L (1999) Reactive oxygen species activate a Ca2+-dependent cell death pathway in the unicellular organism Trypanosoma brucei brucei. Biochem J 340:33–40CrossRefPubMedGoogle Scholar
  61. Roger A, Clark CG, Doolittle WF (1996) A possible mitochondrial gene in the early-branching amitochondriate protist Trichomonas vaginalis. Proc Natl Acad Sci U S A 93:14618–14622CrossRefPubMedGoogle Scholar
  62. Ryley JF (1955) Studies on the metabolism of the protozoa. 5. Metabolism of the parasitic flagellate Tritrichomonas foetus. Biochem J 59:361–369Google Scholar
  63. Sahara S, Aoto M, Eguchi Y, Imamoto N, Yoneda Y, Tsujimoto Y (1999) Acinus is a caspase-3-activated protein required for apoptotic chromatin condensation. Nature 401:168–173PubMedGoogle Scholar
  64. Schirmer RH, Schollhammer T, Eisenbrand G, Krauth-Siegel RL (1987) Oxidative stress as a defense mechanism against parasitic infections. Free Radic Res Commun 3:3–12PubMedGoogle Scholar
  65. Segovia M, Haramaty L, Berges JA, Falkowski PG (2003) Cell death in the unicellular chlorophyte Dunaliella tertiolecta. A hypothesis on the evolution of apoptosis in higher plants and metazoans. Plant Physiol 132:99–105CrossRefPubMedGoogle Scholar
  66. Sereno D, Holzmuller P, Mangot I, Cuny G, Ouaissi A, Lemesre JL (2001) Antimonial-mediated DNA fragmentation in Leishmania infantum amastigotes. Antimicrob Agents Chemother 45:2064–2069CrossRefPubMedGoogle Scholar
  67. Sesso A, Fujiwara DT, Jaeger M, Jaeger R, Li TC, Monteiro MM, Correa H, Ferreira MA, Schumacher RI, Belisario J, Kachar B, Chen EJ (1999) Structural elements common to mitosis and apoptosis. Tissue Cell 31:357–371CrossRefPubMedGoogle Scholar
  68. Slater AF, Stefan C, Nobel I, van den Dobbelsteen DJ, Orrenius S (1995) Signalling mechanisms and oxidative stress in apoptosis. Toxicol Lett 82/83:149–153Google Scholar
  69. Solbach W, Laskay T (2000) The host response to Leishmania infection. Adv Immunol 74:275–317PubMedGoogle Scholar
  70. Sperandio S, de Belle I, Bredesen DE (2000) An alternative, nonapoptotic form of programmed cell death. Proc Natl Acad Sci U S A 97:14376–14381PubMedGoogle Scholar
  71. Steller H (1995) Mechanisms and genes of cellular suicide. Science 267:1445–1449PubMedGoogle Scholar
  72. Stennicke HR, Salvesen GS (2000) Caspases: controlling intracellular signals by protease zymogen activation. Biochim Biophys Acta 1477:299–306PubMedGoogle Scholar
  73. Stroh C, Schulze-Osthoff K (1998) Death by a thousand cuts: an ever increasing list of caspase substrates. Cell Death Differ 5:997–1000PubMedGoogle Scholar
  74. Takahashi A, Alnemri ES, Lazebnik YA, Fernandes-Alnemri T, Litwack G, Moir RD, Goldman RD, Poirier GG, Kaufmann SH, Earnshaw WC (1996) Cleavage of lamin A by Mch2 alpha but not CPP32: multiple interleukin 1 beta-converting enzyme-related proteases with distinct substrate recognition properties are active in apoptosis. Proc Natl Acad Sci U S A 93:8395–8400CrossRefPubMedGoogle Scholar
  75. Uren AG, O'Rourke K, Aravind LA, Pisabarro MT, Seshagiri S, Koonin EV, Dixit VM (2000) Identification of paracaspases and metacaspases: two ancient families of caspase-like proteins, one of which plays a key role in MALT lymphoma. Mol Cell 6:961–967Google Scholar
  76. Vannier-Santos MA, Urbina J, Martini A, Neves A, de Souza W (1995) Alterations induced by the antifungal compounds ketoconadole and terbinafine in Leishmania. J Eukaryot Microbiol 42:337–346PubMedGoogle Scholar
  77. Vardi A, Berman-Frank I, Rozenberg T, Hadas O, Kaplan A, Levine A (1999) Programmed cell death of the dinoflagellate Peridinium gatunense is mediated by CO2 limitation and oxidative stress. Curr Biol 9:1061–1064PubMedGoogle Scholar
  78. Vaux DL, Korsmeyer J (1999) Cell death in development. Cell 96:245–254PubMedGoogle Scholar
  79. Vaux DL, Haecker G, Strasser A (1994) An evolutionary perspective on apoptosis. Cell 76:777–779PubMedGoogle Scholar
  80. Vollgraf U, Wegner M, Richter-Landsberg C (1999) Activation of AP-1 and nuclear factor-kappaB transcription factor is involved in hydrogen peroxide induced apoptotic cell death of oligodendrocytes. J Neurochem 73:2501–2509CrossRefPubMedGoogle Scholar
  81. Welburn SC, Dale C, Ellis D, Beecroft R, Pearson TW (1996) Apoptosis in procyclic Trypanosoma brucei rhodesiense in vitro. Cell Death Differ 3:229–236Google Scholar
  82. Wolf BB, Green DR (1999) Suicidal tendencies: apoptotic cell death by caspase family proteinases. J Biol Chem 274:20049–20052PubMedGoogle Scholar
  83. Wyllie AH, Golstein P (2001) More than one way to go. Proc Natl Acad Sci U S A 98:11–13CrossRefPubMedGoogle Scholar
  84. Yuan J, Shaham S, Ledoux S, Ellis HM, Horvitz HR (1993) The C. elegans cell death gene ced-3 encodes a protein similar to mammalian interleukin-1β-converting enzyme. Cell 75:641–652PubMedGoogle Scholar
  85. Zangger H, Mottram JC, Fasel N (2002) Cell death in Leishmania induced by stress and differentiation: programmed cell death or necrosis? Cell Death Differ 9:1126–1139Google Scholar

Copyright information

© Springer-Verlag 2003

Authors and Affiliations

  • Rafael M. Mariante
    • 2
    • 3
  • Cinthya A. Guimarães
    • 4
    • 5
  • Rafael Linden
    • 4
  • Marlene Benchimol
    • 1
    • 3
    Email author
  1. 1.Rua Jornalista Orlando DantasRio de JaneiroRJ. Brazil
  2. 2.Programa de Ciências MorfológicasUniversidade Federal do Rio de JaneiroRio de JaneiroBrazil
  3. 3.Laboratório de Ultraestrutura CelularUniversidade Santa ÚrsulaRio de JaneiroBrazil
  4. 4.Laboratório de NeurogêneseUniversidade Federal do Rio de JaneiroRio de JaneiroBrazil
  5. 5.Department of Biological Chemistry, Institute of Life SciencesThe Hebrew University of JerusalemJerusalemIsrael

Personalised recommendations