Abstract
Apoptosis, or programmed cell death, is common in a variety of eucaryotes, from unicellular protozoa to vertebrates. The ciliated protozoan Tetrahymena thermophila has a unique apoptosis-like nuclear death during conjugation, called programmed nuclear death. This death program involves nuclear condensation (pyknosis) and oligonucleosomal DNA fragmentation in the parental macronucleus. Subsequently, the condensed nucleus is entirely resorbed in the autophagosome. Here we demonstrate that caspase-8- and -9-like activity was detected, but no caspase-3-like activity, by in vitro assay during the nuclear resorption process, suggesting that caspase-like activity is associated with both programmed cell death and apoptosis-like nuclear death in Tetrahymena. The use of indicator dye to detect the loss of mitochondrial membrane potential suggested the uptake of mitochondria and the degenerating macronucleus by the autophagosome. An involvement of mitochondria in the programmed nuclear death is discussed.
Similar content being viewed by others

Log in or create a free account to read this content
Gain free access to this article, as well as selected content from this journal and more on nature.com
or
Abbreviations
- PND:
-
programmed nuclear death
- Ac-DEVD-pNA:
-
N-acetyl-Asp-Glu-Val-Asp-p-nitroaniline
- Ac-IETD-pNA:
-
N-acetyl-Ile-Glu-Thr-Asp-nitroaniline
- Ac-LEHD-pNA:
-
N-acetyl-Leu-Glu-His-Asp-p-nitroaniline
- Ac-DEVD-CHO:
-
N-acetyl-Asp-Glu-Val-Asp-aldehyde
- Ac-IETD-CHO:
-
N-acetyl-Ile-Glu-Thr-Asp-aldehyde
- Ac-LEHD-CHO:
-
N-acetyl-Leu-Glu-His-Asp-aldehyde
References
Raff MC (1992) Social controls on cell survival and cell death. Nature 356: 397–400
Vaux DL and Krsmeyer SJ (1999) Cell death in development. Cell 96: 245–254
Grüter MG (2000) Caspases: key players in programmed cell death. Curr. Opin. Struct. Biol. 10: 649–655
Nicholson DW and Thornberry NA (1997) Caspases: killer protease. Trends Biochem. Sci. 22: 299–306
Yuan J, Shaham S, Ledoux S, Ellis HM and Horvitz HR (1993) The C. elegans cell death gene ced-3 encodes a protein similar to mammalian interleukin-1 b-converting enzyme. Cell 75: 641–652
Cikala M, Wilm B, Hobmayer E, Böttger A and David CN (1999) Identification of caspases and apoptosis in the simple metazoan Hydra. Curr. Biol. 9: 959–962
Cornillon S, Foa C, Davoust J, Buonavista N, Gross JD and Golstein P (1994) Programmed cell death in Dictyostelium. J. Cell Sci. 107: 2691–2704
Ameisen JC, Idziorek T, Billaut-Mulot O, Loyens M, Tissier JP, Potentier A and Ouaissi A (1995) Apoptosis in a unicellular eukaryotes (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: 285–300
Welburn SC, Dale C, Ellis D, Beecroft R and Pearson TW (1996) Apoptosis in procyclic Trypanosoma brucei rhodiense in vitro. Cell Death Differ. 3: 229–236
Moreira ME, Del Portillo HA, Milder RV, Balanco JM and Barcinski MA (1996) Heat shock induction of apoptosis in promastigotes of the unicellular organism Leishmania (Leishmania) amazonesis. J. Cell Physiol. 167: 305–313
Das M, Mukherjee SB and Shaha C (2001) Hydrogen peroxide induces apoptosis-like death in Leishmania donovani promastigotes. J. Cell Sci. 114: 2461–2469
Vardi A, Berman-Frank I, Rozenberg T, Hadas O, Kaplan A and Levin A (1999) Programmed cell death of the dinoflagellate Peridinium gatunense is mediated by CO2 limitation and oxidative stress. Curr. Biol. 9: 1061–1064
Picot S, Burnod J, Bracchi V, Chumpitazi BFF and Ambroise-Thomas P (1997) Apoptosis related to chloroquine sensitivity of the human malaria parasite Plasmodium falciparum. Trans. R. Soc. Trop. Med. Hyg. 91: 590–591
Nasirudeen AMA, Tan KSW, Singh M and Yap EH (2001) Programmed cell death in a human intestinal parasite, Blastocystis hominis. Parasitology 123: 235–246
Christensen ST, Wheatly DN, Rasmussen MI and Rasmussen L (1995) Mechanisms controlling death, survival and proliferation in a model unicellular eukaryote Tetrahymena thermophila. Cell Death Differ. 2: 301–308
Christensen ST, Sørensen H, Beyer NH, Kristiansen K, Rasmussen L and Rasmussen MI (2001) Cell death in Tetrahymena thermophila: new observations on culture conditions. Cell Biol. Int. 25: 509–519
Christensen ST, Chemnitz J, Straarup EM, Kristiansen K, Wheatley DN and Rasmussen L (1998) Staurosporine-induced cell death Tetrahymena thermophila has mixed characteristics of both apoptotic and autophagic degeneration. Cell Biol. Int. 2: 591–598
Kovács P, Hegyesi H, Köhidai L, Nemes P and Csaba G (1999) Effect of C2 ceramide on the inositol phospholipid metabolism (uptake of 32P, 3H-serine and 3H-palmitic acid) and apoptosis-related morphological changes in Tetrahymena. Comp. Biochem. Physiol. C 122: 215–224
Jaso-Friedmann L, Leary III JH and Evans DL (2000) Role of nonspecific cytotoxic cells in the induction of programmed cell death of pathogenic protozoans: participation of the Fas ligand–Fas receptor system. Exp. Parasitol. 96: 75–88
Davis MC, Ward JG, Herrick G and Allis CD (1992) Programmed nuclear death: apoptotic-like degradation of specific nuclei in conjugating Tetrahymena. Dev. Biol. 154: 419–432
Martindale DW, Allis CD and Bruns PJ (1982) Conjugation in Tetrahymena thermophila; a temporal analysis of cytological stages. Exp. Cell Res. 140: 227–236
Orias E (1986) Ciliate conjugation. In The Molecular Biology of Ciliated Protozoa, Gall JG (ed) (San Diego: Academic Press) pp. 45–84
Weiske-Benner A and Eckert WA (1987) Differentiation of nuclear structure during the sexual cycle in Tetrahymena thermophila; II. Degeneration and autolysis of macro- and micronuclei. Differentiation 34: 1–12
Mpoke S and Wolfe J (1996) DNA digestion and chromatin condensation during nuclear death in Tetrahymena. Exp. Cell Res. 225: 357–365
Ejercito R and Wolfe J (1998) Caspase and nuclear death in Tetrahymena. Mol. Biol. Cell 9S: 245a
Nagata S (2000) Apoptotic DNA fragmentation. Exp. Cell Res. 256: 12–18
Thornberry NA, Rano TA, Peterson EP, Rasper DM, Timkey T, Garcia-Calvo M, Houtzager VM, Nordstorm PA, Roy A, Vaillancourt JP, Chapman KT and Nicholson DW (1997) A combinatorial approach defines specificities of members of the caspase family and granzyme B. J. Biol. Chem. 272: 17907–17911
Budijardjo I, Oliver H, Lutter M, Luo X and Wang X (1999) Biochemical pathways of caspase activation during apoptosis. Annu. Rev. Cell Dev. Biol. 15: 269–290
Du C, Fang M, Li Y, Li L and Wang X (2000) Smac, a mitochondrial protein that promotes cytochrome c-dependent caspase activation by eliminating IAP inhibition. Cell 102: 33–34
Verhagen AM, Ekert PG, Pakusch M, Silke J, Connolly LM, Reid GE, Moritz RL, Simpson RJ and Vaux DL (2000) Identification of DIABLO, a mammalian protein that promotes apoptosis by binding to and antagonizing IAP proteins. Cell 102: 43–53
Chai J, Du C, Wu JW, Kyin S, Wang X and Shi Y (2000) Structural and biochemical basis of apoptotic activation by Smac/DIABLO. Nature 406: 855–862
Schulze-Osthoff K, Ferrari D, Los M, Wesselborg S and Peter ME (1998) Apoptosis signaling by death receptors. Eur. J. Biochem. 254: 439–459
Olie RA, Durrieu F, Cornillon S, Loughran G, Gross J, Earnshaw WC and Golstein P (1998) Apparent caspase independence of programmed cell death in Dictyostelium. Curr. Biol. 8: 955–958
Uren AG, O'Rourke K, Aravind L, Pisabarro MT, Seshagiri S, Koonin EV and Dixit VM (2000) Identification of paracaspase and metacaspase: two ancient families of caspase-like proteins, one of which plays a key role in MALT lymphoma. Mol. Cell 6: 961–967
Loo GV, Gurp MV, Depuyd B, Srinivasula SM, Rodriguez I, Anemri ES, Gevaert K, Vandekerckhove J, Declercq W and Vandenabeele P (2002) The serine protease Omi/HtrA2 is released from mitochondria during apoptosis. Omi interacts with caspase-inhibitor XIAP and induces enhanced caspase activity. Cell Death Differ. 9: 20–26
Li LY, Luo X and Wang X (2001) Endonuclease G is an apoptotic DNase when released from mitochondria. Nature 412: 95–99
Ward JG, Davis MC, Allis CD and Herrick G (1995) Effects of nullisomic chromosome deficiencies on conjugation events in Tetrahymena thermophila: insufficiency of the parental macronucleus to direct postzygotic development. Genetics 140: 989–1005
Mpoke SS and Wolfe J (1997) Differential staining of apoptotic nuclei in living cells: application to macronuclear elimination in Tetrahymena. J. Histochem. Cytochem. 45: 675–683
Lu E and Wolfe J (2001) Lysosomal enzymes in the macronucleus of Tetrahymena during its apoptosis-like degradation. Cell Death Differ. 8: 289–297
Nilsson J (1984) On starvation-induced autophagy in Tetrahymena. Carlsberg Res. Commun. 49: 323–340
Arnoult D, Tatischeff I, Estaquier J, Girard M, Sureau F, Tissier JP, Grodet A, Dellinger M, Traincard F, Kahn A, Ameisen JC and 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: 3016–3030
Arnoult D, Akarid DA, Grodet A, Petit PX, Estaquier J and 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: 65–81
Kobayashi T and Endoh H (1998) Abortive conjugation induced by UV-B irradiation at meiotic prophase in Tetrahymena thermophila. Dev. Genet. 23: 151–157
Sambrook J and Russell DW (2001) Molecular Cloning: a Laboratory Manual, 3rd edn, (NY, Cold Spring Harbor: Cold Spring Harbor Laboratory Press)
Gurtu V, Kain SR and Zhang G (1997) Fluorometric and colorimetric detection of caspase activity associated with apoptosis. Anal. Biochem. 251: 98–102
Acknowledgements
We are very grateful to S Hoshina for caspase assay technical support and to S Sakurai for critical discussion and helpful suggestions.
Author information
Authors and Affiliations
Corresponding author
Additional information
Edited by H Ichijo
Rights and permissions
About this article
Cite this article
Kobayashi, T., Endoh, H. Caspase-like activity in programmed nuclear death during conjugation of Tetrahymena thermophila. Cell Death Differ 10, 634–640 (2003). https://doi.org/10.1038/sj.cdd.4401216
Received:
Revised:
Accepted:
Published:
Issue date:
DOI: https://doi.org/10.1038/sj.cdd.4401216
Keywords
This article is cited by
-
Sirtuin-mediated nuclear differentiation and programmed degradation in Tetrahymena
BMC Cell Biology (2011)
-
Role of apoptosis-inducing factor (AIF) in programmed nuclear death during conjugation in Tetrahymena thermophila
BMC Cell Biology (2010)
-
The effect of phosphoinositide 3-kinase inhibitors on programmed nuclear degradation in Tetrahymena and fate of surviving nuclei
Cell Death & Differentiation (2004)

