Skip to main content
SpringerLink
Log in

Mitochondrial cytochrome c release precedes transmembrane depolarisation and caspase-3 activation during ceramide-induced apoptosis of Jurkat T cells

  • Published:
Apoptosis Aims and scope Submit manuscript

Abstract

Whilst the role of ceramide, a second messenger of the sphingolipid family, in the initiation of receptor-mediated apoptosis is controversial, a growing body of evidence is emerging for a role of ceramide in the amplification of apoptosis via mitochondrial perturbations that culminate in the activation of execution caspases. Treatment of Jurkat T cells with the cell-permeable analog, C2-ceramide, resulted in the rapid onset of apoptosis as evidenced by Annexin V-FITC staining of externalised phosphatidylserine residues. Cells bearing this early apoptotic marker had a reduced mitochondrial transmembrane potential (ΔΨm) that was preceded by the release of cytochrome c from mitochondria. Subsequent activation of caspase-3 provides the link between these ceramide-induced mitochondrial changes and execution caspases that ultimately result in the physical destruction of the cell. Collectively these results demonstrate that ceramide signalling results in caspase-mediated apoptosis via mitochondrial cytochrome c release and are further supportive of the role of ceramide in the amplification of apoptosis.

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. Kolesnick RN, Haimovitz-Friedman A, Fuks Z. The sphingomyelin signal transduction pathway mediates apoptosis for tumor necrosis factor, Fas, and ionizing radiation. Biochem Cell Biol 1994; 72: 471–474.

    Google Scholar 

  2. Cifone MG, De Maria R, Roncaioli P, et al Apoptotic signaling through CD95 (Fas/Apo-1) activates an acidic sphingomyelinase. J Exp Med 1994; 180: 1547–1552.

    Google Scholar 

  3. Cifone MG, Roncaioli P, De Maria R, et al Multiple pathways originate at the Fas/APO-1 (CD95) receptor: Sequential involvement of phosphatidylcholine-specific phospholipase C and acidic sphingomyelinase in the propagation of the apoptotic signal. EMBO J 1995; 14: 5859–5868.

    Google Scholar 

  4. Tepper CG, Jayadev S, Liu B, et al Role for ceramide as an endogenous mediator of Fas-induced cytotoxicity. Proc Natl Acad Sci USA 1995; 92: 8443–8447.

    Google Scholar 

  5. Gulbins E, Bissonnette R, Mahboubi A, et al FAS-induced apoptosis is mediated via a ceramide-initiated RAS signaling pathway. Immunity 1995; 2: 341–351.

    Google Scholar 

  6. Gamen S, Marzo I, Anel A, Pineiro A, Naval J CPP32 inhibition prevents Fas-induced ceramide generation and apoptosis in human cells. FEBS Letters 1996; 390: 232–237.

    Google Scholar 

  7. Sillence DJ, Allan D Evidence against an early signalling role for ceramide in Fas-mediated apoptosis. Biochem J 1997; 324: 29–32.

    Google Scholar 

  8. Watts JD, Gu M, Patterson SD, Aebersold R, Polverino AJ On the complexities of ceramide changes in cells undergoing apoptosis: Lack of evidence for a second messenger function in apoptotic induction. Cell Death Differ 1999; 6: 105–114.

    Google Scholar 

  9. Watts JD, Gu M, Polverino AJ, Patterson SD, Aebersold R Fas-induced apoptosis of T cells occurs independently of ceramide generation. Proc Natl Acad Sci USA 1997; 94: 7292–7296.

    Google Scholar 

  10. Tepper AD, de Vries E, van Blitterswijk WJ, Borst J Ordering of ceramide formation, caspase activation, and mitochondrial changes during CD95-and DNA damage-induced apoptosis. [erratum appears in J Clin Invest 1999 May; 103(9): 1363]. J Clin Invest 1999; 103: 971–978.

    Google Scholar 

  11. Hsu SC, Wu CC, Luh TY, et al Apoptotic signal of Fas is not mediated by ceramide, Blood 1998; 91: 2658–2663.

    Google Scholar 

  12. Susin SA, Zamzami N, Castedo M, et al The central executioner of apoptosis: Multiple connections between protease activation and mitochondria in Fas/APO-1/CD95-and ceramide-induced apoptosis. J Exp Med 1997; 186: 25–37.

    Google Scholar 

  13. Gamen S, Anel A, Pineiro A, Naval J Caspases are the main executioners of Fas-mediated apoptosis, irrespective of the ceramide signalling pathway. Cell Death Differ 1998; 5: 241–249.

    Google Scholar 

  14. Obeid LM, Linardic CM, Karolak LA, Hannun YA Programmed cell death induced by ceramide. Science 1993; 259: 1769–1771.

    Google Scholar 

  15. Jarvis WD, Kolesnick RN, Fornari FA, et al Induction of apoptotic DNA damage and cell death by activation of the sphingomyelin pathway. Proc Natl Acad Sci USA 1994; 91: 73–77.

    Google Scholar 

  16. Ruvolo PP, Deng X, Ito T, Carr BK, May WS Ceramide induces Bcl2 dephosphorylation via a mechanism involving mitochondrial PP2A. J Biol Chem 1999; 274: 20296–20300.

    Google Scholar 

  17. Garcia-Ruiz C, Colell A, Mari M, Morales A, Fernandez-Checa JC Direct effect of ceramide on the mitochondrial electron transport chain leads to generation of reactive oxygen species. Role of mitochondrial glutathione. J Biol Chem 1997; 272: 11369–11377.

    Google Scholar 

  18. Arora AS, Jones BJ, Patel TC, Bronk SF, Gores GJ Ceramide induces hepatocyte cell death through disruption of mitochondrial function in the rat. Hepatology 1997; 25: 958–963.

    Google Scholar 

  19. Di Paola M, Cocco T, Lorusso M Ceramide interaction with the respiratory chain of heart mitochondria. Biochemistry 2000; 39: 6660–6668.

    Google Scholar 

  20. Ghafourifar P, Klein SD, Schucht O, et al Ceramide induces cytochrome c release from isolated mitochondria. Importance of mitochondrial redox state. J Biol Chem 1999; 274: 6080–6084.

    Google Scholar 

  21. Ito A, Uehara T, Tokumitsu A, Okuma Y, Nomura Y Possible involvement of cytochrome c release and sequential activation of caspases in ceramide-induced apoptosis in SK-N-MC cells. Biochim Biophys Acta 1999; 1452: 263–274.

    Google Scholar 

  22. Cuvillier O, Edsall L, Spiegel S Involvement of sphingosine in mitochondria-dependent Fas-induced apoptosis of type II JurkatTcells. Journal of Biological Chemistry 2000; 275: 15691–15700.

    Google Scholar 

  23. Bossy-Wetzel E, Green DR Caspases induce cytochrome c release from mitochondria by activating cytosolic factors. J Biol Chem 1999; 274: 17484–17490.

    Google Scholar 

  24. Waterhouse NJ, Green DR Mitochondria and apoptosis: HQ or high-security prison? J Clin Immunol 1999; 19: 378–387.

    Google Scholar 

  25. Lee HC, Wei YH Mitochondrial role in life and death of the cell. J Biomed Sci 2000; 7: 2–15.

    Google Scholar 

  26. Scarlett JL, Sheard PW, Hughes G, et al Changes in mitochondrial membrane potential during staurosporine-induced apoptosis in Jurkat cells. FEBS Lett 2000; 475: 267–272.

    Google Scholar 

  27. Amarante-Mendes GP, Naekyung Kim C, Liu L, et al Bcr-Abl exerts its antiapoptotic effect against diverse apoptotic stimuli through blockage of mitochondrial release of cytochrome c and activation of caspase-3. Blood 1998; 91: 1700–1705.

    Google Scholar 

  28. Zhang J, Alter N, Reed JC, et al Bcl-2 interrupts the ceramidemediated pathway of cell death. Proc Natl Acad Sci USA 1996; 93: 5325–5328.

    Google Scholar 

  29. Tepper AD, Ruurs P, Wiedmer T, et al Sphingomyelin hydrolysis to ceramide during the execution phase of apoptosis results from phospholipid scrambling and alters cell-surface morphology. [see comments]. J Cell Biol 2000; 150: 155–164.

    Google Scholar 

  30. Green DR Apoptosis and sphingomyelin hydrolysis. The flip side. J Cell Biol 2000; 150: F5-F7.

    Google Scholar 

  31. Annis MG, Zamzami N, Zhu W, et al Endoplasmic reticulum localized Bcl-2 prevents apoptosis when redistribution of cytochrome c is a late event. Oncogene 2001; 20: 1939–1952.

    Google Scholar 

  32. Pervaiz S, Seyed MA, Hirpara JL, Clement MV, Loh KW Purified photoproducts of merocyanine 540 trigger cytochrome c release and caspase 8-dependent apoptosis in human leukemia and melanoma cells. Blood 1999; 93: 4096–4108.

    Google Scholar 

  33. Strasser A, Harris AW, Huang DC, Krammer PH, Cory S Bcl-2 and Fas/APO-1 regulate distinct pathways to lymphocyte apoptosis. EMBO J 1995; 14: 6136–6147.

    Google Scholar 

  34. Sawada M, Nakashima S, Banno Y, et al Ordering of ceramide formation, caspase activation, and Bax/Bcl-2 expression during etoposide-induced apoptosis in C6 glioma cells. Cell Death Differ 2000; 7: 761–772.

    Google Scholar 

  35. Hannun YA, Luberto C Ceramide in the eukaryotic stress response. Trends Cell Biol 2000; 10: 73–80.

    Google Scholar 

  36. Kolesnick RN, Kronke M Regulation of ceramide production and apoptosis. Annu Rev Physiol 1998; 60: 643–665.

    Google Scholar 

  37. Mizushima N, Koike R, Kohsaka H, et al Ceramide induces apoptosis via CPP32 activation. FEBS Lett 1996; 395: 267–271.

    Google Scholar 

  38. Li P, Nijhawan D, Budihardjo I, et al Cytochrome c and dATPdependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade. Cell 1997; 91: 479–489.

    Google Scholar 

  39. Dbaibo GS, Perry DK, Gamard CJ, et al Cytokine response modifier A (CrmA) inhibits ceramide formation in response to tumor necrosis factor (TNF)-alpha: CrmA and Bcl-2 target distinct components in the apoptotic pathway. J Exp Med 1997; 185: 481–490.

    Google Scholar 

  40. Cuvillier O, Rosenthal DS, Smulson ME, Spiegel S Sphingosine 1-phosphate inhibits activation of caspases that cleave poly(ADP-ribose) polymerase and lamins during Fas-and ceramide-mediated apoptosis in Jurkat T lymphocytes. J Biol Chem 1998; 273: 2910–2916.

    Google Scholar 

  41. Chinnaiyan AM, Tepper CG, Seldin MF, et al FADD/MORT1 is a common mediator of CD95 (Fas/APO-1) and tumor necrosis factor receptor-induced apoptosis. J Biol Chem 1996; 271: 4961–4965.

    Google Scholar 

  42. Karasavvas N, Zakeri Z Relationships of apoptotic signaling mediated by ceramide and TNF-alpha in U937 cells. Cell Death Differ 1999; 6: 115–123.

    Google Scholar 

  43. Grullich C, Sullards MC, Fuks Z, Merrill AH Jr, Kolesnick R CD95(Fas/APO-1) signals ceramide generation independent of the effector stage of apoptosis. J Biol Chem 2000; 275: 8650–8656.

    Google Scholar 

  44. Jaffrezou JP, Levade T, Bettaieb A, et al Daunorubicin-induced apoptosis: Triggering of ceramide generation through sphingomyelin hydrolysis. EMBO J 1996; 15: 2417–2424.

    Google Scholar 

  45. Bose R, Verheij M, Haimovitz-Friedman A, et al Ceramide synthase mediates daunorubicin-induced apoptosis: An alternative mechanism for generating death signals. Cell 1995; 82: 405–414.

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Division of Medicine, University of Tasmania, GPO Box 252-34, Hobart, Tasmania, 7001, Australia

    A. C. Hearps, J. Burrows, C. E. Connor, R. M. Lowenthal & S. J. Ragg

  2. Division of Pathology, University of Tasmania, GPO Box 252-29, Hobart, Tasmania, 7001, Australia

    G. M. Woods

Authors
  1. A. C. Hearps
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. J. Burrows
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. C. E. Connor
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. G. M. Woods
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. R. M. Lowenthal
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. S. J. Ragg
    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

Hearps, A.C., Burrows, J., Connor, C.E. et al. Mitochondrial cytochrome c release precedes transmembrane depolarisation and caspase-3 activation during ceramide-induced apoptosis of Jurkat T cells. Apoptosis 7, 387–394 (2002). https://doi.org/10.1023/A:1020034906200

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1023/A:1020034906200

Search

Navigation