Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Inhibition of CED-3 zymogen activation and apoptosis in Caenorhabditis elegans by caspase homolog CSP-3

Nature Structural & Molecular Biology volume 15, pages 1094–1101 (2008)Cite this article

Abstract

Inhibitor of apoptosis (IAP) proteins have a crucial role in apoptosis, through negative regulation of caspases in species from fruitflies to mammals. In Caenorhabditis elegans, however, no IAP homolog or caspase inhibitor has been identified, calling into question how the cell-killing caspase CED-3 can be negatively regulated. Here we show that inactivation of the C. elegans csp-3 gene, which encodes a protein similar to the small subunit of the CED-3 caspase, causes cells that normally live to undergo apoptosis in a CED-3–dependent manner. Biochemical analysis reveals that CSP-3 associates with the large subunit of the CED-3 zymogen and inhibits zymogen autoactivation. However, CSP-3 does not block CED-3 activation induced by CED-4, nor does it inhibit the activity of the activated CED-3 protease. Therefore CSP-3 uses a previously unreported mechanism to protect cells from apoptosis.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: CSP-3 is a cytoplasmic protein with sequence homology to the small subunit of CED-3.
Figure 2: Loss of csp-3 results in increased cell deaths during embryonic development.
Figure 3: Loss of csp-3 causes cells that normally live to undergo programmed cell death.
Figure 4: CSP-3 associates with CED-3 in vitro and in C. elegans.
Figure 5: CSP-3 specifically inhibits the autoactivation of the CED-3 zymogen.

Similar content being viewed by others

References

  1. Steller, H. Mechanisms and genes of cellular suicide. Science 267, 1445–1449 (1995).

    Article  CAS  PubMed  Google Scholar 

  2. Thornberry, N.A. & Lazebnik, Y. Caspases: enemies within. Science 281, 1312–1316 (1998).

    Article  CAS  PubMed  Google Scholar 

  3. Boatright, K.M. & Salvesen, G.S. Mechanisms of caspase activation. Curr. Opin. Cell Biol. 15, 725–731 (2003).

    Article  CAS  PubMed  Google Scholar 

  4. Shi, Y. Mechanisms of caspase activation and inhibition during apoptosis. Mol. Cell 9, 459–470 (2002).

    Article  CAS  PubMed  Google Scholar 

  5. Budihardjo, I., Oliver, H., Lutter, M., Luo, X. & Wang, X. Biochemical pathways of caspase activation during apoptosis. Annu. Rev. Cell Dev. Biol. 15, 269–290 (1999).

    Article  CAS  PubMed  Google Scholar 

  6. Degterev, A., Boyce, M. & Yuan, J. A decade of caspases. Oncogene 22, 8543–8567 (2003).

    Article  CAS  PubMed  Google Scholar 

  7. Zou, H., Henzel, W.J., Liu, X., Lutschg, A. & Wang, X. APAF-1, a human protein homologous to C. elegans CED-4, participates in cytochrome c-dependent activation of caspase-3. Cell 90, 405–413 (1997).

    Article  CAS  PubMed  Google Scholar 

  8. Zou, H., Li, Y., Liu, X. & Wang, X. An APAF-1.cytochrome c multimeric complex is a functional apoptosome that activates procaspase-9. J. Biol. Chem. 274, 11549–11556 (1999).

    Article  CAS  PubMed  Google Scholar 

  9. Jiang, X. & Wang, X. Cytochrome c-mediated apoptosis. Annu. Rev. Biochem. 73, 87–106 (2004).

    Article  CAS  PubMed  Google Scholar 

  10. James, C., Gschmeissner, S., Fraser, A. & Evan, G.I. CED-4 induces chromatin condensation in Schizosaccharomyces pombe and is inhibited by direct physical association with CED-9. Curr. Biol. 7, 246–252 (1997).

    Article  CAS  PubMed  Google Scholar 

  11. Wu, D., Wallen, H.D. & Nunez, G. Interaction and regulation of subcellular localization of CED-4 by CED-9. Science 275, 1126–1129 (1997).

    Article  CAS  PubMed  Google Scholar 

  12. Chinnaiyan, A.M., O'Rourke, K., Lane, B.R. & Dixit, V.M. Interaction of CED-4 with CED-3 and CED-9: a molecular framework for cell death. Science 275, 1122–1126 (1997).

    Article  CAS  PubMed  Google Scholar 

  13. Chen, F. et al. Translocation of C. elegans CED-4 to nuclear membranes during programmed cell death. Science 287, 1485–1489 (2000).

    Article  CAS  PubMed  Google Scholar 

  14. Yan, N. et al. Structure of the CED-4–CED-9 complex provides insights into programmed cell death in Caenorhabditis elegans. Nature 437, 831–837 (2005).

    Article  CAS  PubMed  Google Scholar 

  15. Cory, S., Huang, D.C. & Adams, J.M. The Bcl-2 family: roles in cell survival and oncogenesis. Oncogene 22, 8590–8607 (2003).

    Article  CAS  PubMed  Google Scholar 

  16. Deveraux, Q.L. & Reed, J.C. IAP family proteins—suppressors of apoptosis. Genes Dev. 13, 239–252 (1999).

    Article  CAS  PubMed  Google Scholar 

  17. Deveraux, Q.L., Takahashi, R., Salvesen, G.S. & Reed, J.C. X-linked IAP is a direct inhibitor of cell-death proteases. Nature 388, 300–304 (1997).

    Article  CAS  PubMed  Google Scholar 

  18. Roy, N., Deveraux, Q.L., Takahashi, R., Salvesen, G.S. & Reed, J.C. The c-IAP-1 and c-IAP-2 proteins are direct inhibitors of specific caspases. EMBO J. 16, 6914–6925 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Deveraux, Q.L. et al. IAPs block apoptotic events induced by caspase-8 and cytochrome c by direct inhibition of distinct caspases. EMBO J. 17, 2215–2223 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Meier, P., Silke, J., Leevers, S.J. & Evan, G.I. The Drosophila caspase DRONC is regulated by DIAP1. EMBO J. 19, 598–611 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Hawkins, C.J., Wang, S.L. & Hay, B.A. A cloning method to identify caspases and their regulators in yeast: identification of Drosophila IAP1 as an inhibitor of the Drosophila caspase DCP-1. Proc. Natl. Acad. Sci. USA 96, 2885–2890 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Salvesen, G.S. & Duckett, C.S. IAP proteins: blocking the road to death's door. Nat. Rev. Mol. Cell Biol. 3, 401–410 (2002).

    Article  CAS  PubMed  Google Scholar 

  23. Callus, B.A. & Vaux, D.L. Caspase inhibitors: viral, cellular and chemical. Cell Death Differ. 14, 73–78 (2007).

    Article  CAS  PubMed  Google Scholar 

  24. Srinivasula, S.M. et al. A conserved XIAP-interaction motif in caspase-9 and Smac/DIABLO regulates caspase activity and apoptosis. Nature 410, 112–116 (2001).

    Article  CAS  PubMed  Google Scholar 

  25. Horvitz, H.R. Genetic control of programmed cell death in the nematode Caenorhabditis elegans. Cancer Res. 59, 1701s–1706s (1999).

    CAS  PubMed  Google Scholar 

  26. Shaham, S. Identification of multiple Caenorhabditis elegans caspases and their potential roles in proteolytic cascades. J. Biol. Chem. 273, 35109–35117 (1998).

    Article  CAS  PubMed  Google Scholar 

  27. Reddien, P.W. & Horvitz, H.R. The engulfment process of programmed cell death in Caenorhabditis elegans. Annu. Rev. Cell Dev. Biol. 20, 193–221 (2004).

    Article  CAS  PubMed  Google Scholar 

  28. Hsu, D.R. & Meyer, B.J. The dpy-30 gene encodes an essential component of the Caenorhabditis elegans dosage compensation machinery. Genetics 137, 999–1018 (1994).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Gu, T., Orita, S. & Han, M. Caenorhabditis elegans SUR-5, a novel but conserved protein, negatively regulates LET-60 Ras activity during vulval induction. Mol. Cell. Biol. 18, 4556–4564 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Harbinder, S. et al. Genetically targeted cell disruption in Caenorhabditis elegans. Proc. Natl. Acad. Sci. USA 94, 13128–13133 (1997).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Zahn, T.R., Macmorris, M.A., Dong, W., Day, R. & Hutton, J.C. IDA-1, a Caenorhabditis elegans homolog of the diabetic autoantigens IA-2 and phogrin, is expressed in peptidergic neurons in the worm. J. Comp. Neurol. 429, 127–143 (2001).

    Article  CAS  PubMed  Google Scholar 

  32. Hengartner, M.O., Ellis, R.E. & Horvitz, H.R. Caenorhabditis elegans gene ced-9 protects cells from programmed cell death. Nature 356, 494–499 (1992).

    Article  CAS  PubMed  Google Scholar 

  33. Peden, E., Kimberly, E., Gengyo-Ando, K., Mitani, S. & Xue, D. Control of sex-specific apoptosis in C. elegans by the BarH homeodomain protein CEH-30 and the transcriptional repressor UNC-37/Groucho. Genes Dev. 21, 3195–3207 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Rotonda, J. et al. The three-dimensional structure of apopain/CPP32, a key mediator of apoptosis. Nat. Struct. Biol. 3, 619–625 (1996).

    Article  CAS  PubMed  Google Scholar 

  35. Xue, D. & Horvitz, H.R. Caenorhabditis elegans CED-9 protein is a bifunctional cell-death inhibitor. Nature 390, 305–308 (1997).

    Article  CAS  PubMed  Google Scholar 

  36. Harlin, H., Reffey, S.B., Duckett, C.S., Lindsten, T. & Thompson, C.B. Characterization of XIAP-deficient mice. Mol. Cell. Biol. 21, 3604–3608 (2001).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Wang, L., Miura, M., Bergeron, L., Zhu, H. & Yuan, J. Ich-1, an Ice/ced-3-related gene, encodes both positive and negative regulators of programmed cell death. Cell 78, 739–750 (1994).

    Article  CAS  PubMed  Google Scholar 

  38. Brenner, S. The genetics of Caenorhabditis elegans. Genetics 77, 71–94 (1974).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Riddle, D.L., Blumenthal, T., Meyer, B.J. & Preiss, J.R. (eds.). C. elegans II (Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, 1997).

    Google Scholar 

  40. Gengyo-Ando, K. & Mitani, S. Characterization of mutations induced by ethyl methanesulfonate, UV, and trimethylpsoralen in the nematode Caenorhabditis elegans. Biochem. Biophys. Res. Commun. 269, 64–69 (2000).

    Article  CAS  PubMed  Google Scholar 

  41. Wang, X., Yang, C., Chai, J., Shi, Y. & Xue, D. Mechanisms of AIF-mediated apoptotic DNA degradation in Caenorhabditis elegans. Science 298, 1587–1592 (2002).

    Article  CAS  PubMed  Google Scholar 

  42. Xue, D., Shaham, S. & Horvitz, H.R. The Caenorhabditis elegans cell-death protein CED-3 is a cysteine protease with substrate specificities similar to those of the human CPP32 protease. Genes Dev. 10, 1073–1083 (1996).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank members of the Xue laboratory and N. Yan for comments and discussions, A. Seluzicki for the initial observation, M. Driscoll (Rutgers University) for the bzIs8 strain, J. Hutton (University of Colorado Health Science Center) for the inIs179 strain and T. Blumenthal (University of Colorado) for anti–CstF-64 antibody. This work was supported by US National Institutes of Health R01 grants (GM059083 and GM079097 to D.X. and GM072633 to Yigong S.), a Burroughs Welcome Fund Career Award to D.X. and a grant from Ministry of Education, Culture, Sports, Science and Technology (MEXT) of Japan to S.M. X.G. is supported by University of Colorado Matching Grant to the SCR Training Grant (T32 GM08759).

Author information

Authors and Affiliations

  1. Department of Molecular, Cellular, and Developmental Biology, Campus Box 347, University of Colorado, Boulder, 80309, Colorado, USA

    Xin Geng, Yong Shi, Akihisa Nakagawa & Ding Xue

  2. Department of Physiology, Tokyo Women's Medical University, School of Medicine, and Core Research for Evolutional Science and Technology, Japan Science and Technology Agency, 8-1, Kawada-cho, Shinjuku-ku, Tokyo, 162-8666, Japan

    Sawako Yoshina & Shohei Mitani

  3. Department of Molecular Biology, Washington Road, Princeton University, Princeton, 08544, New Jersey, USA

    Yigong Shi

Authors
  1. Xin Geng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yong Shi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Akihisa Nakagawa
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Sawako Yoshina
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Shohei Mitani
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yigong Shi
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Ding Xue
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

X.G. performed most of the experiments. X.G. and D.X. designed and interpreted all experiments. Yigong S. performed the structural modeling of the CSP-3–CED-3 complex. S.Y. and S.M. isolated csp-3 deletion alleles. Yong S. and A.N. contributed to some of the experiments. X.G. and D.X. wrote the paper and others commented on the manuscript.

Corresponding author

Correspondence to Ding Xue.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1 and 2 and Supplementary Tables 1 and 2 (PDF 1274 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Geng, X., Shi, Y., Nakagawa, A. et al. Inhibition of CED-3 zymogen activation and apoptosis in Caenorhabditis elegans by caspase homolog CSP-3. Nat Struct Mol Biol 15, 1094–1101 (2008). https://doi.org/10.1038/nsmb.1488

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.1488

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing