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Structure of the CED-4–CED-9 complex provides insights into programmed cell death in Caenorhabditis elegans

Nature volume 437, pages 831–837 (2005)Cite this article

Abstract

Interplay among four genes—egl-1, ced-9, ced-4 and ced-3—controls the onset of programmed cell death in the nematode Caenorhabditis elegans. Activation of the cell-killing protease CED-3 requires CED-4. However, CED-4 is constitutively inhibited by CED-9 until its release by EGL-1. Here we report the crystal structure of the CED-4–CED-9 complex at 2.6 Å resolution, and a complete reconstitution of the CED-3 activation pathway using homogeneous proteins of CED-4, CED-9 and EGL-1. One molecule of CED-9 binds to an asymmetric dimer of CED-4, but specifically recognizes only one of the two CED-4 molecules. This specific interaction prevents CED-4 from activating CED-3. EGL-1 binding induces pronounced conformational changes in CED-9 that result in the dissociation of the CED-4 dimer from CED-9. The released CED-4 dimer further dimerizes to form a tetramer, which facilitates the autoactivation of CED-3. Together, our studies provide important insights into the regulation of cell death activation in C. elegans.

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Figure 1: Overall structure of the CED-4–CED-9 complex.
Figure 2: Mechanisms of CED-4 recognition by CED-9, and CED-4 release by EGL-1.
Figure 3: Structural analyses of the CED-4a–CED-4b interface and ATP binding.
Figure 4: Free CED-4 forms a tetramer.
Figure 5: Complete reconstitution of the CED-3 activation pathway using homogeneous proteins of CED-4, CED-9 and EGL-1.

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Acknowledgements

We thank H. R. Horvitz for complementary DNA sequences of egl-1, ced-9, ced-4 and ced-3, and for his encouragement. We thank V. Dixit and X. Yang for providing ced-3 constructs, and M. Becker and A. Saxena for beamtime at NSLS. This research was supported by NIH grants to Y.S. and D.X. H.L. acknowledges support from the Brookhaven National Laboratory LDRD program and a Department of Energy grant. The atomic coordinates of the CED-4–CED-9 complex have been deposited in the Protein Data Bank with the accession number 2A5Y. Author Contributions N.Y. performed the bulk of the experiments. N.Y. and Y.S. designed and interpreted the bulk of the experiments. J.C., E.S.L., L.G., Q.L., J.H., J.-W.W. and D.K. contributed to experiments. J.C., E.S.L., L.G., Q.L., J.H., J.-W.W., D.K., H.L., Q.H. and D.X. contributed to data analysis and interpretation. N.Y. and Y.S. wrote the paper. J.C. and E.S.L. contributed equally to this work.

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Authors and Affiliations

  1. Department of Molecular Biology, Princeton University, Lewis Thomas Laboratory, Washington Road, New Jersey, 08544, Princeton, USA

    Nieng Yan, Jijie Chai, Lichuan Gu, Jia-Wei Wu & Yigong Shi

  2. Department of Molecular, Cellular and Developmental Biology, University of Colorado, Colorado, 80309, Boulder, USA

    Eui Seung Lee, David Kokel & Ding Xue

  3. Department of Life Science, Gwangju Institute of Science and Technology, 500-712, Gwangju, Republic of Korea

    Eui Seung Lee

  4. Molecular Biology and Genetics, Cornell University, New York, 14853, Ithaca, USA

    Qun Liu & Quan Hao

  5. Biology Department, Brookhaven National Laboratory, New York, 11973, Upton, USA

    Jiaqing He & Huilin Li

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Supplementary information

Supplementary Table 1

Summary of Crystallographic Analysis. (DOC 24 kb)

Supplementary Table 2

Biological and biochemical characterization of CED-4 mutants. (DOC 48 kb)

Supplementary Figure 1

Mapping of the surface features onto the primary sequences of CED-4 (JPG 448 kb)

Supplementary Figure 2

Mechanism of CED-4 release induced by EGL-1 binding. (JPG 266 kb)

Supplementary Figure 3

CED-4a and CED-4b are structurally similar. (JPG 514 kb)

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Yan, N., Chai, J., Lee, E. et al. Structure of the CED-4–CED-9 complex provides insights into programmed cell death in Caenorhabditis elegans. Nature 437, 831–837 (2005). https://doi.org/10.1038/nature04002

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