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Dark is a Drosophila homologue of Apaf-1/CED-4 and functions in an evolutionarily conserved death pathway

Nature Cell Biology volume 1, pages 272–279 (1999)Cite this article

Abstract

Here we identify a new gene, dark, which encodes a Drosophila homologue of mammalian Apaf-1 and Caenorhabditis elegans CED-4, cell-death proteins. Like Apaf-1, but in contrast to CED-4, Dark contains a carboxy-terminal WD-repeat domain necessary for interactions with the mitochondrial protein cytochrome c. Dark selectively associates with another protein involved in apoptosis, the fly apical caspase, Dredd. Dark-induced cell killing is suppressed by caspase-inhibitory peptides and by a dominant-negative mutant Dredd protein, and enhanced by removal of the WD domain. Loss-of-function mutations in dark attenuate programmed cell deaths during development, causing hyperplasia of the central nervous system, and other abnormalities including ectopic melanotic tumours and defective wings. Moreover, ectopic cell killing by the Drosophila cell-death activators, Reaper, Grim and Hid, is substantially suppressed in dark mutants. These findings establish dark as an important apoptosis effector in Drosophila and raise profound evolutionary considerations concerning the relationship between mitochondrial components and the apoptosis-promoting machinery.

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Figure 1: The predicted amino-acid sequence of Dark and comparison to Apaf-1 and CED-4.
Figure 2: Dark-induced cell killing is suppressed by an active-site Dredd mutant.
Figure 3: Dark interacts with the fly apical caspase, Dredd, and cytochrome c.
Figure 4: Expression of dark during development.
Figure 5: dark loss-of-function phenotypes.
Figure 6: Reduced apoptosis in dark mutants.
Figure 7: Dark function is required for reaper-, grim-, and hid-induced cell death.

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Acknowledgements

We thank J. Chapo, S.I. Ho and Y. Li for technical expertise; C. Klingerberg and T. Barrientos for help with mutagenesis; G. Lawton for electron microscopy; J. Waddle for photographic help; the Bloomington stock center for fly strains; C. Thummel for cDNA libraries; A. Fraser and G. Evans for drICE plasmids; and J. Agapite and H. Steller for the P[GMR-hid]-1M strain. This work was supported by grants from the NIA/NIH (R01 AG12466) and the NSF (MCB-9816841) (to J.M.A.) and from the American Cancer Society (Re258), NIH (GMR01-57158) and Welch Foundation (I1412) to X.W.

Correspondence and requests for materials should be addressed to J.M.A. The Drosophila genomic region containing the dark gene is contained in DNA located in GenBank under accession number AC004335. The dark cDNA has been submitted to GenBank under accession number AF162659.

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  1. Antony Rodriguez, Holt Oliver, Hua Zou and Po Chen: These authors contributed equally to this work

Authors and Affiliations

  1. Department of Cell Biology and Neuroscience, The University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, 75235-9039, Texas, USA

    Antony Rodriguez, Po Chen & John M. Abrams

  2. Howard Hughes Medical Institute and Department of Biochemistry, The University of Texas Southwestern Medical Center, 5323 Harry Hines Boulevard, Dallas, 75235-9039, Texas, USA

    Holt Oliver, Hua Zou & Xiaodong Wang

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Correspondence to John M. Abrams.

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Rodriguez, A., Oliver, H., Zou, H. et al. Dark is a Drosophila homologue of Apaf-1/CED-4 and functions in an evolutionarily conserved death pathway. Nat Cell Biol 1, 272–279 (1999). https://doi.org/10.1038/12984

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