Disruption of the CED-9.CED-4 complex by EGL-1 is a critical step for programmed cell death in Caenorhabditis elegans.
In the nematode Caenorhabditis elegans, the apoptotic machinery is composed of four basic elements: the caspase CED-3, the Apaf-1 homologue CED-4, and the Bcl-2 family members CED-9 and EGL-1. The ced-9(n1950) gain-of-function mutation prevents most, if not all, somatic cell deaths in C. elegans. It encodes a CED-9 protein with a glycine-to-glutamate substitution at position 169, which is located within the highly conserved Bcl-2 homology 1 domain. We performed biochemical analyses with the CED-9G169E protein to gain insight into the mechanism of programmed cell death. We find that CED-9G169E retains the ability to bind both EGL-1 and CED-4, although its affinity for EGL-1 is reduced. In contrast to the behavior of wild-type CED-9, the interaction between CED-9G169E and CED-4 is not disrupted by expression of EGL-1. Furthermore, CED-4 and CED-9G169E co-localizes with EGL-1 to the mitochondria in mammalian cells, and expression of EGL-1 does not induce translocation of CED-4 to the cytosol. Finally, the ability of EGL-1 to promote apoptosis is impaired by the replacement of wild-type CED-9 with CED-9G169E, and this effect is correlated with the inability of EGL-1 to induce the displacement of CED-4 from the CED-9.CED-4 complex. These studies suggest that the release of CED-4 from the CED-9.CED-4 complex is a necessary step for induction of programmed cell death in C. elegans.
del Peso
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L
Department of Pathology and Comprehensive Cancer Center and the The University of Michigan, Ann Arbor, Michigan 48109, USA.
Gonzalez
V M
VM
Inohara
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N
Ellis
R E
RE
Núñez
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G
D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D051017:Apoptosis Regulatory Proteins; D017173:Caenorhabditis elegans / Q000166:cytology* / Q000378:metabolism; D029742:Caenorhabditis elegans Proteins; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D015801:Helminth Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2; D012097:Repressor Proteins / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D051017:Apoptosis Regulatory Proteins; D029742:Caenorhabditis elegans Proteins; D002135:Calcium-Binding Proteins; C089224:Ced-4 protein, C elegans; C087233:Ced-9 protein, C elegans; C112486:EGL-1 protein, C elegans; D015801:Helminth Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012097:Repressor Proteins