Caenorhabditis elegans EGL-1 disrupts the interaction of CED-9 with CED-4 and promotes CED-3 activation.
In the nematode Caenorhabditis elegans, programmed cell death is implemented by the protease CED-3 whose activity is inhibited by CED-9 through physical associations with the regulator CED-4. The product of a recently described gene, egl-1, binds to and inhibits CED-9. In the present studies, we have addressed the molecular mechanism by which EGL-1 regulates CED-9 function and promotes cell death. Expression of CED-4 and CED-3 resulted in decreased survival and apoptosis of mammalian cells, activities that could be inhibited by CED-9. Importantly, this protective effect of CED-9 was antagonized by EGL-1. Immunoprecipitation analysis showed that EGL-1 binding to CED-9 disrupts the association between CED-4 and CED-9, an activity that required the BH3 motif of EGL-1. Consistent with these results, expression of EGL-1 promoted CED-4-dependent processing of CED-3, and this activity of EGL-1 was mediated through inhibition of CED-9. In mammalian cells, CED-9 is known to target the subcellular localization of CED-4 from the cytosol to intracellular membranes. Expression of EGL-1 resulted in redistribution of CED-4 from intracellular membranes, where it co-localized with CED-9, to the cytoplasm, providing further evidence that EGL-1 regulates CED-4 through CED-9. Finally, the levels of EGL-1 were greatly enhanced by co-expression of CED-9 in both mammalian cells and in a cell-free system, suggesting a role for CED-9 in the expression and/or stabilization of EGL-1. These studies provide a mechanism for how EGL-1 functions to antagonize pro-survival of CED-9 and to promote CED-3 activation and programmed cell death.
del Peso
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Department of Pathology and Comprehensive Cancer Center, the University of Michigan Medical School, Ann Arbor, Michigan 48109, USA.
González
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Núñez
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D000818:Animals; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D001483:Base Sequence; D017173:Caenorhabditis elegans / Q000378:metabolism*; D029742:Caenorhabditis elegans Proteins; D002135:Calcium-Binding Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D020169:Caspases; D002460:Cell Line; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D017931:DNA Primers; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D015801:Helminth Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D011485:Protein Binding; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*
D051017:Apoptosis Regulatory Proteins; D029742:Caenorhabditis elegans Proteins; D002135:Calcium-Binding Proteins; C089224:Ced-4 protein, C elegans; C087233:Ced-9 protein, C elegans; D017931:DNA Primers; C112486:EGL-1 protein, C elegans; D015801:Helminth Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012097:Repressor Proteins; D020169:Caspases; D003546:Cysteine Endopeptidases; C089223:ced-3 protein, C elegans