A GH3-like domain in reaper is required for mitochondrial localization and induction of IAP degradation.

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TitleA GH3-like domain in reaper is required for mitochondrial localization and induction of IAP degradation.
Publication TypeJournal Article
Year of Publication2003
AuthorsOlson, MR, Holley, CL, Gan, EC, Colón-Ramos, DA, Kaplan, B, Kornbluth, S
JournalJ Biol Chem
Volume278
Issue45
Pagination44758-68
Date Published2003 Nov 7
ISSN0021-9258
KeywordsAnimals, Apoptosis, bcl-X Protein, Caspases, Cell Nucleus, Cells, Cultured, Drosophila melanogaster, Drosophila Proteins, Enzyme Inhibitors, Gene Expression, Green Fluorescent Proteins, Inhibitor of Apoptosis Proteins, Luminescent Proteins, Microscopy, Confocal, Mitochondria, Mutagenesis, Neuropeptides, Peptide Fragments, Protein Structure, Secondary, Proto-Oncogene Proteins c-bcl-2, Recombinant Fusion Proteins, Saccharomyces cerevisiae, Structure-Activity Relationship, Transfection
Abstract

Reaper is a potent pro-apoptotic protein originally identified in a screen for Drosophila mutants defective in apoptotic induction. Multiple functions have been ascribed to this protein, including inhibition of IAPs (inhibitors of apoptosis); induction of IAP degradation; inhibition of protein translation; and when expressed in vertebrate cells, induction of mitochondrial cytochrome c release. Structure/function analysis of Reaper has identified an extreme N-terminal motif that appears to be sufficient for inhibition of IAP function. We report here that this domain, although required for IAP destabilization, is not sufficient. Moreover, we have identified a small region of Reaper, similar to the GH3 domain of Grim, that is required for localization of Reaper to mitochondria, induction of IAP degradation, and potent cell killing. Although a mutant Reaper protein lacking the GH3 domain was deficient in these properties, these defects could be fully rectified by appending either the C-terminal mitochondrial targeting sequence from Bcl-xL or a homologous region from the pro-apoptotic protein HID. Together, these data strongly suggest that IAP destabilization by Reaper in intact cells requires Reaper localization to mitochondria and that induction of IAP instability by Reaper is important for the potent induction of apoptosis in Drosophila cells.

DOI10.1074/jbc.M308055200
Alternate JournalJ. Biol. Chem.
PubMed ID12917412