title: Cellular inhibitor of apoptosis protein 1 ubiquitinates endonuclease G but does not affect endonuclease G-mediated cell death.
abstract: Inhibitors of Apoptosis Proteins (IAPs) are evolutionarily well conserved and have been recognized as the key negative regulators of apoptosis. Recently, the role of IAPs as E3 ligases through the Ring domain was revealed. Using proteomic analysis to explore potential target proteins of DIAP1, we identified Drosophila Endonuclease G (dEndoG), which is known as an effector of caspase-independent cell death. In this study, we demonstrate that human EndoG interacts with IAPs, including human cellular Inhibitor of Apoptosis Protein 1 (cIAP1). EndoG was ubiquitinated by IAPs in vitro and in human cell lines. Interestingly, cIAP1 was capable of ubiquitinating EndoG in the presence of wild-type and mutant Ubiquitin, in which all lysines except K63 were mutated to arginine. cIAP1 expression did not change the half-life of EndoG and cIAP1 depletion did not alter its levels. Expression of dEndoG 54310, in which the mitochondrial localization sequence was deleted, led to cell death that could not be suppressed by DIAP1 in S2 cells. Moreover, EndoG-mediated cell death induced by oxidative stress in HeLa cells was not affected by cIAP1. Therefore, these results indicate that IAPs interact and ubiquitinate EndoG via K63-mediated isopeptide linkages without affecting EndoG levels and EndoG-mediated cell death, suggesting that EndoG ubiquitination by IAPs may serve as a regulatory signal independent of proteasomal degradation.
authors.affiliation: Department of Biology, Research Institute for Basic Sciences, Kyung Hee University, Seoul 130-701, Republic of Korea.
authors.forename: Tae Woong
authors.initials: TW
authors.lastname: Seo
authors.affiliation: Department of Nanopharmaceutical Life Sciences, Research Institute for Basic Sciences, Kyung Hee University, Seoul 130-701, Republic of Korea.
authors.forename: Ji Sun
authors.initials: JS
authors.lastname: Lee
authors.affiliation: Department of Biology, Research Institute for Basic Sciences, Kyung Hee University, Seoul 130-701, Republic of Korea; Department of Nanopharmaceutical Life Sciences, Research Institute for Basic Sciences, Kyung Hee University, Seoul 130-701, Republic of Korea. Electronic address: yoosoonji@khu.ac.kr.
authors.forename: Soon Ji
authors.initials: SJ
authors.lastname: Yoo
mesh_terms: D000818:Animals; D017209:Apoptosis; D016923:Cell Death; D004330:Drosophila; D029721:Drosophila Proteins / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology; D025801:Ubiquitin / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D054875:Ubiquitination
chemical_list: C111891:DIAP1 protein, Drosophila; D029721:Drosophila Proteins; D051034:Inhibitor of Apoptosis Proteins; D025801:Ubiquitin; C494833:BIRC2 protein, human; D044767:Ubiquitin-Protein Ligases; D004706:Endodeoxyribonucleases; C052391:endonuclease G
keywords: Cell death; E3 ligase; EndoG; K63-linked ubiquitination; cIAP1