title: RIPK3 regulates p62-LC3 complex formation via the caspase-8-dependent cleavage of p62.
abstract: RIPK3 is a key molecule for necroptosis, initially characterized by necrotic cell death morphology and the activation of autophagy. Cell death and autophagic signaling are believed to tightly regulate each other. However, the associated recruitment of signaling proteins remains poorly understood. p62/sequestosome-1 is a selective autophagy substrate and a selective receptor for ubiquitinated proteins. In this study, we illustrated that both mouse and human RIPK3 mediate p62 cleavage and that RIPK3 interacts with p62, resulting in complex formation. In addition, RIPK3-dependent p62 cleavage is restricted by the inhibition of caspases, especially caspase-8. Moreover, overexpression of A20, a ubiquitin-editing enzyme and an inhibitor of caspase-8 activity, inhibits RIPK3-dependent p62 cleavage. To further investigate the potential role of RIPK3 in selective autophagy, we analyzed p62-LC3 complex formation, revealing that RIPK3 prevents the localization of LC3 and ubiquitinated proteins to the p62 complex. In addition, RIPK3-dependent p62-LC3 complex disruption is regulated by caspase inhibition. Taken together, these results demonstrated that RIPK3 interacts with p62 and regulates p62-LC3 complex formation. These findings suggested that RIPK3 serves as a negative regulator of selective autophagy and provides new insights into the mechanism by which RIPK3 regulates autophagic signaling.
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Yu
authors.initials: Y
authors.lastname: Matsuzawa
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan. Electronic address: soshima.gast@tmd.ac.jp.
authors.forename: Shigeru
authors.initials: S
authors.lastname: Oshima
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Nibe
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Masanori
authors.initials: M
authors.lastname: Kobayashi
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Chiaki
authors.initials: C
authors.lastname: Maeyashiki
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Nemoto
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Nagaishi
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan; Center for Stem Cell and Regenerative Medicine, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Ryuichi
authors.initials: R
authors.lastname: Okamoto
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan; Department of Advanced Therapeutics for GI Diseases, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Kiichiro
authors.initials: K
authors.lastname: Tsuchiya
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan; Department of Advanced Therapeutics for GI Diseases, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Nakamura
authors.affiliation: Department of Gastroenterology and Hepatology, Graduate School, Tokyo Medical and Dental University (TMDU), Tokyo, Japan. Electronic address: mamoru.gast@tmd.ac.jp.
authors.forename: Mamoru
authors.initials: M
authors.lastname: Watanabe
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis; D001343:Autophagy; D053181:Caspase 8 / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000737:chemistry; D015971:Gene Expression Regulation, Enzymologic; D057809:HEK293 Cells; D006360:Heat-Shock Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D010588:Phagosomes / Q000378:metabolism; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000378:metabolism; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000378:metabolism*; D000071456:Sequestosome-1 Protein
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D004791:Enzyme Inhibitors; D006360:Heat-Shock Proteins; D034741:RNA, Small Interfering; C099718:SQSTM1 protein, human; D000071456:Sequestosome-1 Protein; C101343:Sqstm1 protein, mouse; C119705:RIPK3 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C485615:Ripk3 protein, mouse; D053181:Caspase 8
keywords: Autophagy; Caspase-8; LC3; Necroptosis; RIPK3; p62