title: Diverse sequence determinants control human and mouse receptor interacting protein 3 (RIP3) and mixed lineage kinase domain-like (MLKL) interaction in necroptotic signaling.
abstract: Receptor interacting protein 3 (RIP3) is a protein kinase essential for TNF-induced necroptosis. Phosphorylation on Ser-227 in human RIP3 (hRIP3) is required for its interaction with human mixed lineage kinase domain-like (MLKL) in the necrosome, a signaling complex induced by TNF stimulation. RIP1 and RIP3 mediate necrosome aggregation leading to the formation of amyloid-like signaling complexes. We found that TNF induces Thr-231 and Ser-232 phosphorylation in mouse RIP3 (mRIP3) and this phosphorylation is required for mRIP3 to interact with mMLKL. Ser-232 in mRIP3 corresponds to Ser-227 in hRIP3, whereas Thr-231 is not conserved in hRIP3. Although the RIP3-MLKL interaction is required for necroptosis in both human and mouse cells, hRIP3 does not interact with mMLKL and mRIP3 cannot bind to hMLKL. The species specificity of the RIP3-MLKL interaction is primarily determined by the sequence differences in the phosphorylation sites and the flanking sequence around the phosphorylation sites in hRIP3 and mRIP3. It appears that the RIP3-MLKL interaction has been selected as an evolutionarily conserved mechanism in mediating necroptosis signaling despite that differing structural and mechanistic bases for this interaction emerged simultaneously in different organisms. In addition, we further revealed that the interaction of RIP3 with MLKL prevented massive abnormal RIP3 aggregation, and therefore should be crucial for formation of the amyloid signaling complex of necrosomes. We also found that the interaction between RIP3 and MLKL is required for the translocation of necrosomes to mitochondria-associated membranes. Our data demonstrate the importance of the RIP3-MLKL interaction in the formation of functional necrosomes and suggest that translocation of necrosomes to mitochondria-associated membranes is essential for necroptosis signaling.
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Wanze
authors.initials: W
authors.lastname: Chen
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Zhenru
authors.initials: Z
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Lisheng
authors.initials: L
authors.lastname: Li
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Chuan-Qi
authors.initials: CQ
authors.lastname: Zhong
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Xinru
authors.initials: X
authors.lastname: Zheng
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Xiurong
authors.initials: X
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Yingying
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Huan
authors.initials: H
authors.lastname: Ma
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Deli
authors.initials: D
authors.lastname: Huang
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005.
authors.forename: Wenjuan
authors.initials: W
authors.lastname: Li
authors.affiliation: Life Sciences Institute, Zhejiang University, Hangzhou, Zhejiang 310058, China.
authors.forename: Zongping
authors.initials: Z
authors.lastname: Xia
authors.affiliation: State Key Laboratory of Cellular Stress Biology and School of Life Sciences, Xiamen University, Xiamen, Fujian 361005. Electronic address: jhan@xmu.edu.cn.
authors.forename: Jiahuai
authors.initials: J
authors.lastname: Han
mesh_terms: D000682:Amyloid / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D006801:Humans; D051379:Mice; D008931:Mitochondria, Muscle / Q000378:metabolism / Q000473:pathology; D032342:Muscle Cells / Q000201:enzymology* / Q000473:pathology; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D009336:Necrosis / Q000201:enzymology / Q000235:genetics / Q000473:pathology; D010766:Phosphorylation / Q000235:genetics; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D053422:Receptor-Interacting Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D000682:Amyloid; D009124:Muscle Proteins; C570693:MLKL protein, human; C570694:MLKL protein, mouse; D011494:Protein Kinases; C119705:RIPK3 protein, human; D053422:Receptor-Interacting Protein Serine-Threonine Kinases; C485615:Ripk3 protein, mouse
keywords: Cell Death; MLKL; Necroptosis; Necrosis (Necrotic Death); Protein Kinases; Protein Phosphorylation; Protein-Protein Interactions; RIP; RIP3; Tumor Necrosis Factor (TNF)