Combinatorial ubiquitin code degrades deubiquitylation-protected substrates

Abstract Protein ubiquitylation is maintained by a dynamic balance of the conjugation and deconjugation of ubiquitin. It remains unclear how deubiquitylation-stabilized substrates are directed for degradation. Branched ubiquitin chains promote substrate degradation through the proteasome. TRIP12 and...

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Main Authors: Mai Morita, Miyu Takao, Honoka Tokuhisa, Ryotaro Chiba, Shota Tomomatsu, Yoshino Akizuki, Takuya Tomita, Akinori Endo, Yasushi Saeki, Yusuke Sato, Fumiaki Ohtake
Format: Article
Language:English
Published: Nature Portfolio 2025-03-01
Series:Nature Communications
Online Access:https://doi.org/10.1038/s41467-025-57873-9
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author Mai Morita
Miyu Takao
Honoka Tokuhisa
Ryotaro Chiba
Shota Tomomatsu
Yoshino Akizuki
Takuya Tomita
Akinori Endo
Yasushi Saeki
Yusuke Sato
Fumiaki Ohtake
author_facet Mai Morita
Miyu Takao
Honoka Tokuhisa
Ryotaro Chiba
Shota Tomomatsu
Yoshino Akizuki
Takuya Tomita
Akinori Endo
Yasushi Saeki
Yusuke Sato
Fumiaki Ohtake
author_sort Mai Morita
collection DOAJ
description Abstract Protein ubiquitylation is maintained by a dynamic balance of the conjugation and deconjugation of ubiquitin. It remains unclear how deubiquitylation-stabilized substrates are directed for degradation. Branched ubiquitin chains promote substrate degradation through the proteasome. TRIP12 and UBR5 are HECT-type E3 ubiquitin ligases, which are specific for lysine 29 (K29) and lysine 48 (K48) linkages, respectively. Here, we show that the deubiquitylase (DUB) OTUD5 is cooperatively modified by TRIP12 and UBR5, resulting in conjugation of K29/K48 branched ubiquitin chains and accelerated proteasomal degradation. TRIP12–OTUD5 antagonism regulates TNF-α–induced NF-κB signaling. Mechanistically, OTUD5 readily cleaves K48 linkages, but does not affect K29 linkages. Consequently, K29 linkages overcome OTUD5 DUB activity to facilitate UBR5-dependent K48-linked chain branching. This mechanism is applicable to other OTUD5-associated TRIP12 substrates. Thus, the combination of DUB-resistant and proteasome-targeting ubiquitin linkages promotes the degradation of deubiquitylation-protected substrates, underscoring the role of branched ubiquitin chains in shifting the ubiquitin conjugation/deconjugation equilibrium.
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spelling doaj-art-7c742fa9ec344d2894f6c241b4d7ea2a2025-08-20T02:10:10ZengNature PortfolioNature Communications2041-17232025-03-0116111710.1038/s41467-025-57873-9Combinatorial ubiquitin code degrades deubiquitylation-protected substratesMai Morita0Miyu Takao1Honoka Tokuhisa2Ryotaro Chiba3Shota Tomomatsu4Yoshino Akizuki5Takuya Tomita6Akinori Endo7Yasushi Saeki8Yusuke Sato9Fumiaki Ohtake10Laboratory of Protein Degradation, Institute for Advanced Life Sciences, Hoshi University, 2-4-41 Ebara, Shinagawa-kuLaboratory of Protein Degradation, Institute for Advanced Life Sciences, Hoshi University, 2-4-41 Ebara, Shinagawa-kuDepartment of Chemistry and Biotechnology, Graduate School of Engineering, Tottori University, 4-101 Koyama-cho Minami, Tottori-shiLaboratory of Protein Degradation, Institute for Advanced Life Sciences, Hoshi University, 2-4-41 Ebara, Shinagawa-kuLaboratory of Protein Degradation, Institute for Advanced Life Sciences, Hoshi University, 2-4-41 Ebara, Shinagawa-kuLaboratory of Protein Degradation, Institute for Advanced Life Sciences, Hoshi University, 2-4-41 Ebara, Shinagawa-kuDivision of Protein Metabolism, The Institute of Medical Science, The University of Tokyo, 4-6-1, Shirokanedai, Minato-kuProtein Metabolism Project, Tokyo Metropolitan Institute of Medical Sciences, 2-1-6 Kamikitazawa, Setagaya-kuDivision of Protein Metabolism, The Institute of Medical Science, The University of Tokyo, 4-6-1, Shirokanedai, Minato-kuDepartment of Chemistry and Biotechnology, Graduate School of Engineering, Tottori University, 4-101 Koyama-cho Minami, Tottori-shiLaboratory of Protein Degradation, Institute for Advanced Life Sciences, Hoshi University, 2-4-41 Ebara, Shinagawa-kuAbstract Protein ubiquitylation is maintained by a dynamic balance of the conjugation and deconjugation of ubiquitin. It remains unclear how deubiquitylation-stabilized substrates are directed for degradation. Branched ubiquitin chains promote substrate degradation through the proteasome. TRIP12 and UBR5 are HECT-type E3 ubiquitin ligases, which are specific for lysine 29 (K29) and lysine 48 (K48) linkages, respectively. Here, we show that the deubiquitylase (DUB) OTUD5 is cooperatively modified by TRIP12 and UBR5, resulting in conjugation of K29/K48 branched ubiquitin chains and accelerated proteasomal degradation. TRIP12–OTUD5 antagonism regulates TNF-α–induced NF-κB signaling. Mechanistically, OTUD5 readily cleaves K48 linkages, but does not affect K29 linkages. Consequently, K29 linkages overcome OTUD5 DUB activity to facilitate UBR5-dependent K48-linked chain branching. This mechanism is applicable to other OTUD5-associated TRIP12 substrates. Thus, the combination of DUB-resistant and proteasome-targeting ubiquitin linkages promotes the degradation of deubiquitylation-protected substrates, underscoring the role of branched ubiquitin chains in shifting the ubiquitin conjugation/deconjugation equilibrium.https://doi.org/10.1038/s41467-025-57873-9
spellingShingle Mai Morita
Miyu Takao
Honoka Tokuhisa
Ryotaro Chiba
Shota Tomomatsu
Yoshino Akizuki
Takuya Tomita
Akinori Endo
Yasushi Saeki
Yusuke Sato
Fumiaki Ohtake
Combinatorial ubiquitin code degrades deubiquitylation-protected substrates
Nature Communications
title Combinatorial ubiquitin code degrades deubiquitylation-protected substrates
title_full Combinatorial ubiquitin code degrades deubiquitylation-protected substrates
title_fullStr Combinatorial ubiquitin code degrades deubiquitylation-protected substrates
title_full_unstemmed Combinatorial ubiquitin code degrades deubiquitylation-protected substrates
title_short Combinatorial ubiquitin code degrades deubiquitylation-protected substrates
title_sort combinatorial ubiquitin code degrades deubiquitylation protected substrates
url https://doi.org/10.1038/s41467-025-57873-9
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