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Mol Cell. 2017 Mar 2;65(5):941-955.e8. doi: 10.1016/j.molcel.2017.01.004. Epub 2017 Feb 9.

An Interaction Landscape of Ubiquitin Signaling.

Author information

1
Department of Molecular Biology, Faculty of Science, Radboud Institute for Molecular Life Sciences, Radboud University Nijmegen, Nijmegen 6525, the Netherlands. Electronic address: x.zhang@science.ru.nl.
2
Department of Molecular Biology, Faculty of Science, Radboud Institute for Molecular Life Sciences, Radboud University Nijmegen, Nijmegen 6525, the Netherlands.
3
Division of Cell Biology II, Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam 1066CX, the Netherlands; Department of Chemical Immunology, Leiden University Medical Center, Albinusdreef 2, Leiden 2333ZA, the Netherlands.
4
Division of Cell Biology II, Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam 1066CX, the Netherlands; Department of Chemical Immunology, Leiden University Medical Center, Albinusdreef 2, Leiden 2333ZA, the Netherlands. Electronic address: h.ovaa@lumc.nl.
5
Department of Molecular Biology, Faculty of Science, Radboud Institute for Molecular Life Sciences, Radboud University Nijmegen, Nijmegen 6525, the Netherlands. Electronic address: michiel.vermeulen@science.ru.nl.

Abstract

Intracellular signaling via the covalent attachment of different ubiquitin linkages to protein substrates is fundamental to many cellular processes. Although linkage-selective ubiquitin interactors have been studied on a case-by-case basis, proteome-wide analyses have not been conducted yet. Here, we present ubiquitin interactor affinity enrichment-mass spectrometry (UbIA-MS), a quantitative interaction proteomics method that makes use of chemically synthesized diubiquitin to enrich and identify ubiquitin linkage interactors from crude cell lysates. UbIA-MS reveals linkage-selective diubiquitin interactions in multiple cell types. For example, we identify TAB2 and TAB3 as novel K6 diubiquitin interactors and characterize UCHL3 as a K27-linkage selective interactor that regulates K27 polyubiquitin chain formation in cells. Additionally, we show a class of monoubiquitin and K6 diubiquitin interactors whose binding is induced by DNA damage. We expect that our proteome-wide diubiquitin interaction landscape and established workflows will have broad applications in the ongoing efforts to decipher the complex language of ubiquitin signaling.

KEYWORDS:

DNA damage; TAB2; UCHL3; UbIA-MS; diubiquitin; interaction proteomics; ubiquitin signaling

PMID:
28190767
DOI:
10.1016/j.molcel.2017.01.004
[Indexed for MEDLINE]
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