Insights into ubiquitin chain architecture using Ub-clipping

Kirby N. Swatek, Joanne L. Usher, Anja F. Kueck, Christina Gladkova, Tycho E.T. Mevissen, Jonathan Pruneda, Tim Skern, David Komander

Research output: Contribution to journalLetter

Abstract

Protein ubiquitination is a multi-functional post-translational modification that affects all cellular processes. Its versatility arises from architecturally complex polyubiquitin chains, in which individual ubiquitin moieties may be ubiquitinated on one or multiple residues, and/or modified by phosphorylation and acetylation1–3. Advances in mass spectrometry have enabled the mapping of individual ubiquitin modifications that generate the ubiquitin code; however, the architecture of polyubiquitin signals has remained largely inaccessible. Here we introduce Ub-clipping as a methodology by which to understand polyubiquitin signals and architectures. Ub-clipping uses an engineered viral protease, Lbpro∗, to incompletely remove ubiquitin from substrates and leave the signature C-terminal GlyGly dipeptide attached to the modified residue; this simplifies the direct assessment of protein ubiquitination on substrates and within polyubiquitin. Monoubiquitin generated by Lbpro∗ retains GlyGly-modified residues, enabling the quantification of multiply GlyGly-modified branch-point ubiquitin. Notably, we find that a large amount (10–20%) of ubiquitin in polymers seems to exist as branched chains. Moreover, Ub-clipping enables the assessment of co-existing ubiquitin modifications. The analysis of depolarized mitochondria reveals that PINK1/parkin-mediated mitophagy predominantly exploits mono- and short-chain polyubiquitin, in which phosphorylated ubiquitin moieties are not further modified. Ub-clipping can therefore provide insight into the combinatorial complexity and architecture of the ubiquitin code.

Original languageEnglish (US)
Pages (from-to)533-537
Number of pages5
JournalNature
Volume572
Issue number7770
DOIs
StatePublished - Aug 22 2019

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Ubiquitin
Polyubiquitin
Ubiquitination
Mitochondrial Degradation
Dipeptides
Post Translational Protein Processing
Mass Spectrometry
Mitochondria
Polymers
Proteins
Peptide Hydrolases
Phosphorylation

ASJC Scopus subject areas

  • General

Cite this

Swatek, K. N., Usher, J. L., Kueck, A. F., Gladkova, C., Mevissen, T. E. T., Pruneda, J., ... Komander, D. (2019). Insights into ubiquitin chain architecture using Ub-clipping. Nature, 572(7770), 533-537. https://doi.org/10.1038/s41586-019-1482-y

Insights into ubiquitin chain architecture using Ub-clipping. / Swatek, Kirby N.; Usher, Joanne L.; Kueck, Anja F.; Gladkova, Christina; Mevissen, Tycho E.T.; Pruneda, Jonathan; Skern, Tim; Komander, David.

In: Nature, Vol. 572, No. 7770, 22.08.2019, p. 533-537.

Research output: Contribution to journalLetter

Swatek, KN, Usher, JL, Kueck, AF, Gladkova, C, Mevissen, TET, Pruneda, J, Skern, T & Komander, D 2019, 'Insights into ubiquitin chain architecture using Ub-clipping', Nature, vol. 572, no. 7770, pp. 533-537. https://doi.org/10.1038/s41586-019-1482-y
Swatek KN, Usher JL, Kueck AF, Gladkova C, Mevissen TET, Pruneda J et al. Insights into ubiquitin chain architecture using Ub-clipping. Nature. 2019 Aug 22;572(7770):533-537. https://doi.org/10.1038/s41586-019-1482-y
Swatek, Kirby N. ; Usher, Joanne L. ; Kueck, Anja F. ; Gladkova, Christina ; Mevissen, Tycho E.T. ; Pruneda, Jonathan ; Skern, Tim ; Komander, David. / Insights into ubiquitin chain architecture using Ub-clipping. In: Nature. 2019 ; Vol. 572, No. 7770. pp. 533-537.
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