Insights into ubiquitin chain architecture using Ub-clipping

Swatek, KN; Usher, JL; Kueck, AF; Gladkova, C; Mevissen, TET; Pruneda, JN; Skern, T; Komander, D

Author(s): Swatek, KN; Usher, JL; Kueck, AF; Gladkova, C; Mevissen, TET; Pruneda, JN; Skern, T; Komander, D;
Details: Publication Year 2019-08-14,Volume 572,Issue #7770,Page 553-537
Journal Title: Nature
Publication Type: Journal Article
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 acetylation(1-3). 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, Lb(pro) *, 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 Lb(pro) * 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.
Publisher: Springer Nature
Research Division(s): Ubiquitin Signalling
PubMed ID: 31413367
Publisher's Version: https://doi.org/10.1038/s41586-019-1482-y
Terms of Use/Rights Notice: Refer to copyright notice on published article.

Creation Date: 2019-08-21 11:19:31

Last Modified: 2019-08-23 02:19:47