Deubiquitinating enzymes and the proteasome regulate preferential sets of ubiquitin substrates

Trulsson, F.; Akimov, V.; Robu, M.; Overbeek, N. van; Berrocal, D.A.P.; Shah, R.G.; Cox, J.; Shah, G.M.; Blagoev, B.; Vertegaal, A.C.O.

doi:doi:10.1038/s41467-022-30376-7

The ubiquitin-proteasome axis has been extensively explored at a system-wide level, but the impact of deubiquitinating enzymes (DUBs) on the ubiquitinome remains largely unknown. Here, we compare the contributions of the proteasome and DUBs on the global ubiquitinome, using UbiSite technology, inhibitors and mass spectrometry. We uncover large dynamic ubiquitin signalling networks with substrates and sites preferentially regulated by DUBs or by the proteasome, highlighting the role of DUBs in degradation-independent ubiquitination. DUBs regulate substrates via at least 40,000 unique sites. Regulated networks of ubiquitin substrates are involved in autophagy, apoptosis, genome integrity, telomere integrity, cell cycle progression, mitochondrial function, vesicle transport, signal transduction, transcription, pre-mRNA splicing and many other cellular processes. Moreover, we show that ubiquitin conjugated to SUMO2/3 forms a strong proteasomal degradation signal. Interestingly, PARP1...Show moreThe ubiquitin-proteasome axis has been extensively explored at a system-wide level, but the impact of deubiquitinating enzymes (DUBs) on the ubiquitinome remains largely unknown. Here, we compare the contributions of the proteasome and DUBs on the global ubiquitinome, using UbiSite technology, inhibitors and mass spectrometry. We uncover large dynamic ubiquitin signalling networks with substrates and sites preferentially regulated by DUBs or by the proteasome, highlighting the role of DUBs in degradation-independent ubiquitination. DUBs regulate substrates via at least 40,000 unique sites. Regulated networks of ubiquitin substrates are involved in autophagy, apoptosis, genome integrity, telomere integrity, cell cycle progression, mitochondrial function, vesicle transport, signal transduction, transcription, pre-mRNA splicing and many other cellular processes. Moreover, we show that ubiquitin conjugated to SUMO2/3 forms a strong proteasomal degradation signal. Interestingly, PARP1 is hyper-ubiquitinated in response to DUB inhibition, which increases its enzymatic activity. Our study uncovers key regulatory roles of DUBs and provides a resource of endogenous ubiquitination sites to aid the analysis of substrate specific ubiquitin signalling.Deubiquitinases (DUBs) remove ubiquitin from its target proteins. Here, authors compare the regulatory effects of the proteasome and DUBs on the ubiquitinated proteome. They find preferential sets of substrates regulated by DUBs or by the proteasome. Moreover, they find that PARP1 is hyper-ubiquitinated in response to DUB inhibition, which increases its enzymatic activity.Show less

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Deubiquitinating enzymes and the proteasome regulate preferential sets of ubiquitin substrates