PXD035045-2

PXD035045 is an original dataset announced via ProteomeXchange.

Title	Ubiquitinome profiling reveals in vivo UBE2D3 targets and implicates UBE2D3 in protein quality control.
Description	Ubiquitination has crucial roles in many cellular processes and dysregulation of ubiquitin machinery enzymes can result in various forms of pathogenesis. Cells only have a limited set of ubiquitin-conjugating (E2) enzymes to support the ubiquitination of many cellular targets. As individual E2 enzymes have many different substrates and interactions between E2 enzymes and their substrates can be transient, it is challenging to define all in vivo substrates of an individual E2 and the cellular processes it affects. Particularly challenging in this respect is UBE2D3, an E2 enzyme with promiscuous activity in vitro but less defined roles in vivo. Here, we set out to identify in vivo targets of UBE2D3 by using SILAC-based and label-free quantitative ubiquitin diGly proteomics to study global proteome and ubiquitinome changes associated with UBE2D3 depletion. UBE2D3 depletion changed the global proteome, with proteins from metabolic pathways, in particular retinol metabolism, being the most affected. The impact of UBE2D3 depletion on the ubiquitinome was much more prominent. Interestingly, molecular pathways related to mRNA translation were the most affected. Indeed, we find that ubiquitination of the ribosomal proteins RPS10 and RPS20, critical for ribosome-associated protein quality control (RQC), is dependent on UBE2D3. Moreover, we show by TULIP2 methodology that RPS10 and RPS20 are direct targets of UBE2D3 and demonstrate that UBE2D3’s catalytic activity is required to ubiquitinate RPS10 in vivo. Collectively, our findings show that depletion of an E2 enzyme in combination with quantitative diGly-based ubiquitinome profiling is a powerful tool to identify new in vivo E2 substrates, as we have done here for UBE2D3. Together with our identification of RQC factors as novel in vivo targets of UBE2D3, our work provides an important resource for further studies on the in vivo functions of UBE2D3.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_08:59:31.558.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Jeroen Demmers
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationList	ubiquitinylated lysine
Instrument	Q Exactive HF; Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2022-06-30 07:35:23	ID requested
1	2023-04-13 06:56:03	announced
⏵ 2	2023-11-14 08:59:32	announced	2023-11-14: Updated project metadata.

Dataset with its publication pending

submitter keyword: Ubiquitinome, SILAC, UBE2D3

Jeroen Demmers
contact affiliation	Erasmus MC
contact email	j.demmers@erasmusmc.nl
lab head
Jeroen Demmers
contact affiliation	Proteomics Center, Erasmus University Medical Center, Rotterdam, The Netherlands
contact email	j.demmers@erasmusmc.nl
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD035045
     1. Label: PRIDE project
     2. Name: Ubiquitinome profiling reveals in vivo UBE2D3 targets and implicates UBE2D3 in protein quality control.