PXD068808

PXD068808 is an original dataset announced via ProteomeXchange.

Title	Global ISGylome analysis to identify substrates of SARS-CoV-2 Nsp3-PLpro
Description	Interferon-stimulated gene 15 (ISG15) regulates diverse cellular processes, including antiviral immunity, through its conjugation to proteins (ISGylation). Several viruses, including SARS-CoV-2, reverse the process of protein ISGylation by encoding specialized enzymes harboring deISGylating enzymatic activity. The papain-like protease (PLpro), which is part of the nonstructural protein 3 (Nsp3) of SARS-CoV-2, antagonizes mammalian host innate defense pathways by removing ISG15 from certain cellular substrates. However, the full substrate landscape of Nsp3-PLpro’s deISGylating activity is unknown. Moreover, PLpro, as part of Nsp3, localizes to double-membrane vesicles (DMVs), the sites where coronaviruses replicate inside cells; thus, this unique subcellular localization of Nsp3-PLpro is expected to restrain its substrate specificity. Here, we expressed in human cells a SARS-CoV-2 Nsp3-4 fusion construct that produces mature Nsp3 and Nsp4 via PLpro-mediated autoprocessing and promotes DMV formation, followed by IFNα treatment to mimic an infected state. Cells expressing empty vector with or without IFNα treatment, as well as cells expressing Nsp3-4 without IFNα stimulation were included for comparison. LC-MS/MS analysis identified numerous substrates of PLpro-mediated deISGylation that play roles in several major biological processes, including innate immune signaling, translational control, RNA metabolism, and oxidative stress responses. This global ISGylome dataset provides novel insights into how ISGylation regulates host cells at the molecular level and, further, how SARS-CoV-2 Nsp3-PLpro-mediated deISGylation influences the host ISGylome.
HostingRepository	PRIDE
AnnounceDate	2026-05-05
AnnouncementXML	Submission_2026-05-05_14:54:51.418.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Junji Zhu
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: NEWT:9606;
ModificationList	No PTMs are included in the dataset
Instrument	timsTOF Pro 2

Revision	Datetime	Status	ChangeLog Entry
0	2025-09-24 19:53:05	ID requested
⏵ 1	2026-05-05 14:54:52	announced

Dataset with its publication pending

submitter keyword: SARS-CoV-2, PLpro, ISG15, deISGylation, ISGylome, LC-MS/MS

Michaela Gack
contact affiliation	Florida Research and Innovation Center, Cleveland Clinic, Port Saint Lucie, FL 34987, USA
contact email	GACKM@ccf.org
lab head
Junji Zhu
contact affiliation	Florida Research and innovation center, Cleveland Clinic
contact email	zhuj6@ccf.org
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2026/05/PXD068808

PRIDE project URI

• PRIDE
  1. PXD068808
     1. Label: PRIDE project
     2. Name: Global ISGylome analysis to identify substrates of SARS-CoV-2 Nsp3-PLpro