PXD008411

PXD008411 is an original dataset announced via ProteomeXchange.

Title	Patient-derived KEAP1 superbinder mutants stabilize KEAP1 structure and sequester NRF2 in p62-dependent clusters
Description	Cancer-derived loss-of-function mutations in the KEAP1 tumor suppressor gene stabilize the NRF2 transcription factor, resulting in a pro-survival gene expression program that alters cellular metabolism and neutralizes oxidative stress. In a previous study of KEAP1 mutations observed in lung cancer, we classified 40% of the mutations as ‘superbinders’ (superbinders). These mutants bind and ubiquitylate NRF2 but do not promote NRF2 degradation. Here, we further investigated the molecular mechanism(s) driving the superbinder phenotype. BioID-based quantitative proteomic analysis of the R320Q and R470C superbinder mutations revealed increased co-complexed NRF2 without significant alteration to other KEAP1-associated proteins, including CUL3, VCP, and several ubiquitin receptors within the proteasome lid. Dynamic simulation modeling and limited proteolysis analyses suggest that superbinder mutations stabilize residues in KEAP1 that contact NRF2. In cells, KEAP1 R320Q and R470C mutants co-localize with NRF2, p62/SQSTM1 and polyubiquitin in spherical clusters that rapidly fuse and dissolve; KEAP1-NRF2 localization to these clusters requires p62. Expression of R320Q and R470C in lung cancer cells provided resistance to the reactive oxygen species-inducing drug bleomycin. We present a model wherein superbinder mutations alter the conformational dynamics of the KEAP1-NRF2 complex to alter the cycling of KEAP1 between open and closed conformations, thus inhibiting NRF2 degradation.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_04:42:22.292.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Dennis Goldfarb
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	biotinylated residue; phosphorylated residue; monohydroxylated residue; acetylated residue; iodoacetamide derivatized residue
Instrument	LTQ Orbitrap Elite

Revision	Datetime	Status	ChangeLog Entry
0	2017-12-11 06:04:45	ID requested
1	2018-11-07 01:38:43	announced
⏵ 2	2024-10-22 04:42:30	announced	2024-10-22: Updated project metadata.

Cloer EW, Siesser PF, Cousins EM, Goldfarb D, Mowrey DD, Harrison JS, Weir SJ, Dokholyan NV, Major MB, p62-Dependent Phase Separation of Patient-Derived KEAP1 Mutations and NRF2. Mol Cell Biol, 38(22):(2018) [pubmed]

10.1128/mcb.00644-17;

curator keyword: Biomedical

submitter keyword: NRF2,KEAP1, superbinders, p62, clusters, mutations, lung cancer

Ben Major
contact affiliation	Department of Cell Biology and Physiology, University of North Carolina at Chapel Hill School of Medicine, Chapel Hill, United States.
contact email	ben_major@med.unc.edu
lab head
Dennis Goldfarb
contact affiliation	Cell Biology and Physiology Institute for Informatics
contact email	d.goldfarb@wustl.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD008411
     1. Label: PRIDE project
     2. Name: Patient-derived KEAP1 superbinder mutants stabilize KEAP1 structure and sequester NRF2 in p62-dependent clusters