PXD061073

PXD061073 is an original dataset announced via ProteomeXchange.

Title	Dissecting Branch-Specific Unfolded Protein Response Activation in Drug-Tolerant BRAF-Mutant Melanoma using Data-Independent Acquisition
Description	Cells rely on the Unfolded Protein Response (UPR) to maintain ER homeostasis (proteostasis) when faced with elevated levels of misfolded and aggregated proteins, which can hinder the secretion of properly folded proteins. The UPR is composed of three main branches—ATF6, IRE1, and PERK—that coordinate the synthesis of proteins involved in folding, trafficking, and degradation of nascent proteins to restore ER function. Dysregulation of the UPR is linked to numerous diseases, including neurodegenerative disorders, cancer, and diabetes. Despite its importance, identifying UPR targets has been challenging due to their heterogeneous induction, which varies by cell type and tissue. Additionally, defining the magnitude and range of UPR-regulated genes is difficult because of intricate temporal regulation, feedback between UPR branches, and extensive cross-talk with other stress-signaling pathways. To comprehensively identify UPR-regulated proteins and determine their branch specificity, we developed a data-independent acquisition (DIA) liquid-chromatography mass spectrometry (LC-MS) pipeline. Our optimized workflow improved protein identifications of low-abundant UPR proteins and leveraged an automated SP3-based protocol on the Biomek i5 liquid handler for label-free peptide preparation. Using engineered stable cell lines that enable selective pharmacological activation of each UPR branch without triggering global UPR activation, we identified branch-specific UPR proteomic targets. These targets were subsequently applied to investigate proteomic changes in multiple BRAF-mutant melanoma cell lines, representing different genetic backgrounds, treated with a BRAF inhibitor (PLX4720, i.e., vemurafenib). Our findings revealed differential regulation of the XBP1s branch of the UPR in the BRAF-mutant melanoma cell lines after PLX4720 treatment, likely due to calcium activation, suggesting that the UPR plays a role as a non-genetic mechanism of drug tolerance in melanoma. In conclusion, the validated branch-specific UPR proteomic targets identified in this study provide a robust framework for investigating this pathway across different cell types, drug treatments, and disease conditions in a high-throughput manner.
HostingRepository	PRIDE
AnnounceDate	2025-08-25
AnnouncementXML	Submission_2025-08-25_08:21:53.753.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD061073
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Lea Barny
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	iodoacetamide derivatized residue
Instrument	Orbitrap Exploris 480

Revision	Datetime	Status	ChangeLog Entry
0	2025-02-21 20:14:53	ID requested
⏵ 1	2025-08-25 08:21:54	announced

Barny LA, Hermanson JN, Garcia SK, Stauffer PE, Plate L, Dissecting Branch-Specific Unfolded Protein Response Activation in Drug-Tolerant BRAF-Mutant Melanoma Using Data-Independent Acquisition Mass Spectrometry. Mol Cell Proteomics, 24(8):101036(2025) [pubmed]

10.6019/PXD061073;

10.1016/j.mcpro.2025.101036;

submitter keyword: Proteomics automation, Activating Transcription Factor 6,Data-Independent Acquisition (DIA), Inositol requiring enzyme 1, Protein Kinase R-like ER Kinase, Unfolded Protein Response

Lars Plate
contact affiliation	Vanderbilt University Department of Chemistry
contact email	lars.plate@vanderbilt.edu
lab head
Lea Barny
contact affiliation	Vanderbilt University
contact email	lea.a.barny@vanderbilt.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD061073
     1. Label: PRIDE project
     2. Name: Dissecting Branch-Specific Unfolded Protein Response Activation in Drug-Tolerant BRAF-Mutant Melanoma using Data-Independent Acquisition