PXD053863-1

PXD053863 is an original dataset announced via ProteomeXchange.

Title	Deep phosphotyrosine characterization of mouse primary T cells using Broad Spectrum Optimization of Selective Triggering
Description	Sequencing the tyrosine phosphoproteome using MS-based proteomics is challenging due to the low abundance of tyrosine phosphorylation in cells, a challenge compounded in scarce samples like primary cells or clinical samples. The broad-spectrum optimization of selective triggering (BOOST) method was recently developed to increase phosphotyrosine sequencing in low protein input samples by leveraging tandem mass tags (TMT), phosphotyrosine enrichment, and a phosphotyrosine-loaded carrier channel. Here, we demonstrate the viability of BOOST in T cell receptor (TCR)-stimulated primary murine T cells by benchmarking the accuracy and precision of the BOOST method and discerning significant alterations in the phosphoproteome associated with receptor stimulation. Using 1 milligram of protein input (about 20 million cells) and BOOST, we identify and precisely quantify more than 2,000 unique pY sites compared to about 300 unique pY sites in non-BOOST control samples. We show that although replicate variation increases when using the BOOST method, BOOST does not jeopardise quantitative precision or the ability to determine statistical significance for peptides measured in triplicate. Many pY previously uncharacterised sites on important T cell signalling proteins are quantified using BOOST, and we identify new TCR responsive pY sites observable only with BOOST. Finally, we determine that the phase-spectrum deconvolution method on Orbitrap instruments can impair pY quantitation in BOOST experiments.
HostingRepository	PRIDE
AnnounceDate	2025-12-08
AnnouncementXML	Submission_2025-12-07_17:27:55.773.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Arthur Salomon
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: NEWT:10090;
ModificationList	phosphorylated residue
Instrument	Orbitrap Eclipse

Revision	Datetime	Status	ChangeLog Entry
0	2024-07-10 15:40:10	ID requested
⏵ 1	2025-12-07 17:27:56	announced

10.1002/pmic.202400106;

Callahan A, Chua XY, Griffith AA, Hildebrandt T, Fu G, Hu M, Wen R, Salomon AR, Deep phosphotyrosine characterisation of primary murine T cells using broad spectrum optimisation of selective triggering. Proteomics, 24(23-24):e2400106(2024) [pubmed]

submitter keyword: T cell,Mus musculus, TMT

Arthur Robert Salomon
contact affiliation	Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Providence, Rhode Island, USA
contact email	drsalomon@gmail.com
lab head
Arthur Salomon
contact affiliation	Brown University
contact email	art@drsalomon.com
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/2025/12/PXD053863

PRIDE project URI

• PRIDE
  1. PXD053863
     1. Label: PRIDE project
     2. Name: Deep phosphotyrosine characterization of mouse primary T cells using Broad Spectrum Optimization of Selective Triggering