PXD069927

PXD069927 is an original dataset announced via ProteomeXchange.

Title	IP-MS analysis for identifying NSD3-short-interacting proteins
Description	To identify NSD3-short–interacting proteins, HA-tagged NSD3-short was overexpressed in 293T cells. Immunoprecipitation was performed using antibodies against HA and NSD3, followed by mass spectrometry analysis to identify NSD3-short–associated proteins. Cells were lysed on ice for 1 h in buffer containing 50 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1% Triton X-100, 5 mM EDTA, and 8% glycerol, supplemented with protease inhibitors and 1 mM DTT. Lysates were incubated overnight at 4 °C with Protein A/G beads (25 μL per IP) pre-bound to HA-tag or anti-NSD3 antibody. Beads were washed five times with washing buffer (20 mM Tris-HCl, pH 7.5, 150 mM NaCl, 0.1% Triton X-100, 1 mM EDTA), and bound proteins were eluted by heating in 1× SDS loading buffer at 100 °C for 10 min. Bead-bound proteins were reduced with 10 mM DTT for 1 h and alkylated with 25 mM iodoacetamide for 20 min in the dark at room temperature. After washing with 25 mM ammonium bicarbonate, on-bead digestion was performed with sequencing-grade trypsin at 37 °C. Peptides were desalted, dissolved in 0.1% formic acid, and analyzed on an Easy-nLC 1200 system coupled to a Q Exactive Plus mass spectrometer. Peptides were separated using a linear acetonitrile gradient (2–35% solvent B in 0.1% formic acid) at a flow rate of 300 nL/min. MS1 spectra (m/z 350–1800) were acquired at a resolution of 70,000, and the top 20 precursor ions (charge state ≥ +1) were fragmented by higher-energy collisional dissociation (HCD) and analyzed at a resolution of 17,500. Raw data were processed with Proteome Discoverer 3.0 using label-free quantification (Top3 method), and differential protein abundance was determined by one-way ANOVA.
HostingRepository	iProX
AnnounceDate	2025-10-27
AnnouncementXML	Submission_2025-10-26_19:33:40.105.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Zhengke Lian
SpeciesList	scientific name: Homo sapiens; NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	Q Exactive Plus

Revision	Datetime	Status	ChangeLog Entry
0	2025-10-26 19:33:25	ID requested
⏵ 1	2025-10-26 19:33:40	announced

Dataset with its publication pending

submitter keyword: NSD3-short, HDAC complex

Xiufeng Pang
contact affiliation	East China Normal University
contact email	xfpang@bio.ecnu.edu.cn
lab head
Zhengke Lian
contact affiliation	East China Normal University
contact email	Valient17@outlook.com
dataset submitter

iProX dataset URI