PXD072518

PXD072518 is an original dataset announced via ProteomeXchange.

Title	DIA-PASEF–based total proteome profiling of HeLa cells expressing nuclear or cytosolic p62 condensates
Description	Cells were lysed in 6 M guanidine-HCl, 100 mM HEPES-NaOH, pH7.5, 10 mM TCEP, and 40 mM CAA. The lysates were dissolved by heating and sonication, followed by centrifugation at 20,000 × g for 15 min at 4 °C. The supernatants were recovered, and proteins (20 µg each) were purified by the SP3 method and digested with 0.2 µg trypsin/Lys-C mix (Promega) at 37 °C overnight. The resulting peptide solutions were desalted using GL-Tip SDB (GL Sciences), evaporated in a SpeedVac concentrator, and re-dissolved in 0.1% TFA and 3% ACN. LC-MS/MS analysis of the resultant peptides was performed on a nanoElute 2 coupled with a timsTOF HT mass spectrometer (Bruker). The peptides were separated on a 75-μm inner diameter × 150 mm C18 reversed-phase column (Nikkyo Technos) with a linear 5%–35% ACN gradient for 0–60 min followed by an increase to 95% ACN for 1 min and a final hold at 80% ACN for 4 min. Data acquisition was conducted in dia-PASEF mode with an MS1 m/z range of 100–1700 and an ion mobility (1/K0) range of 0.6–1.6. The ramp time was set to 100 ms with a 100% duty cycle. The MS2 polygon was manually defined based on the region where peptides were predominantly detected, specifically using the following four vertices: Point 1 (m/z = 300, 1/K0 = 0.64), Point 2 (m/z = 300, 1/K0 = 0.8), Point 3 (m/z = 770, 1/K0 = 0.97), and Point 4 (m/z = 770, 1/K0 = 1.15). Within the defined polygon, a 8 Da isolation window was employed with an overlap of 0.5 Da for each window, resulting in 62 windows per cycle and a cycle time of 3.36 s. Protein identification was carried out in DIA-NN (version 1.9) against a human in silico spectral library. Parameters for constructing the library included: trypsin as the digestion enzyme, one allowed missed cleavage, peptide lengths ranging from 7 to 45 amino acids, precursor charges of 2 to 4, and a fragment ion m/z range of 200–1800. Features such as library-free search, deep learning-based retention time and ion mobility predictions, N-terminal methionine cleavage, and cysteine carbamidomethylation were enabled. In the DIA-NN search settings, MS1 and MS2 mass accuracies were set to auto, neural network classifiers were configured for single-pass mode, and the quantification strategy was set to QuantUMS (high precision).
HostingRepository	jPOST
AnnounceDate	2025-12-30
AnnouncementXML	Submission_2026-02-07_03:54:37.860.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Hidetaka Kosako
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	L-methionine sulfoxide; Acetyl; S-carboxamidomethyl-L-cysteine
Instrument	instrument

Revision	Datetime	Status	ChangeLog Entry
0	2025-12-30 05:10:42	ID requested
1	2025-12-30 05:11:46	announced
⏵ 2	2026-02-07 03:54:38	announced	2026-02-07: Updated PubMed.

Lulu-Shimron C, Luo Z, Brekhman V, Huang L, Livneh I, Kosako H, Cohen-Kaplan V, Ciechanover A, Proteasomal proteolysis in p62 condensates directs tumor suppression or growth depending on their subcellular localization. Proc Natl Acad Sci U S A, 123(3):e2529422123(2026) [pubmed]

submitter keyword: DIA-PASEF, HeLa, p62 condensates

Aaron Ciechanover
lab head
Hidetaka Kosako
contact affiliation	Tokushima University
dataset submitter

jPOST dataset URI

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