PXD036793

PXD036793 is an original dataset announced via ProteomeXchange.

Title	Proteomic Analysis for Cul5 E3 Ligase Complex as a Key Negative Feedback Regulator of TCR/Il2 Signaling and Anti-Tumor Activity in Cd8+ T Cells
Description	To investigate the mechanism by which Cul5 regulates CD8+ T cell cytokine-dependent differentiation and TCR-dependent activation, we performed quantitative mass spectrometry (MS) by data-independent acquisition (DIA)-MS of total proteins in the Cul5 KO and NC primary CD8+ T cells in the following conditions: 1) Cytokine dependent expansion and differentiation (T0); 2) 8 hours cytokine withdraw prior to TCR stimulation (T8); and 3) 16 hours TCR stimulation post 8-hour cytokine withdraw (T16). Principal component analysis (PCA) and correlation analysis revealed that replicates in each condition clustered together while different conditions separated from each other, suggesting significant proteomic changes among different conditions of the same T cells as well as between Cul5 KO and NC T cells in each condition. Together with similar total protein quantities among all detected samples, DIA-MS analyses were of high quality. Additionally, the strong reductions of Cul5 abundances in the Cul5 KO cells from all three conditions compared to the NC cells confirms high KO efficiency. Of note, the proteomic analysis showed that the Cul5 protein level was significantly increased upon TCR stimulation post cytokine starvation in the NC cells, suggesting a potential negative feedback regulatory role of Cul5 in CD8+ T cell activation. Consistent with this idea, we observed more markedly upregulation of Cul5 expression upon of TCR stimulation of na•ve primary CD8+ cells. To identify Cul5 interacting proteins in CD8+ T cells, we overexpressed Cul5 with a C terminal HA-tag (Cul5-HA) in mouse primary CD8+ T cells by retroviral transduction. The cells were subjected to TCR stimulation for 12 hr, and Cul5-HA was immunoprecipitated by anti-HA, followed by DIA-MS analysis (co-IP-MS). Compared to the negative control samples (cells transduced with the empty vector), 65 proteins were enriched (p value <0.05 and fold change >1.5) in the anti-HA IP samples. Altogether, we report that Cul5 KO alters CD8+ T cell proteome and the Cul5 interactome.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_06:19:35.362.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Wenxue Li
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationList	acetylated residue; iodoacetamide derivatized residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2022-09-17 02:53:51	ID requested
1	2024-01-08 09:43:15	announced
⏵ 2	2024-10-22 06:19:35	announced	2024-10-22: Updated project metadata.

10.1101/2022.11.16.516824;

submitter keyword: CRISPR,Proteomics, Cul5,CD8 T cells

Yansheng Liu
contact affiliation	Yale Cancer Biology Institute, Yale University, West Haven, CT 06516, USA.
contact email	yansheng.liu@yale.edu
lab head
Wenxue Li
contact affiliation	Yale University
contact email	wenxue.li@yale.edu
dataset submitter

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PRIDE project URI

• PRIDE
  1. PXD036793
     1. Label: PRIDE project
     2. Name: Proteomic Analysis for Cul5 E3 Ligase Complex as a Key Negative Feedback Regulator of TCR/Il2 Signaling and Anti-Tumor Activity in Cd8+ T Cells