Dataset Information

Neutrophil-derived migrasomes are an essential part of the coagulation system

ABSTRACT: Neutrophil-derived migrasomes are an essential part of the coagulation system, human

INSTRUMENT(S): Orbitrap Ascend

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood

SUBMITTER: Dong Jiang

LAB HEAD: Li Yu

PROVIDER: PXD051231 | Pride | 2024-05-07

REPOSITORIES: Pride

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Dataset's files

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			Action	DRS
	20221219_174341_MS221726-Report.xls	Xls
	MS221726-MIGRASOME-1.raw	Raw
	MS221726-MIGRASOME-2.raw	Raw
	MS221726-MIGRASOME-3.raw	Raw
	MS221726-PLATELET-1_20221213010717.raw	Raw

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Project 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.

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Neutrophil-derived migrasomes are an essential part of the coagulation system

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