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Gene expression profiles of peptide-primed OT-1 cells with or without corticosteroid

ABSTRACT: We report the gene expression profiles of FACS-sorted OT-I cells stimulated with N4 or Y3 peptide with or without corticosteroids.

ORGANISM(S): Mus musculus

PROVIDER: GSE136236 | GEO | 2019/08/24

REPOSITORIES: GEO

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Gene expression profiles of peptide-primed OT-1 cells with or without corticosteroid

Project description:Gene expression profiles of peptide-primed OT-1 cells with or without corticosteroid

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Single-cell RNA sequencing of OT-I CD8+ T cells after stimulation with different affinity ligands

Project description:Activation of CD8+ T cells depends exquisitely on the affinity of the T cell receptor (TCR) for a peptide MHC (pMHC) ligand complex. Here, we activated OT-I transgenic CD8+ T cells with pure peptide and examined early activation responses by single-cell RNA-sequencing. T cells were activated with the high affinity OT-I cognate peptide (N4=SIINFEKL) for 1, 3 or 6 hours, or with reduced affinity peptides (T4=SIITFEKL and G4=SIIGFEKL) or the non-binding peptide (NP68=ASNENMDAM) for 6 hours. Cells were then sorted into 96-well plates by FACS and RNA was sequenced following an adapted Smart-Seq2 protocol.

2018-06-25 | E-MTAB-6051 | biostudies-arrayexpress

Transcriptional effects of IL-6 on peptide-stimulated OT-I CD8 T cells after 2 or 7 days

Project description:Interleukin 6 (IL-6) is a pleiotropic cytokine with diverse roles in homeostasis, inflammation, and cancer. To identify transcriptional patterns and differentiation states induced by IL-6 in CD8 T cells, we performed RNAseq profiling of murine OT-I CD8+ T cells stimulated with SIINFEKL peptide in the presence of exogenous IL-6 (10 ng/ml) or anti-IL6R antibody (to block endogenous IL-6 signaling). Bulk OT-I splenocytes were stimulated with SIINFEKL peptide and CD8 T cells were FACS-sorted for RNA extraction after 2 and 7 days. We found that IL-6 potently repressed classical effector differentiation and promoted a gene expression pattern similar to that of memory precursor cells.

2022-11-04 | GSE199044 | GEO

Time-resolved phosphoproteomic analysis of OT-I CD8+ T cells stimulated with MHC-I tetramers loaded with N4 agonist peptide

Project description:T cell receptor (TCR) signaling is essential for the function of T cells. Here we monitor and quantify the dynamics of phosphorylation sites in OT-I CD8+ TCR transgenic T cells that have been stimulated with pMHC-I tetramers conjugated with the SIINFEKL (N4) peptide for 0, 15, 30, 120, 300 and 600 seconds. Cells were lysed in urea, proteins were digested into tryptic peptides, and phosphorylated peptides were enriched using titanium (TiO2) beads. A fraction of the eluate was analyzed by nanoLC-MS/MS, while the remaining sample was further enriched using phospho-Tyrosine (pTyr) immuno-precipitation. The eluate of the anti-pTyr immuno-precipitation was analyzed by nanoLC-MS/MS in a second serie of MS runs. We performed 3 independent experiments, each containing 6 time-points of stimulation. All samples were analyzed in triplicate MS runs. This dataset contains the raw LC-MS/MS files of the TiO2-enriched peptides (TiO2, 18 samples, 54 MS raw files), and the pTyr enriched peptides (Ptyr, 18 samples, 54 MS raw files).

2020-07-09 | PXD016583 | Pride

Time-resolved phosphoproteomic analysis of murine CD8+ T cells upon stimulation with ligands of different affinity to the TCR

Project description:T cells have the remarkable ability to sense and discriminate between a wide range of antigenic peptides bound to major histocompatibility complex molecules (pMHC). Here, we use phosphoproteomics to monitor in a time dependent manner the signalling cascade taking place upon stimulation of murine CD8+ T cells with ligands of different affinity to the TCR. We used the mouse OT-I model in which T cells specifically recognize the ovalbumin OVA257-264 peptide (SIINFEKL, also named N4) bound to the MHC-I molecule H-2Kb (pMHC). The OT-I TCR also recognizes altered peptide ligands amongst which SIITFEKL (T4) and SIIGFEKL (G4) with sub-optimal binding capacities (ligand affinity N4>T4>G4). CD8+ T cells were purified from pooled lymph nodes and spleens, shortly expanded, and were left unstimulated or stimulated with N4, T4 and G4 soluble pMHC tetramers for 30, 120, 300 or 600s. We conducted six independent multiplexed experiments, each containing either independent stimulations with the N4 and T4 peptides (N4-T4 experiments, 3 replicates) or with the N4 and G4 peptides (N4-G4 experiments, 3 replicates). Samples from each experiment were labeled by TMT10plex, pooled and subjected to phospho-enrichment using first TiO2 beads and then phospho-Tyrosine (pY) immuno-precipitation (IP). Samples were injected before TiO2 enrichment, after TiO2 and after TiO2 followed by pY IP for analysis of the proteome, serine/threonine phospho-proteome and tyrosine phospho-proteome, respectively.

2022-07-05 | PXD030080 | Pride

Analysis of the interactomes of ZAP70, CD247 and SLP76 upon stimulation of murine CD8+ T cells with ligands of different affinity to the TCR

Project description:T cells have the remarkable ability to sense and discriminate between a wide range of antigenic peptides bound to major histocompatibility complex molecules (pMHC). Here, we use interactomics to monitor in a time dependent manner the early molecular events taking place upon stimulation of murine CD8+ T cells with ligands of different affinity to the TCR. We used the mouse OT-I model in which T cells specifically recognize the ovalbumin OVA 257-264 peptide (SIINFEKL, also named N4) bound to the MHC-I molecule H-2Kb (pMHC). The OT-I TCR also recognizes altered peptide ligands amongst which SIITFEKL (T4) and SIIGFEKL (G4) with sub-optimal binding capacities (ligand affinity N4>T4>G4). For affinity purification (AP) of signalling complexes, we crossed OT-I TCR transgenic mice onto mice expressing an endogenous tagged (One-Strep-Tag: OST) version of the CD3z (also called CD247), ZAP70 or SLP76 molecules. CD8+ T cells were purified from pooled lymph nodes and spleens, shortly expanded, and were left unstimulated or stimulated with N4, T4 and G4 soluble pMHC tetramers for 30, 120 or 300 s. Complexes formed around either CD247, ZAP70 or SLP76 were then purified and analysed by mass spectrometry. Each AP-MS purification is associated with a corresponding control (purification from OT-I CD8+ T cells expressing only the endogenous version of each bait, cultured and stimulated in the same conditions). The number of replicate biological experiments was n=3 for all time-points, and each sample was analyzed twice by single-run nano LC-MS. In addition, the whole proteome of OT-I cell was analyzed by label-free shotgun proteomics in order to calculate cellular protein abundances for each interactor.

2022-07-05 | PXD029974 | Pride

Expression data from OT-I CD8 T cells activated with DC in the presence or absence of CpG-Induced Inflammation

Project description:Inflammatory cytokines promote the accumulation of activated CD8 T cells. Here, we transfer 600 OT-I CD8 T cells iv into naÃ¯ve C57BL/6 hosts. One day later, 500,000 LPS-matured and OVA257 peptide-coated DC were injected iv into OT-I CD8 T cell seeded hosts with (DC+CpG) or without (DC). Other seeded mice were infected with 2x10^4 virulent Listeria monocytogenes (vLM-OVA) iv. OT-I CD8 T cells were harvested from the spleen, flow sort purified, then RNA was extracted using RNeasy (Qiagen) kit. Naive OT-I CD8 T cells (Naive) were purified from the spleens of OT-I transgenic mice. Each group had three independent biological replicates.Transcriptomes were compared using DAVID analysis (with genes scoring FDR<0.01) and GSEA analysis. 3 biological replicates per group. Groups included NaÃ¯ve OT-I CD8 T cells, DC+CpG OT-I CD8 T cells, DC OT-I CD8 T cells, and vLM-OVA OT-I CD8 T cells. Most comparisons used NaÃ¯ve OT-I CD8 T cells as a baseline comparison

2013-12-10 | E-GEOD-53200 | biostudies-arrayexpress

Desulfosporosinus sp. OT strain:OT

Project description:Desulfosporosinus sp. OT genome sequencing

| PRJNA61253 | ENA

Effect of loss of PKC theta and p50+cRel on gene expression post T-cell stimulation

Project description:OT-1 Transgenic CD8 T-cells were isolated from spleens of WT, PKCθ theta KO, and p50 cRel DKO mice. The T-cells were either cultured with non-pulsed DC (WT only and signified as "WT - UN") or with BMDCs pulsed with the OVA peptide SIINFEKL (N4) (WT, PKCθ theta KO, and p50 cRel DKO and signified as 'genotype - N4') at a ratio of 1:10 (DC:T-cell) for 18 hours. DCs then were depleted from the culture and RNA was made from the T-cells to measure gene expression at the early / late stage of T-cell activation

2014-12-01 | GSE61758 | GEO

Redirection of therapeutic T cells to a bone marrow niche improves their persistence and anti-tumor functions: transcription profiling by array of resting memory OT-I T_CXCR4) or OT-I T_Control cells isolated from the spleen 90 days following DC vaccination.

Project description:This study reports the improvement in function and antitumor efficacy of CXCR4 overexpression in therapeutic CD8+ T cells. Polyclonal CD8+ T cells from OT-I C57BL/6 (B6) mice were transduced with a modified pMP71 retroviral vector containing murine Cxcr4 and Gfp reporter sequences (T_CXCR4) or with a control vector containing Gfp alone (T_Control), and co-transferred into to B6 mice that were then vaccinated with peptide-pulsed DC. 90 days following DC vaccination, lymphocytes were isolated from the spleen and resting memory OT-I T_CXCR4 and OT-I T_Control cells were FACS sorted to perform gene expression profiling.

2018-03-28 | E-MTAB-6091 | biostudies-arrayexpress

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