Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Total RNA: 24h activated mouse OT-1 T lymphocytes, WT vs Lck-/- (OFF)

ABSTRACT: This experiment aims to study the transcriptome of activated T cells at 24h, using the same biological samples as the polysome profiling in GSE84781. A T cell receptor (TCR) transgenic mouse model (OT-1) posessing a homogenous alpha-beta CD8 T lymphocyte population was used, and cells with intact TCR signalling pathways (WT) were compared to those with defective signal transduction (Lck-/-, OFF). Ex vivo OT-1 T cells were isolated from the superficial cervicals, axillary, brachial, mesenteric, and inguinal lymph nodes and then activated for 24h with cognate peptide antigen SIINFEKL. Total RNA was extracted from the cells, labelled with Cy3, and hybridised onto an Agilent-074809 SurePrint G3 Mouse GE v2 8x60K Microarray.

ORGANISM(S): Mus musculus

PROVIDER: GSE84856 | GEO | 2016/11/30

SECONDARY ACCESSION(S): PRJNA335377

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Similar Datasets

Total RNA: naïve ex vivo mouse OT-1 T lymphocytes, WT vs Lck-/- (OFF)

Project description:This experiment aims to study the transcriptome of naïve T cells at the steady state. A T cell receptor (TCR) transgenic mouse model (OT-1) posessing a homogenous alpha-beta CD8 T lymphocyte population was used, and cells with intact TCR signalling pathways (WT) were compared to those with defective signal transduction (Lck-/-, OFF). Naïve (CD8b+, CD44low) ex vivo T cells were isolated from the superficial cervicals, axillary, brachial, mesenteric, and inguinal lymph nodes. Total RNA was extracted from the cells, labelled with Cy3, and hybridised onto an Agilent-074809 SurePrint G3 Mouse GE v2 8x60K Microarray.

2016-11-30 | GSE84855 | GEO

Polysome profiling: Mouse OT-1 T cells WT vs Lck-/- (OFF)

Project description:In order to identify gene targets of translational regulation during T cell activation, a polysome analysis was performed in a T cell receptor (TCR) transgenic model (OT-1) possessing a homogenous alpha-beta CD8 T lymphocyte population. Cells with defective TCR signal tranduction (Lck-/-, OFF) were compared to WT cells to visualise targets regulated by TCR signalling. Total cytoplasmic ribonucleoprotein was extracted from ex vivo OT-1 transgenic T lymphocytes stimulated with SIINFEKL peptide antigen for 24h, extracts were fractionated using a sucrose density gradient, and separated into a sub-polysomal and a polysomal fraction. Total RNA from each fraction was extracted, equal volume of RNA from each fraction was individually labelled (Sub-poly: Cy3; Poly: Cy5), and hybridised onto a Agilent SurePrint G3 Mouse GE v2 8x60K Microarray.

2016-11-30 | GSE84781 | GEO

OT-I T cell activation in Lrrc8af/f and Lrrc8af/f Cd4-Cre;OT-I mice

Project description:Splenocytes were separated from Lrrc8af/f and Lrrc8af/f Cd4-Cre;OT-I mice. The splenocytes were stimulated with indicated peptides (unstime/ SIINFEKL-10pM(NL)/ SIINFEKL-1000pM(NH)/ SIIGFEKL-1000nM(G4)) for 6 hours. The OT-I CD8+ T cells were labeled with Va2-PE/CD8-A700 and sorted for the sequencing.

2023-09-26 | GSE243985 | GEO

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

Transcription profiling of mouse splenocytes from OT-1 TCR transgenic mice to assess the effect of IL-12 on CTL gene expression

Project description:The goal was to determine how IL-12 affects gene expression by murine CTL. Experiment Overall Design: Following RBC lysis, splenocytes from OT-1 TCR transgenic mice (5 million/ml) were cultured in six well-plates (5-6ml/well) in IMDM supplemented with 10% FCS and 55Î¼M 2-ME with 1Î¼M SIINFEKL. After 4 days, the cells were harvested by purification with Ficoll-Paque PLUS and placed in fresh medium containing 0 or 20ng/ml rmIL-12. Twenty-four hours later the cells were harvested and used in experiments. The live cells harvested were >98% CD8+VÎ±2+ (OT-1).

2009-02-08 | E-GEOD-13173 | biostudies-arrayexpress

OT-1 RNA seq day36

Project description:OT-1 RNA seq day36 MCMV-SIINFEKL vs RAE-1gMCMV-SIINFEKL

| PRJEB44407 | ENA

Quantitative interactomics of Lck-TurboID in living human T cells unveils T cell receptor stimulation-induced proximal Lck interactors

Project description:While Lck has been widely recognized to play a pivotal role in the initiation of the T cell receptor (TCR) signaling pathway, an understanding of the precise regulation of Lck in T cells upon TCR activation remains elusive. Investigation of protein-protein interaction (PPI) using proximity labeling techniques such as TurboID has the potential to provide valuable molecular insights into Lck regulatory networks. By expressing Lck-TurboID in Jurkat T cells, we have uncovered a dynamic, short-range Lck protein interaction network upon 30 minutes of TCR stimulation. In this novel application of TurboID, we detected 27 TCR stimulation-induced Lck-proximal interactors in living T cells, including both bona fide and uncanonical Lck interactors, validating the discovery power of this tool. Our results revealed previously unappreciated Lck PPI which may be associated with cytoskeletal rearrangement, ubiquitination of TCR signaling proteins, activation of the MAPK cascade, coalescence of the LAT signalosome, and formation of the immunological synapse. In this study, we demonstrated for the first time in immune cells and for the kinase Lck that TurboID can be utilized to unveil PPI dynamics in living cells at a time scale consistent with TCR signaling.

2020-11-12 | PXD020759 | Pride

Gene expression profiles of OT-1 T cells co-cultured with MHC-I Negative Tumors and Antigen Loaded Macrophages

Project description:OT-1 T-cells were co-cultured either alone in T-cell media supplemented with IL-2 (50IU/mL IL-2), with OVA loaded macrophages, or with both OVA loaded macrophages and CT2A-TRP2-β2mKO tumor cells. Cells were cultured at a 5:1 T-cell to tumor ratio, and 2:1 T-cell to macrophage ratio. After 24 h of co-culture, CD8 T-cells were FACS sorted, and RNA extracted (RNeasy Mini Kit, Qiagen). RNA was analyzed on an nCounter MAX Analysis System (Nanostring) with the PanCancer Immune profiling panel (Nanostring) according to manufacturer instructions. Expression dat OT-1 T-cells were isolated from OT-1 mice by culturing OT-1 splenocytes in T-cell media supplemented with 50IU/mL IL-2 and 1 μM OVA SIINFEKL peptide (Anaspec) for 48 h. Cells were purified for CD8 T-cells as described above and subsequently cultured in TCM with 50IU/mL IL-2, splitting every 24h for a total of 4 days.

2023-05-22 | GSE220960 | GEO

Lck promotes Zap70-dependent LAT phosphorylation by bridging Zap70 to LAT

Project description:T cell antigen receptor (TCR) signaling depends upon the kinases Lck and Zap70. Lck phosphorylates the TCR, facilitating Zap70 recruitment to the stimulated TCR. Lck also phosphorylates Zap70, relieving its auto-inhibition and activating its catalytic domain. Zap70 then phosphorylates the critical adaptors LAT and SLP76 which serve to nucleate key effector molecules required for downstream responses. However, mechanisms facilitating the interaction of Zap70 with its substrates have not been described. We report an evolutionarily conserved proline-rich motif in LAT is important for Zap70-induced phosphorylation of LAT and downstream signaling. This LAT proline-rich motif associated with the Lck SH3 domain, thereby facilitating Zap70-mediated phosphorylation of LAT and downstream functions. Our results suggest Lck orchestrates multiple steps in TCR signaling including the newly described facilitation of the interaction of Zap70 with its substrate LAT. This previously unrecognized feature of TCR proximal signaling may contribute to the development of more immunomodulatory therapies.

2019-11-12 | PXD008258 | Pride

Lck phosphorylation status of Lck Tyr192 and Tyr192/Tyr505 mutantsLck phosphorylation status of Lck Tyr192 and Tyr192/Tyr505 mutants

Project description:Lymphocyte-specific protein tyrosine kinase (Lck) is crucial for signaling from the T cell receptor (TCR) and is controlled through tyrosine phosphorylations. Phosphorylated Tyr505 (pTyr505) promotes a closed, inactive conformation of Lck, while pTyr394 is critical for kinase activity. Additionally, pTyr192 has been suggested to regulate Lck activity by changing the specificity of the Lck Src-homology 2 (SH2) domain and/or by affecting Lck association with CD45 thus drastically increasing pTyr505. However, little is known about how pTyr192 affects endogenously expressed Lck. Here we used CRISPR/Cas9 genome editing to generate Jurkat cell lines expressing Lck Glu192 mimicking Lck pTyr192, or Lck Phe192 mimicking unphosphorylated Lck Tyr192. We confirmed that Lck Glu192 is hyperphosphorylated on Tyr505, possibly explaining reduced association of Lck Glu192 with prototypic Lck-SH2 ligands. To isolate the effect of Lck Tyr192 mutations from the effect on Lck pTyr505, we subsequently generated Jurkat cells doubly mutated on Lck Tyr192 and Lck Tyr505. Both Lck Phe192/Phe505 and Lck Glu192/Phe505 mutants co-precipitated similar amounts of binding partners. Moreover, both mutants displayed hyperphosphorylation of Tyr394. Our results indicate that CD45 is the main phosphatase controlling Lck pTyr394 in steady-state T cells. Additionally, our data demonstrate that the prototypic specificity of the Lck SH2 domain (owned by the Lck Phe192 mutants) promotes transphosphorylation of Tyr394. These observations pinpoint the fundamental role of Tyr192 in regulation of Lck activity and simultaneously reveal the most potent Lck mutants so far described

2021-04-02 | PXD014470 | Pride

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data