Effect of depletion of TFII-I or TRIM24 on Jurkat T cell activation.
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ABSTRACT: To investigate the cooperative function TFII-I and TRIM24 in the regulation of T cell activation regulated genes, we established Jurkat Tat cells in which TFII-I was depleted by shRNA and TRIM24 was knocked out by CRISPR-Cas9. We then performed gene expression profiling analysis using data obtained from RNA-seq of cell lines stimulated by PMA and Ionomycin.
Project description:To investigate the cooperative function USF factors for the regulation of T cell transcriptom, we established Jurkat mHIV-Luciferase cells in which USF1 or USF2 was knocked out by CRISPR-Cas9. We then performed gene expression profiling analysis using data obtained from RNA-seq of cell lines either left untreated or stimulated by PMA and Ionomycin co-treatment.
Project description:To assess the effects of small molecules SP100030 and Selinexor on Jurkat T cells activated by PMA/Ionomycin after 6 hours of activation and treatment
Project description:In order to determine the calcineurin inhibitory effect of CABIN1 peptide, we performed RNA-sequencing in Jurkat T cells expressing negative contorl (HA-mCherry) or HA-mCherry-CABIN1 peptide. Jurkat T cells were activated by treatment 40 nM PMA and 1 μM Ionomycin for 8 hr. 0.5 μM FK506 (Tacrolimus, Tac) was pretreated for 1 hr before treatment with PMA and Ionomycin. Total RNA was extracted from these cells. Extracted RNA was used to prepare an mRNA sequencing library using TruSeq Stranded mRNA sample preparation kit. All samples were sequenced on Illumina NextSeq 500 with a 75 bp paired end read.
Project description:We used microarray to monitor the differentially expresed genes during Jurkat T cell activaiton. Jurkat T cells with control shRNA or IKKe shRNA were treated with solvent or PMA and ionomycin for 3 h, and then RNA was extracted and applied to microarray analysis
Project description:A quantitative label-free secretome analysis protocol using a click chemistry-based approach for the enrichment of secreted glycoproteins was adapted for and applied to a T cell model. There, Jurkat cells were activated via PMA/ionomycin and the dynamic modulation of the T cell secretome was investigated and compared to the dynamic modulation of the T cell proteome of the same model.
Project description:Purpose: study the role of MALT1 auto-proteolysis in T cell receptor mediated activation of NF-kB. Methods: Jurkat cells were generated that express wild type MALT1, the auto-cleavage deficient MALT1-R149A mutant, the catalytic inactive MALT1-C464A mutant or the R149A-C464A double mutant (RACA). Expression of endogenous MALT1 was inactivated using TALEN technology for the Jurkat cells expressing MALT1-R149A (JDM-RA) and MALT1-C464A (JDM-CA). Illumina HISeq 2000 deep sequencing was performed to determine the mRNA profiles for MALT1, JDM-RA, JDM-CA and RACA cells in unstimulated conditions or after treatment with 75ng/ml PMA and 150 ng/ml ionomycin for 3 or 18 hrs. Results: PMA ionomycin stimulation of the MALT1 auto-cleavage defective JDM-RA cells fails to activate NF-kB-dependent transcription like for the MALT1 catalytic inactive JDM-CA cells and the double RACA mutant cells. Conclusion: MALT1 autoproteolysis is essential for transcription of NF-kB target genes
Project description:The transcription factor (TF) Forkhead Box P3 (FOXP3) is constitutively expressed in high levels in natural occurring CD4+CD25+ regulatory T cells (nTreg) and is not only the most accepted marker for that cell population, but is considered lineage determinative. Chromatin immunoprecipitation (ChIP) of transcription factors in combination with genomic tiling microarray analysis (ChIP-on-Chip) has been shown to be an appropriate tool to identify FOXP3 transcription factor binding sites (TFBS) on a genome-wide scale. In combination with microarray expression analysis the ChIP-on-Chip technique allows to identify direct FOXP3 target genes. ChIP-on-Chip analysis of human FOXP3M-NM-^T2 isoform expressed in resting and PMA / ionomycin stimulated Jurkat T cells revealed several thousand putative FOXP3 binding sites and importance of intronic regions for FOXP3 binding. Knowledge of general distribution patterns of FOXP3 TFBS in the human genome under resting and activated conditions contributes to a better understanding of this TF and its influence on direct target genes with importance for Treg cell phenotype and function. ChIP-DNA from FOXP3(M-NM-^T2) expressing Jurkat T cells under resting and PMA / ionomycin stimulated conditions from duplicate experiments was analyzed. FOXP3-specific tiling array data were analyzed in reference to an individual isotype control dataset (J-FOXP3 ChIP'd with FOXP3 antibody vs. J-FOXP3 ChIP'd with isotype control antibody). In total 8 tiling array analyses were performed (2x resting J-FOXP3 with FOXP3-IP, 2x resting J-FOXP3 with isotype-IP, 2x PMA/iono J-FOXP3 with FOXP3-IP, 2x PMA/iono J-FOXP3 with isotype-IP)
Project description:The transcription factor (TF) Forkhead Box P3 (FOXP3) is constitutively expressed in high levels in natural occurring CD4+CD25+ regulatory T cells (nTreg) and is not only the most accepted marker for that cell population, but is considered lineage determinative. Chromatin immunoprecipitation (ChIP) of transcription factors in combination with genomic tiling microarray analysis (ChIP-on-Chip) has been shown to be an appropriate tool to identify FOXP3 transcription factor binding sites (TFBS) on a genome-wide scale. In combination with microarray expression analysis the ChIP-on-Chip technique allows to identify direct FOXP3 target genes. This dataset shows expression data of resting and mitogen stimulated (PMA / ionomycin) retrovirally transduced Jurkat T cells either expressing FOXP3(M-NM-^T2) (J-FOXP3) or an empty vector control (J-GFP). Expression profile of resting and PMA/ionomycin stimulated J-GFP and J-FOXP3 cells was analyzed (one microarray per condition).