Project description:To investigate the internal regulatory mechanisms governing the CD28 gene region during T cell activation and to screen for truly active dynamic chromatin regulatory elements responsive to stimulation, we employed the CRISPRa gene editing technology to design specific sgRNAs targeting this region and conducted a tilling CRISPRa screen sequencing on the CD28 gene region.
Project description:To meticulously explore how local chromatin architecture influences T cell activation, we conducted high-resolution regional capture Hi-C by designing tiling capture probes that span the CD28, CTLA4, and ICOS genes along with their distal regulatory sequences (chr2:204300000-205000000, GRCh37) in both resting and stimulated Jurkat cells.
Project description:We previously performed a tiling CRISPR activation (CRISPRa) screen in Jurkat T cells to discover enhancers controlling IL2RA expression. This screen identified six regions where recruitment of dCas9-VP64 was sufficient to drive increased expression of IL2RA. We named these putative enhancers CRISPRa Responsive Elements (CaREs). To examine transcriptome-wide consequences of dCas9-VP64 recruitment to IL2RA CaREs, we performed RNA-Seq on HuT78 cells stably expressing dCas9-VP64 and gRNAs targeting the IL2RA TSS, CaRE3, or CaRE4, or stimulated with anti-CD3/CD28 antibodies.
Project description:To finely dissect the dynamic changes in the epigenome and chromatin state during T cell activation, we utilized the Jurkat cell line as a model and performed ChIP-seq on both resting and anti-CD3/CD28-stimulated Jurkat cells, targeting various histone modifications (H3K27ac, H3K4me1, H3K4me3, H3K27me3), RNA PolII, and CTCF.
Project description:The majority of genetic risk variants associated with autoimmune diseases are located in chromatin regions that are accessible during T cell stimulation. To systematically investigate the dynamic changes in chromatin accessibility during T cell activation, we utilized the Jurkat cell line as a model and performed ATAC-seq on both resting and anti-CD3/CD28-stimulated Jurkat cells.
Project description:T lymphocytes are orchestrators of adaptive immunity. Naïve T cells may differentiate into the Th1, Th2, Th17 or iTreg phenotype, depending on environmental co-stimulatory signals. In order to identify the genes and pathways involved in differentiation of Jurkat T cells towards Th1 and Th2 subtypes we performed comprehensive transcriptome analyses of Jurkat T cells stimulated with various stimuli an pathway inhibitors Jurkat T cells were treated with CD3/CD28 and CD3/PMA and CD28/PMA. RNA was isolated after 1 and 8 hrs stumalation.
Project description:Fe-IMAC phosphoproteomics using TMT 11plex for quant, of Jurkat T cells stimulated with CD3 and CD28 agonist antibodies for 0, 3, 9, or 27 minutes.
Project description:T lymphocytes are orchestrators of adaptive immunity. Naïve T cells may differentiate into the Th1, Th2, Th17 or iTreg phenotype, depending on environmental co-stimulatory signals. In order to identify the genes and pathways involved in differentiation of Jurkat T cells towards Th1 and Th2 subtypes we performed comprehensive transcriptome analyses of Jurkat T cells stimulated with various stimuli an pathway inhibitors Jurkat T cells were treated with CD3, CD28 and PMA and all pairwise combinations, in the presence of DMSO (control) or kinase inhibitors. RNA was isolated after 8 hrs incubation.
Project description:To investigate the interacting loci within the region containing rs5837875, we employed the circularized chromosome conformation capture (4C) assay to elucidate the spatial interactions and regulatory networks associated with rs5837875, utilizing this specific region as the bait in Jurkat cells.