ABSTRACT: To better understand the mechanisms downstream of IL-27 signaling, we performed RNA sequencing on repetitively stimulated OT-I T cells treated with IL-27.
Project description:To better understand the mechanisms downstream of IL-27 signaling, we performed ATAC sequencing on repetitively stimulated OT-I T cells treated with IL-27.
Project description:The precise timing and pathway of memory CD8+ T cells differentiation from naïve T cells have remained undetermined. We found the smaller cell-size and slower cell cycling cells were segregated from the proliferative larger cell-size activated T cell pool at the peak of infection. Gene signature of the smaller cell-size slower cycling cells and the large cell-size proliferative cells was compared to the signature of naïve, effector, central and effector memory CD8+ T cells. Total RNA samples were prepared from sorted populations of larger or smaller cell-sized cells from spleens of influenza virus PR8-OVA-infected mice on day 7 p.i. or from in vitro 7 days culture after stimulation with plate-bound anti-CD3ε (1.0 μg ml−1) and anti-CD28 mAb (0.5 μg ml−1). Effector T-cell control samples were prepared from SIINFEKL (100 ng ml−1) stimulated OT-I cells after 4 days of in vitro culture with rIL-2 (10 ng/ml) and sorted as CD8+CD44hiCD62Llo. Control bona fide effector memory and central memory T cells were sorted from the spleens of PR8-OVA-infected mice on day 42 p.i. Naive cells were sorted as CD8+CD44loCD62Lhi cells from uninfected C57BL/6 mice.
Project description:Memory T cells are critical to protect us from recurring infections. Their instantaneous reactivity to pathogens is empowered by persistent expression of cytokine mRNA. How aberrant protein production of this pre-formed mRNA is prevented in the absence of infection, however, remains unresolved. We show that protein production in memory T cells is blocked through a 3’untranslated region (3’UTR)-mediated process, and that AU-rich elements (AREs) are key herein. Germ-line deletion of AREs leads to chronic IFN- production in bona fide memory T cells. Strikingly, the aberrant protein production does not result from increased mRNA levels and/or half-life. Rather, AREs block the recruitment of cytokine mRNA to ribosomes, a process that is mediated by the ARE-binding protein ZFP36L2. Thus, AREs are crucial elements for translational repression that allow murine and human memory T cells to contain pre-formed cytokine mRNAs, while preventing undesirable protein production in the absence of infection.
Project description:An unresolved issue in immunology is the extent to which inflammatory effects are needed for robust T cell responses. In this study, mice were immunized by iv injection using either high toxicity lipopolysaccharide (LPS) or low toxicity monophosphoryl lipid A (MPL) as adjuvant. Six hours after iv immunization, whole spleens were harvested and gene expression was measured in unfractionated splenic populations of cells. The analysis indicated that the low toxicity adjuvanticity of MPL was associated with TLR4-mediated signaling that was biased to the TRIF branch of TLR4, while LPS generated balanced MyD88 and TRIF-associated outcomes. Experiment Overall Design: B6.SJL mice adoptively transferred with T cells from B6.OTI and OTII transgenic TCR transgenic were immunized via intravenous injection with ova peptides alone ("Ova"), or with LPS ("OvaLPS") or MPL as adjuvant ("OvaMPL"). Six hours after immunization, the mice were euthanized by cervical dislocation, the spleens were removed and immediately lysed in guanidium chloride, and kept frozen until being used to extract total cellular RNA. Three mice each were given the indicated treatments, with independent processing and analysis of nine samples total.
Project description:How systemic metabolic alterations during acute infections impact immune-cell function remains poorly understood. Hess and colleagues demonstrate that acetate rapidly increases during infections, which drives acetylation of GAPDH in memory CD8+ T cells and thereby catalyzes the rapid recall response. Comparison of mRNA profile of control vs. 3d acetate exposed memory OT-I T cells
Project description:NaM-CM-/ve, liver- and gut-activated CD8 OT-I T cells show differential migration behaviour. To analyze which genes could be responsible for different migration patterns, naM-CM-/ve, liver-activated and gut-activated CD8 T cells were isolated and compared for their gene expression profile. Gene expression profiles of naM-CM-/ve CD8 OT-I T cells versus liver-activated T cells and gut-activated T cells, respectively. NaM-CM-/ve CD8 OT-I T cells were isolated from lymph nodes and spleens of OT-I mice. CD8 OT-I T cells activated by liver-derived and gut-derived antigen for three days in vivo, positive for CFSE were sorted from livers of TF-OVA mice or from mesenteric lymph nodes of iFABP-OVA mice. After total RNA extraction, reverse transcription, cDNA extraction, the biotinylated cRNA was transcribed, fragmented, and 15 M-BM-5g cRNA hybridized in duplicates for each of the three groups to the GeneChip arrays. Group1: naM-CM-/ve, Group2: liver-activated Group3: gut-activated. Lists of differentially regulated genes were created using High Performance Chip Data Analysis (HPCDA) with Bioretis database (http://www.bioretis-analysis.de). Worldwide data sharing is possible via Bioretis, please ask the authors.
Project description:In vitro differentiated Th17 have a distinct expression profile compared to in vivo differentiated Th17 Several conditions of Th17 cells were tested, with 2-3 replicates per condition.
Project description:We demonstrate that transcription factor IRF4 is induced in a T cell receptor (TCR) affinity-dependent manner and functions as a dose-dependent regulator of the metabolic function of activated T cells. IRF4 regulates the expression of key molecules required for aerobic glycolysis of effector T cells, and is essential for clonal expansion and maintenance of effector function of antigen-specific CD8+ T cells. Examination of gene expression profiles in six types of samples