Total and cholera toxin B-tetramer specific T cells from Peyer's patches and mesenteric lymph nodes
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ABSTRACT: T cells were separately isolated from MLN and PP from two naive mice, and T cells binding to a cholera toxin B subunit specific MHC II tetramer from mice orally immunized with cholera toxin 7 days or boosted 5 days with a second dose of cholera toxin. Cells were purified using flow cytometric cell sorting and 3´ (n=5) or 5´ (n=5) libraries were constructed using the 10X workflow. From 5´libraries TCR libraries were also constructed and sequenced.
Project description:CD8+ T cells in chronic viral infections like HIV develop functional defects such as loss of IL-2 secretion and decreased proliferative potential that are collectively termed exhaustion1. Exhausted T cells express increased levels of multiple inhibitory receptors, such as Programmed Death 1 (PD-1). PD-1 inhibition contributes to impaired virus-specific T cell function in chronic infection because antibody-mediated blockade of its ligand, Programmed Death Ligand 1 (PD-L1) is sufficient to improve T cell function and reduce viral replication in animal models. Reversing PD-1 inhibition is therefore an attractive therapeutic target, but the cellular mechanisms by which PD-1 ligation results in T cell inhibition are not fully understood. PD-1 is thought to limit T cell activation by attenuating T cell receptor (TCR) signaling. It is not known whether PD-1 ligation also acts by upregulating genes in exhausted T cells that impair their function. Here, we analyzed gene-expression profiles from HIV-specific CD8+ T cells in patients with HIV and show that PD-1 coordinately upregulates a program of genes in exhausted CD8+ T cells from humans and mice. This program includes upregulation of basic leucine transcription factor, ATF-like (BATF), a transcription factor in the AP-1 family. Enforced expression of BATF was sufficient to impair T cell proliferation and cytokine secretion, while BATF knockdown reduced PD-1 inhibition. Silencing BATF in CD4+ and CD8+ T cells from chronic viremic patients rescued HIV-specific T cell function. Thus inhibitory receptors can cause T cell exhaustion by upregulating genes â such as BATF â that inhibit T cell function. We sorted HIV-specific CD8+ T cells from 18 progressors and 24 controllers, cohorts with a two-log difference in mean viral load
Project description:Mucosal-Associated Invariant T cells (MAIT cells) have a unique specificity for the microbial metabolite 5-OP-RU presented by the non-classical presentation molecule MR1. Upon activation, they release cytotoxic mediators and engage an antimicrobial activity. As a subset of T lymphocytes, MAIT development occurs in the thymus where they acquire their effector phenotype under the control of the key transcription factor ZBTB16. This particular maturation process is in contrast with conventional T cells that egress the thymus with a naive phenotype before populating the secondary lymphoid organs, and the molecular events driving the MAIT lineage decision are poorly known. In the present work, we evaluated the transcriptional events and the role of the slam-SAP pathway on the lineage decision of MR1-restricted T cells by single cell RNAseq. MAIT cells undergoing positive selection were FACS-sorted with a MR1:5-OP-RU labeled tetramer, from thymus of wild-type and sapKO mice. Their transcriptomes were captured using a 10x chromium system.
Project description:This SuperSeries is composed of the following subset Series: GSE24026: Comparison of gene expression profiles of Jurkat cells with or without PD-1 ligation GSE24081: Comparison of gene expression profiles of HIV-specific CD8 T cells from controllers and progressors Refer to individual Series
Project description:Xbp1 is a major transcription factor in the unfolded protein response. To uncover its function in DCs we generated a conditional KO for Xbp1 in dendritic cells. We here compare the expression of mRNAs in two different splenic DC subpopulations, CD8a and CD11b DCs in both WT and KO mice. Reference: Inositol-requiring enzyme 1-alpha regulates CD8a dendritic cell function via regulated mRNA decay. Osorio et al, Nature Immunology (2014) Primary DC subsets were isolated and sorted from spleens from 3 different WT or CD11c-cre Xbp-1fl/fl mice. RNA was isolated, converted to cDNA and then hybridised on Affymetrix GeneChip Mouse Gene 1.0 ST Arrays (GPL6246).
Project description:Single cell proteomics data containing quantitative information of THP1 and U937 monocytes that were treated or not to undergo a macrophage-like differentiation. Dataset also contains "Mix" samples generated by mixing in equal proportions THP1 and U937 peptides in the single-cell range or by processing together 1 THP1 cell and 1 U937 cell. Samples run on the Orbitrap Fusion Lumos Tribrid and the Exploris 240 were acquired by the de Duve institute, UCLouvain. Samples run on the timsTOF SCP were acquired by the GIGA institute, ULiège.
Project description:Transcriptional profiling of Langerhans cells isolated from CD11c-Cre x Dicer wt/wt (Dicer +/+ ) and CD11c-Cre x Dicer fl/fl (Dicer-/-) mice. Langerhans cells are isolated via enzymatic digestion of skin followed by antibody staining and FACS sorting
Project description:Comparison of the germinal center B cell receptor repertoire in Cxcl13-Cre/TdTom EYFP (control) and Cxcl13-Cre/TdTom EYFP Cxcl12fl/fl mice on day following subcutaneous immunization with NP-KLH.
Project description:Human cytomegalovirus infection (CMV) can stimulate robust human leukocyte antigen (HLA)-E restricted CD8 T cell responses. These T cells recognize a peptide from UL40, which differs by as little as a single methyl group from self-peptides that also bind HLA-E, challenging their capacity to avoid self-reactivity. We showed in one donor 2 distinct populations of UL40/HLA-E T cells, with vastly different T cell receptor (TCR) affinities for the UL40/HLA-E complex. However, paradoxically, lower cytokine responses were observed from UL40/HLA-E T cells bearing TCRs with high affinity for HLA-E. To identify why these T cells bearing high affinity T cell receptors were less responsive to antigens, we performed single cell RNAseq analysis. These 2 distinct populations of T cells were single-cell sorted into 96-well plates prior to single cell RNA-seq analysis.
Project description:Semi-invariant natural killer T (NKT) cells are thymus-derived innate lymphocytes that modulate microbial and tumour immunity as well as autoimmune diseases. These immunoregulatory properties of NKT cells are acquired during their development. Much has been learnt regarding the molecular and cellular cues that promote NKT cell development, yet how these cells are maintained in the thymus and the periphery and how they acquire functional competence are incompletely understood. We found that IL-15 induced several Bcl-2 family survival factors in thymic and splenic NKT cells in vitro. Yet, IL15-mediated thymic and peripheral NKT cell survival critically depended on Bcl-xL expression. Additionally, IL-15 regulated thymic developmental stage 2 (ST2) to ST3 lineage progression and terminal NKT cell differentiation. Global gene expression analyses and validation revealed that IL-15 regulated Tbx21 (T-bet) expression in thymic ST3 NKT cells. The loss of IL15-dependent T-bet expression resulted in poor expression of IFN-γ and several NK cell receptors in NKT cells. Taken together, our findings reveal a critical role for IL-15 in NKT cell survival, which is mediated by Bcl-xL, and effector differentiation, which is regulated by T-bet. Gene expression was measured in NKT cells sorted from pooled thymi of wild-type (3 replicates) or IL-15 deficient (2 replicates) mice.
Project description:Normal thymic T cell development is enabled by a stromal microenvironment most importantly composed of distinct epithelial cell populations in cortex and medulla. Their differentiation, growth and function require the expression of the transcription factor Foxn1. Direct targets of Foxn1 have, however, remained largely undefined. Utilizing newly created static and inducible genetic model systems, we now provide a genome wide map of Foxn1 target genes and the sequences bound by this master regulator. Foxn1 controls not only essential steps early in intrathymic lymphoid development including T cell lineage commitment but is also indispensable for later stages in T cell maturation such as the selection of CD4 and CD8 T cells. Thus, Foxn1 function critically choreographs both early and late events in thymic lympho-stromal cross-talk. Foxn1 ChIP-seq and RNA-seq in mouse models of hypofunctional or conditional knock-out of Foxn1 Brief sample descriptions are shown below: Foxn1 ChIP-seq (GSM1945905) - chromatin immunoprecipitated using an antibody against FOXN1-FLAG (wt*); 2 samples Input ChIP-seq (GSM1945906) - input chromatin; 2 samples 42cT (GSM1945907) - RNA-seq on wt*/-::nu/nu cTEC; 3 samples 42mT (GSM1945908) - RNA-seq on wt*/-::nu/nu mTEC; 2 samples 96cT (GSM1945909) - RNA-seq on wt*/wt*::nu/nu cTEC; 3 samples 96mT (GSM1945910) - RNA-seq on wt*/wt*::nu/nu mTEC; 2 samples NcT (GSM1945911) - RNA-seq on Dox-treated TetO- iFoxn1(del7,8) cTEC; 5 samples PcT (GSM1945912) - RNA-seq on Dox-treated TetO+ iFoxn1(del7,8) cTEC; 5 samples C57BL/6 mice (GSM1945913) - ATAC-seq on wild-type cTEC; 1 sample Please see each sample for more detailed information.