T cell self-reactivity during thymic development dictates the timing of positive selection
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE164896 | GEO | 2021/01/16
REPOSITORIES: GEO
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Project description:Background. T cells in the thymus undergo opposing positive and negative selection processes so that the only T cells entering circulation are those bearing a T cell receptor (TCR) with a low affinity for self. The mechanism differentiating negative from positive selection is poorly understood, despite the fact that inherited defects in negative selection underlie organ-specific autoimmune disease in AIRE-deficient people and the non obese diabetic (NOD) mouse strain. Results. Here we use homogeneous populations of T cells undergoing either positive or negative selection in vivo together with genome-wide transcription profiling on microarrays to identify the gene expression differences underlying negative selection to an Aire-dependent organ-specific antigen, including the upregulation of a genomic cluster in the cytogenetic band 2F. Analysis of defective negative selection in the autoimmune-prone NOD strain demonstrates a global impairment in the induction of the negative selection response gene set, but little difference in positive selection response genes. Combining expression differences with genetic linkage data we identify differentially expressed candidate genes including Bim, Bnip3, Smox, Pdrg1, Id1, Pdcd1, Ly6c, Pdia3, Trim30 and Trim12. Conclusions. The data provide a molecular map of the negative selection response in vivo, and by analysis of deviations from this pathway in the autoimmune susceptible NOD strain, suggest that susceptibility arises from small expression differences in genes acting at multiple points in the pathway between the TCR and cell death. Experiment Overall Design: Analysis of two thymic populations - early single positive thymocytes (CD4+CD8lowCD69+TCRhi) and early double positive thymocytes (CD4+CD8+CD69-TCRlow). Samples purified from the 3A9 TCR transgenic (TCR) to characterise positive selection and the 3A9 TCR x insHEL double transgenic (Dbl) to characterise negative selection towards an Aire-dependent organ-specific antigen. Both the non autoimmune C57BL10.H2k (B10) and autoimmune Non obese diabetic (NOD) strains were analysed in this method. Three biological replicates were used for each of these eight different groups.
2008-06-12 | E-GEOD-3997 | biostudies-arrayexpress
Project description:Abstract of publicaton: CD4/CD8 double-positive (DP) thymocytes express the transcriptional repressor Histone Deacetylase 7 (HDAC7), a class IIa HDAC that is exported from the cell nucleus after T cell receptor (TCR) engagement. Through signal-dependent nuclear export, class IIa HDACs such as HDAC7 mediate signal-dependent changes in gene expression that are important to developmental fate decisions in multiple tissues. We report that HDAC7 is exported from the cell nucleus during positive selection in thymocytes, and regulates genes mediating the coupling between TCR engagement and downstream events that determine cell survival. Thymocytes lacking HDAC7 are inefficiently positively selected due to a severely shortened lifespan and exhibit a truncated repertoire of TCR Jalpha segments. The expression of multiple important mediators and modulators of the response to TCR engagement is altered in HDAC7-deficient thymocytes, resulting in increased tonic MAP kinase activity that contributes to the observed loss of viability. Remarkably, the activity of Protein Kinase D, the kinase that mediates nuclear export of HDAC7 in response to TCR signaling, is also increased in HDAC7-deficient thymocytes, suggesting that HDAC7 nuclear export governs a self-sustaining auto-excitatory loop. These experiments add to the understanding of the life/death decision in thymic T cell development, define a novel function for class IIa HDACs, and point to a novel feed-forward mechanism whereby these molecules regulate their own state and mediate stable developmental transitions. Title of manuscript: Nuclear Export of Histone Deacetylase 7 During Thymic Selection Mediates Immune Self-tolerance. abstract of manuscript: Histone Deacetylase 7 (HDAC7) is a TCR signal-dependent regulator of differentiation that is highly expressed in CD4/CD8 double-positive (DP) thymocytes. Here we examine the effect of blocking TCR-dependent nuclear export of HDAC7 during thymic selection, through expression of a signal-resistant mutant of HDAC7 (HDAC7-?P) in thymocytes. We find that HDAC7-?P Transgenic thymocytes exhibit a profound block in negative thymic selection, but can still undergo positive selection, resulting in the escape of autoreactive T cells into the periphery. Gene expression profiling reveals a comprehensive suppression of the negative selection-associated gene expression program in DP thymocytes, associated with a defect in the activation of MAP kinase pathways by TCR signals. The consequence of this block in vivo is a lethal autoimmune syndrome involving the exocrine pancreas and other abdominal organs. These experiments establish a novel molecular model of autoimmunity and cast new light on the relationship between thymic selection and immune self-tolerance. Goal of Microarray experiment: We did these experiments to determine how alteration of the function of HDAC7, a site-specific and signal-dependent repressor of transcription, changes gene expression in CD4/CD8 DP thymocytes.
2011-04-06 | GSE26488 | GEO
Project description:Developmental checkpoints in stem/progenitor cells are critical to the determination, commitment and differentiation into distinct lineages. Cancer cells often retain expression of lineage-specific checkpoint proteins, but their potential impact in cancer remains elusive. T lymphocytes mature in the thymus following a highly orchestrated developmental process that entails the successive rearrangements and expression of T-cell receptor (TCR) genes. Low affinity recognition of self-peptide/MHC complexes (self-pMHC) presented by thymic epithelial cells by the TCR of CD4+CD8+ (DP) cortical thymocytes transduces positive selection signals that ultimately shape the developing T cell repertoire. DP thymocytes not receiving these signals die by lack of stimulation whereas those that recognize self-pMHC with high affinity undergo TCR-mediated apoptosis and negative selection. In T-cell acute lymphoblastic leukaemia (T-ALL), leukaemic transformation of maturating thymocytes results from the acquisition of multiple genetic and epigenetic alterations in oncogenes and tumour suppressor genes, that disrupt the normal regulatory circuits and drive clonal expansion of differentiation-arrested lymphoblasts. We show here that TCR triggering by negatively-selecting self-pMHC prevented T-ALL development and leukaemia maintenance in mice. Induction of TCR signalling by high affinity self-pMHC or treatment with monoclonal antibodies to the CD3 signalling chain (anti-CD3) caused massive leukaemic cell death and a gene expression program resembling that of thymocyte negative selection. Importantly, anti-CD3 treatment hampered leukaemogenesis in mice transplanted with either mouse or patient-derived T-ALLs. These data provide a rationale for targeted therapy based on anti-CD3 treatment of T-ALL patients and demonstrate that endogenous developmental checkpoint proteins are amenable to therapeutic intervention in cancer cells. Gene expression data from four culture conditions was performed for the following cells: ALL-SIL-TCRα/β-GFP co-cultured on OP9-DL1 for 48 h, ALL-SIL-TCRα/β-GFP without co-culture, ALL-SIL cells transduced with TLX shRNA (sh-TLX), and ALL-SIL transduced with sh-control vectors. All conditions are performed in two replicates.
2016-08-20 | E-GEOD-65496 | biostudies-arrayexpress
Project description:Developmental checkpoints in stem/progenitor cells are critical to the determination, commitment and differentiation into distinct lineages. Cancer cells often retain expression of lineage-specific checkpoint proteins, but their potential impact in cancer remains elusive. T lymphocytes mature in the thymus following a highly orchestrated developmental process that entails the successive rearrangements and expression of T-cell receptor (TCR) genes. Low affinity recognition of self-peptide/MHC complexes (self-pMHC) presented by thymic epithelial cells by the TCR of CD4+CD8+ (DP) cortical thymocytes transduces positive selection signals that ultimately shape the developing T cell repertoire. DP thymocytes not receiving these signals die by lack of stimulation whereas those that recognize self-pMHC with high affinity undergo TCR-mediated apoptosis and negative selection. In T-cell acute lymphoblastic leukaemia (T-ALL), leukaemic transformation of maturating thymocytes results from the acquisition of multiple genetic and epigenetic alterations in oncogenes and tumour suppressor genes, that disrupt the normal regulatory circuits and drive clonal expansion of differentiation-arrested lymphoblasts. We show here that TCR triggering by negatively-selecting self-pMHC prevented T-ALL development and leukaemia maintenance in mice. Induction of TCR signalling by high affinity self-pMHC or treatment with monoclonal antibodies to the CD3 signalling chain (anti-CD3) caused massive leukaemic cell death and a gene expression program resembling that of thymocyte negative selection. Importantly, anti-CD3 treatment hampered leukaemogenesis in mice transplanted with either mouse or patient-derived T-ALLs. These data provide a rationale for targeted therapy based on anti-CD3 treatment of T-ALL patients and demonstrate that endogenous developmental checkpoint proteins are amenable to therapeutic intervention in cancer cells.
2016-08-20 | GSE65496 | GEO
Project description:The T cell receptor (TCR) determines the specificity and affinity for both foreign and self-peptides presented by MHC. It is established that self-pMHC reactivity impacts T cell function, but it has been challenging to identify TCR sequence features that predict T cell fate. To discern patterns distinguishing TCRs from naïve CD4+ T cells with low versus high self-pMHC reactivity, we used data from 42 mice to train a machine learning (ML) algorithm that predicts self-reactivity directly from TCRβ sequences. This approach revealed that n-nucleotide additions and acidic amino acids weaken selfreactivity. We tested our ML predictions of TCRβ sequence self-reactivity using retrogenic mice. Extrapolating our analyses to independent datasets, we found high predicted self-reactivity for regulatory CD4+ T cells and low predicted self-reactivity for T cells responding to chronic infection. Our analyses suggest a potential trade-off between repertoire diversity and self-reactivity intrinsic to the architecture of a TCR repertoire.
2023-01-09 | GSE221703 | GEO
Project description:Title of Publication: Histone Deacetylase 7 Regulates Cell Survival and TCR Signaling in CD4/CD8 Double-Positive Thymocytes Abstract of publicaton: CD4/CD8 double-positive (DP) thymocytes express the transcriptional repressor Histone Deacetylase 7 (HDAC7), a class IIa HDAC that is exported from the cell nucleus after T cell receptor (TCR) engagement. Through signal-dependent nuclear export, class IIa HDACs such as HDAC7 mediate signal-dependent changes in gene expression that are important to developmental fate decisions in multiple tissues. We report that HDAC7 is exported from the cell nucleus during positive selection in thymocytes, and regulates genes mediating the coupling between TCR engagement and downstream events that determine cell survival. Thymocytes lacking HDAC7 are inefficiently positively selected due to a severely shortened lifespan and exhibit a truncated repertoire of TCR J segments. The expression of multiple important mediators and modulators of the response to TCR engagement is altered in HDAC7-deficient thymocytes, resulting in increased tonic MAP kinase activity that contributes to the observed loss of viability. Remarkably, the activity of Protein Kinase D, the kinase that mediates nuclear export of HDAC7 in response to TCR signaling, is also increased in HDAC7-deficient thymocytes, suggesting that HDAC7 nuclear export governs a self-sustaining auto-excitatory loop. These experiments add to the understanding of the life/death decision in thymic T cell development, define a novel function for class IIa HDACs, and point to a novel feed-forward mechanism whereby these molecules regulate their own state and mediate stable developmental transitions. Goal of Microarray experiment: We did these experiments to determine how alteration of the function of HDAC7, a site-specific and signal-dependent repressor of transcription, changes gene expression in CD4/CD8 DP thymocytes. Three biological replicate samples were prepared for each non-wild type mouse strain (Samples 7-18). Six biological replicates were prepared for the wild type strain (Samples 1-6). Total RNA was prepared from isolated CD4/CD8 Double-positive thymocytes and labeled with the Affymetrix Whole-Transcript labeling protocol. Labeled probes were hybridized to one Affymetrix Mouse Gene 1.0ST array each. Data from .cel files were normalized using RMA, in normalization groups representing each of the binary comparisons made. These binary comparisons between sample groups represent gene expression changes due to loss of HDAC7 (samples 1-3 vs. 7-9), transgenic expression of an HDAC7-VP16 fusion protein (samples 4-6 vs. 10-12), positive thymic selection (samples 1-6 vs. samples 11-15), and negative thymic selection (Samples 11-15 vs. samples 16-18).
2011-04-06 | E-GEOD-26488 | biostudies-arrayexpress
Project description:Themis1, a recently described T-lineage specific protein, is essential for thymic positive and negative selection. Although Themis1 has been clearly identified as a component of the T cell antigen receptor (TCR) signalosome, its precise role in TCR signaling remains unclear. Here, we used quantitative proteomic and TCR signaling reporter mice to gain insight into Themis1 signaling function. Mass spectrometry analysis of the Themis1 interactome identified Grb2, SHP1 and Vav1 as the principal interacting partners of Themis1 in thymocytes. The dataset contains mass spectrometry results from the analysis of 6 different kind of AP-MS purifications (based on immunoprecipitation using a Themis1 antibody) starting from the following samples: - thymocytes from WT mice, non stimulated (noted WT NS) - thymocytes from WT mice, stimulated with pervanadate (noted WT P) - thymocytes from GRB2 +/- mice (with decreased expression of GRB2), non stimulated (noted GRB2 NS) - thymocytes from GRB2 +/- mice (with decreased expression of GRB2), stimulated with pervanadate (noted GRB2 P) - thymocytes from Themis1 -/- mice (knock-out for Themis1), non stimulated (noted KO NS) - thymocytes from Themis1 -/- mice (knock-out for Themis1), stimulated with pervanadate (noted KO P) Three biological replicates were prepared for these 6 different conditions (noted, 1,2,3), yielding 18 analyzed samples. Three technical nanoLC-MS runs were acquired for each sample (noted R1, R2, R3), leading to the 54 nanoLC-MS raw files contained in the dataset.
2017-02-06 | PXD004072 | Pride
Project description:Dr. Jameson's research focus is on development and regulation of lymphocytes, especially T cells. Recent work has suggested that differential glycosylation effects the sensitivity of the T cell receptors and its coreceptors, suggesting that regulation of glycosylation may be a critical element in controlling T cell development, survival and functional activity. Determination of how glycosylation enzymes/substrates change in gene expression during development of mouse CD8 T cells. Highly purified pre-selection CD4+8+ thymocytes (using transgenic/knockout mice in our colony) are compared to mature CD8 T cells from the lymph node. The goal is to build on data suggesting that this developmental step involves regulated expression of sialyltransferases (and/or neuraminidases). Highly purified pre-selection CD4+8+ thymocytes (using transgenic/knockout mice in our colony) are compared to mature CD8 T cells from the lymph node. Pre-selection CD4+8+ (DP) thymocytes were sorted from TCRa-/- thymi. Post-selection (post-positive selection) DP thymocytes came from an OT-I TCR transgenic mouse. Naïve (CD44lo) CD8 T cells from lymph node of OT-I mice were used as naïve CD8 T cells. For activated cells, naïve OT-I T cells were activated for 48 hours in vitro with cognate antigen (SIINFEKL peptide/Kb) (displayed on cell sized latex beads) in the presence of IL-2 and IL-12.
2010-12-11 | E-GEOD-26012 | biostudies-arrayexpress
Project description:The TAL1/SCL and LMO1 oncogenic transcription factors establish a pre-leukemic state by reprogramming thymocytes into self-renewing pre-leukemic stem cells (pre-LSCs). Pre-TCR signaling accelerates the progression to T-cell acute lymphoblastic leukemia (T-ALL). To directly address the importance of pre-TCR signaling in driving progression to T-ALL, we leverage on Cd3-deficient mice in which pre-TCR signaling and progression through β-selection is abrogated. In the absence of pre-TCR signaling in Cd3ε-deficient SCL-LMO1 transgenic mice, T-ALL onset is delayed by 150 days. Despite the absence of pre-TCR/CD3 signaling in these mice, we show that leukemic thymocytes exhibit the gene expression profiles of thymocytes that have undergone β-selection, i.e. exhibiting a re-activation of pre-TCR-driven proliferation signature, and a down regulation of HEB/TCF12 target genes. Lastly, monoallelic deletion of Heb is sufficient to accelerate T-ALL onset in Cd3ε-deficient SCL-LMO1 transgenic mice. Together, these results underscore the role of HEB/TCF12 as a tumor suppressor in T-ALL.
2022-03-27 | GSE198506 | GEO
Project description:Sustained Ca2+ entry into CD4+CD8+ double-positive thymocytes is required for positive selection. We identified a voltage-gated Na+ channel (VGSC), essential for positive selection of CD4+ T cells. Pharmacological inhibition of VGSC activity inhibited sustained Ca2+ influx induced by positive-selecting ligands and in vitro positive selection of CD4+ but not CD8+ T cells. In vivo shRNA knockdown of Scn5a specifically inhibited positive selection of CD4+ T cells. Ectopic expression of VGSC in peripheral AND CD4+ T cells bestowed the ability to respond to a positively selecting ligand, directly demonstrating VGSC expression was responsible for increased sensitivity. Thus active VGSCs in thymocytes provides a mechanism by which a weak positive selecting signal can induce sustained Ca2+ signals required for CD4+ T cell development.
2012-07-01 | GSE38909 | GEO