Mus musculus
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA139449 | ENA |
REPOSITORIES: ENA
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Project description:During thymocyte development, double negative (DN) thymocytes go through consecutive rounds of Tcr and Tcr gene recombination that involve the generation of DNA double strand breaks (DSB), addition of N-nucleotides between coding ends, and DNA repair with the final goal of enhancing the diversity of the TCR repertoire. How DN thymocytes can survive such a genomic instability during this complex process and generate functional Tcr and Tcr genes remain unclear. Performing scRNAseq analyses of isolated DN3 and DN4 thymocytes we show here a defined trajectory from DN3 through DN4 stages that is delineated by a G1 cell cycle checkpoint, but also by a G2M cell cycle checkpoint to help DNA repair during recombination of Tcr and Tcr genes prior to cell proliferation. We have previously shown that inactivation of GSK3 by phosphorylation on Ser389 occurs selectively in response to DNA DSB to enhance survival during DNA repair. In this study we show that failure to inactivate GSK3 by Ser389-phosphorylation during the transition from DN3 to DN4 stages compromises Tcr and Tcr repertoire diversity. Inactivation of GSK3 during V(D)J recombination is needed to prevent cell death by necroptosis. Thus, cell cycle checkpoints to ensure proper DNA repair during V(D)J recombination as well as the use of unique survival pathways to avoid necroptosis during this process are critical to warrant a large diversity of pre-selection TCR repertoire.
2025-05-29 | GSE210735 | GEO
Project description:It is established that E2A and its antagonist, Id3, modulate developmental progression at the pre-TCR and TCR checkpoints. Here we show at a global scale how E2A promotes commitment to the T cell lineage and how pre-TCR mediated signalling affects E2A genome-wide occupancy. We find aberrant development of CD4 memory-like and TFH-like cells, T-B cell conjugates and, remarkably, B cell follicles in Id3-/-thymi. We also find that Id3-/-CD4 splenocytes exhibit increased numbers of TFH-like cells. We propose a model in which Id3 modulates the naive versus effector/memory cell fate. Collectively, these data show how E2A acts globally to orchestrate T-lineage development and that Id3 antagonizes E2A activity beyond the pre-TCR checkpoint to enforce the naive T cell fate. ChIP-Seq was performed in thymocytes isolated from either untreated Rag2-/-mice (DN3 cells) or Rag2-/- mice injected with anti-CD3e antibody (DN4 cells). ChIP used antibodies against either E2A or H3K4me1.
2011-07-31 | E-GEOD-30518 | biostudies-arrayexpress
Project description:We performed a single-cell transcriptome analysis of double-negative developing thymocytes from the DN2, DN3 and DN4 populations
2019-02-04 | GSE118292 | GEO
Project description:In order to understand the molecular mechanisms of DN thymocyte development, it may be also of use to clarify how these developmental processes are regulated in terms of their entire gene expression, to which cell differentiation is ultimately ascribed. In the current study, we approached this issue by investigating gene expression profiles in discrete subsets of DN thymocytes under development, in which DN2, DN3, and DN4 thymocytes were sorted and subjected to expression profiling analysis with high-density oligonucleotide microarrays. Experiment Overall Design: The DN2, DN3, and DN4 populations were FACS-sorted from DN thymocytes harvested from four C57BL/6 mice and analyzed by Affymetrix® Mouse Genome 430 2.0 Array® for gene expression. Four independent experiments were performed using 16 mice.
2008-06-15 | E-GEOD-7784 | biostudies-arrayexpress
Project description:T cell development proceeds in a series of developmental stages, which is precisely orchestrated by multiple signaling and molecular networks. Here we found a zinc finger protein Zfp335 intrinsically controls DN to DP transition, as T cell-specific deficiency in Zfp335 leads to a substantial accumulation of DN3 along with reduction of DP, CD4+ and CD8+ thymocytes. This developmental blockade at DN stage results from the impaired intracellular TCRβ expression as well as increased susceptibility to apoptosis in thymocytes. Transcriptomic and ChIP-seq analyses revealed a direct regulation of transcription factors Bcl6 and Rorc by Zfp335. Importantly, enhanced expression of TCRβ and Bcl6/RorγT restores the developmental defect during DN3 to DN4 transition and improves thymocytes survival, respectively. These findings identify a critical role of Zfp335 in controlling T cell development by maintaining intracellular TCR expression-mediated β-selection and independently activating cell survival signaling.
2021-12-30 | GSE184705 | GEO
Project description:T cell development proceeds in a series of developmental stages, which is precisely orchestrated by multiple signaling and molecular networks. Here we found a zinc finger protein Zfp335 intrinsically controls DN to DP transition, as T cell-specific deficiency in Zfp335 leads to a substantial accumulation of DN3 along with reduction of DP, CD4+ and CD8+ thymocytes. This developmental blockade at DN stage results from the impaired intracellular TCRβ expression as well as increased susceptibility to apoptosis in thymocytes. Transcriptomic and ChIP-seq analyses revealed a direct regulation of transcription factors Bcl6 and Rorc by Zfp335. Importantly, enhanced expression of TCRβ and Bcl6/RorγT restores the developmental defect during DN3 to DN4 transition and improves thymocytes survival, respectively. These findings identify a critical role of Zfp335 in controlling T cell development by maintaining intracellular TCR expression-mediated β-selection and independently activating cell survival signaling.
2021-12-30 | GSE184532 | GEO
Project description:Transcriptomic profiling of KN6 DN4 TCR Tg thymocytes adopting the γδ fate (Lig+) or the αβ fate (Lig-)
2014-12-02 | GSE63730 | GEO
Project description:T cell development is accompanied by epigenetic changes that ensure the silencing of stem cell-related, and the activation of lymphocyte-specific programs. How transcription factors influence these changes remains unclear. We show that the Ikaros transcription factor interacts with the Polycomb Repressive Complex 2 (PRC2) in CD4-CD8- thymocytes, and allows its binding to >200 developmentally-regulated genes, many of which are expressed in hematopoietic stem cells. Loss of Ikaros in CD4-CD8- cells leads to diminished histone H3 Lys27 (H3K27) trimethylation and ectopic expression of these genes. Ikaros binding triggers PRC2 recruitment and H3K27 trimethylation. Furthermore, Ikaros interacts with PRC2 independently of the Nucleosome Remodeling and Deacetylation complex. Our results identify Ikaros as a fundamental regulator of PRC2 function in developing T cells. Genome-wide comparison of different histone modifications, Ikaros, Suz12 and NuRD binding in different stages of T cell development in WT and Ikaros mutant mice. Profiling of H3K27me3 in DN1, DN2, DN3, DN4 and DP thymocytes and hematopoietic stem and progenitor cells (LSK cells) of WT and Ikaros mutant mice. Profiling of H3K4me3 and H3ac in WT and Ikaros mutant DP thymocytes. Global analysis of Ikaros binding in WT DN3, DN4 and DP cells, Suz12 binding in WT and Ikaros mutant DN3 cells, and Mta2 and Mi2beta binding in WT DN3 cells. Genome-wide profiling of Ikaros binding and H3K27me3 upon Ikaros activation in Ikaros-deficient leukemic T cells.
2015-11-03 | E-GEOD-61148 | biostudies-arrayexpress
Project description:The regulation of post-transcriptional modifications of pre-mRNA by alternative splicing is important for cellular function, development and immunity The receptor tyrosine phosphatase CD45, which is expressed on all hematopoietic cells, is known for its role in the development and activation of T cells. To investigate the role of hnRNP L further, we have generated conditional hnRNP L knockout mice carrying floxed alleles and the T-cell specific Lck-Cre recombinase transgene. The Lck-Cre transgene is active in DN3, DN4 as well as in DP and SP cells. We found that deletion of hnRNP L results in a decreased thymic cellularity caused by a partial block at the transition stage between DN4 and DP cells. In addition, hnRNP L-/- thymocytes express aberrant levels of the CD45RA splice isoform and show high levels of phosphorylated Lck at the activator tyrosine Y394 but lacking phosphorylation of the inhibitory tyrosine Y505. This is indicative of an increased basal Lck activation and correlated with a higher proliferation rate of DN4 cells in hnRNP L-/- mice. Deletion of hnRNP L also blocked egress of SP cells to peripheral lymphoid organs and the migration of SP thymocytes in response to the chemokines CCL21 and SDF-1a. Since we found that actin polymerization was also compromised in hnRNP L-/- SP cells, we propose that a defect in the signal transduction cascade downstream of the chemokine receptors CCR7 and CXCR4 caused by the absence of hnRNP L is responsible for this phenotype. Our results indicate that hnRNPL regulates pre-T cell development and migration by regulating CD45 pre-mRNA splicing and chemokine receptor signaling. RNA-seq from Thymocytes from WT mice compared to KO (hnRNP L) mice
2012-04-23 | E-GEOD-33306 | biostudies-arrayexpress
Project description:In order to understand the molecular mechanisms of DN thymocyte development, it may be also of use to clarify how these developmental processes are regulated in terms of their entire gene expression, to which cell differentiation is ultimately ascribed. In the current study, we approached this issue by investigating gene expression profiles in discrete subsets of DN thymocytes under development, in which DN2, DN3, and DN4 thymocytes were sorted and subjected to expression profiling analysis with high-density oligonucleotide microarrays. Keywords: developmental stages
2007-05-17 | GSE7784 | GEO