CD4CD8 Lineage Commitment Is Regulated by a Silencer Element at the ThPOK Transcription-Factor Locus

Fox Chase Cancer Center, 7701 Burholme Ave., Philadelphia, PA 19111, USA.
Immunity (Impact Factor: 21.56). 03/2008; 28(3):346-358. DOI: 10.1016/j.immuni.2008.02.006


The transcription factor ThPOK is necessary and sufficient to trigger adoption of the CD4 lymphocyte fate. Here we investigate the regulation of ThPOK expression and its subsequent control of CD4+ T cell commitment. Treatment of immature thymocytes with anti-TCR (T cell receptor) showed that TCR signals were important in ThPOK induction and that the CD4+8lo stage was the likely target of the inductive TCR signal. We identified at the ThPOK locus a key distal regulatory element (DRE) that mediated its differential expression in class I- versus II-restricted CD4+8lo thymocytes. The DRE was both necessary for suppression of ThPOK expression in class I-restricted thymocytes and sufficient for its induction in class II-restricted thymocytes. Mutagenesis analysis defined an essential 80bp core DRE sequence and its potential regulatory motifs. We propose a silencer-dependent model of lineage choice, whereby inactivation of the DRE silencer by a strong TCR signal leads to CD4 commitment, whereas continued silencer activity leads to CD8 commitment.

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Available from: Xiao He, Oct 27, 2015
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    • "Transgenic reporter expression assays and the analysis of how Runx complexes regulate ThPOK expression revealed the existence of several cis-regulatory elements essential for ThPOK expression and gene targeting approaches confirmed the importance of these cis-elements in the regulation of Thpok expression [64, 68, 69]. The proximal Thpok enhancer (designated PE), located about 3.6 kb downstream of exon 1A of the Thpok gene, directs expression in CD4SP and peripheral CD4+ T cells [68]. Interestingly, even though ThPOK expression was normally induced in PE-deficient thymocytes, ThPOK expression was gradually lost in PE-deficient CD4SP cells and CD4+ T cells, indicating that PE is essential for the up-modulation of ThPOK during CD4 lineage differentiation [64]. "
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    ABSTRACT: The differentiation and function of peripheral helper and cytotoxic T cell lineages is coupled with the expression of CD4 and CD8 coreceptor molecules, respectively. This indicates that the control of coreceptor gene expression is closely linked with the regulation of CD4/CD8 lineage decision of DP thymocytes. Research performed during the last two decades revealed comprehensive mechanistic insight into the developmental stage- and subset/lineage-specific regulation of Cd4, Cd8a and Cd8b1 (Cd8) gene expression. These studies provided important insight into transcriptional control mechanisms during T cell development and into the regulation of cis-regulatory networks in general. Moreover, the identification of transcription factors involved in the regulation of CD4 and CD8 significantly advanced the knowledge of the transcription factor network regulating CD4/CD8 cell-fate choice of DP thymocytes. In this review, we provide an overview of the identification and characterization of CD4/CD8 cis-regulatory elements and present recent progress in our understanding of how these cis-regulatory elements control CD4/CD8 expression during T cell development and in peripheral T cells. In addition, we describe the transcription factors implicated in the regulation of coreceptor gene expression and discuss how these factors are integrated into the transcription factor network that regulates CD4/CD8 cell-fate choice of DP thymocytes.
    Full-text · Article · Jun 2013 · Cellular and Molecular Life Sciences CMLS
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    • "Furthermore, the ratio of H3K4me3 to H3K27me3 depositions at the Thpok gfp:242–401RM allele in CD8 þ T cells of Thpok þ /gfp:242–401RM mice was reversed compared to wild type (Figure 6D). This finding is inconsistent with previous results detecting substantial silencer activity in a mutant silencer fragment lacking Runx sites in a transgenic reporter expression assay (He et al, 2008). Given that silencer activity was enhanced by increasing its copy number, we next tested whether insertion of three copies of the 242–401RM mutant silencer restores Thpok silencing by generating a Thpok gfp:3xRM allele (Figure 6A). "
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    ABSTRACT: CD4+ helper and CD8+ cytotoxic T cells differentiate from common precursors in the thymus after T-cell receptor (TCR)-mediated selection. Commitment to the helper lineage depends on persistent TCR signals and expression of the ThPOK transcription factor, whereas a ThPOK cis-regulatory element, ThPOK silencer, represses Thpok gene expression during commitment to the cytotoxic lineage. Here, we show that silencer-mediated alterations of chromatin structures in cytotoxic-lineage thymocytes establish a repressive state that is epigenetically inherited in peripheral CD8+ T cells even after removal of the silencer. When silencer activity is enhanced in helper-lineage cells, by increasing its copy number, a similar heritable Thpok silencing occurs. Epigenetic locking of the Thpok locus may therefore be an independent event from commitment to the cytotoxic lineage. These findings imply that long-lasting TCR signals are needed to establish stable Thpok expression activity to commit to helper T-cell fate and that full commitment to the helper lineage requires persistent reversal of silencer activity during a particular time window.
    Full-text · Article · Mar 2013 · The EMBO Journal
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    • "Therefore, ThPOK expression can be used as a marker of CD4 + lineage identity. We crossed Tcf12 f/f Tcfe2a f/f Cd4cre + mice with a ThPOK reporter strain (denoted here as TR) that expresses GFP under the control of Thpok regulatory elements (He et al., 2008). Analysis of Tcf12 f/f Tcfe2a f/f Cd4cre + TR + mice demonstrated no significant GFP + population within the CD8 SP TCRβ + or CD8 SP TCRβ − gates above the background observed in cre − TR + CD8 SP TCRβ + controls (Figure 6A). "
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    ABSTRACT: The double-positive (DP) to single-positive (SP) transition during T cell development is initiated by downregulation of the E protein transcription factors HEB and E2A. Here, we have demonstrated that in addition to regulating the onset of this transition, HEB and E2A also play a separate role in CD4(+) lineage choice. Deletion of HEB and E2A in DP thymocytes specifically blocked the development of CD4(+) lineage T cells. Furthermore, deletion of the E protein inhibitors Id2 and Id3 allowed CD4(+) T cell development but blocked CD8(+) lineage development. Analysis of the CD4(+) lineage transcriptional regulators ThPOK and Gata3 placed HEB and E2A upstream of CD4(+) lineage specification. These studies identify an important role for E proteins in the activation of CD4(+) lineage differentiation as thymocytes undergo the DP to SP transition.
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