Regulation of positive and negative selection and TCR signaling during thymic T cell development by capicua

  1. Soeun Kim
  2. Guk-Yeol Park
  3. Jong Seok Park
  4. Jiho Park
  5. Hyebeen Hong
  6. Yoontae Lee

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    Evaluation Summary:

    This paper focuses on the transcriptional regulation of the T cell receptor (TCR) signaling cascade and would be of interest to those studying T cell development and differentiation. The authors employ a conditional deletion of the Capicua (Cic) gene, a transcriptional repressor previously shown to be involved in regulating autoimmunity and follicular helper T (Tfh) cell differentiation, and now show that loss of CIC in hematopoietic cells leads to defects in TCR-beta selection as well as in positive and negative selection of developing thymocytes. The overall conclusions are well supported by the findings.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)

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Abstract

Central tolerance is achieved through positive and negative selection of thymocytes mediated by T cell receptor (TCR) signaling strength. Thus, dysregulation of the thymic selection process often leads to autoimmunity. Here, we show that Capicua (CIC), a transcriptional repressor that suppresses autoimmunity, controls the thymic selection process. Loss of CIC prior to T-cell lineage commitment impairs both positive and negative selection of thymocytes. CIC deficiency attenuated TCR signaling in CD4 + CD8 + double-positive (DP) cells, as evidenced by a decrease in CD5 and phospho-ERK levels and calcium flux. We identified Spry4 , Dusp4 , Dusp6, and Spred1 as CIC target genes that could inhibit TCR signaling in DP cells. Furthermore, impaired positive selection and TCR signaling were partially rescued in Cic and Spry4 double mutant mice. Our findings indicate that CIC is a transcription factor required for thymic T cell development and suggests that CIC acts at multiple stages of T cell development and differentiation to prevent autoimmunity.

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  1. Version published to 10.7554/elife.71769 on eLife
  2. eLife

    Evaluation Summary:

    This paper focuses on the transcriptional regulation of the T cell receptor (TCR) signaling cascade and would be of interest to those studying T cell development and differentiation. The authors employ a conditional deletion of the Capicua (Cic) gene, a transcriptional repressor previously shown to be involved in regulating autoimmunity and follicular helper T (Tfh) cell differentiation, and now show that loss of CIC in hematopoietic cells leads to defects in TCR-beta selection as well as in positive and negative selection of developing thymocytes. The overall conclusions are well supported by the findings.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 agreed to share their name with the authors.)

  3. eLife

    Reviewer #1 (Public Review):

    The authors extend their previous work on Capicua (Cic), which focused on peripheral T cell differentiation and function, and now focus on T cell development by making use a Vav-cre Cic-f/f mouse model to conditionally delete Cic in all hematopoietic cells. Their findings show that in the absence of Cic there is an apparent block at the TCR-beta selection checkpoint and during negative and positive selection of CD4+CD8+ (double-positive, DP) thymocytes. Additionally, they show that the TCR repertoire of regulatory T (Treg) cells is altered in Cic deficient mice. Transcriptomic analysis point to the underlying mechanism for these defects in TCR mediated outcomes as being due to a de-repression of several genes responsible for inhibiting ERK/MAPK signaling, as well as other related signaling outcomes. The authors further examine the roles of DUSP4/6 and Spry4 as critical players in Cic-deficient thymocytes. Overall, the work is clearly presented and makes use of several TCR-transgenic mouse models to further delineate the defects in T cell selection.

  4. eLife

    Reviewer #2 (Public Review):

    In this work, Kim et al. investigate the impact of transcriptional repressor CIC deletion on thymocyte development. They use a CIC flox/flox mouse crossed with different Cre transgenic models expressing the Cre enzyme at various stages of T cell development and show that early CIC deletion impacts both DN and DP stages of development. They continue their study by demonstrating the impact of CIC on TCR signaling in DP thymocytes and showed by RNAseq that CIC deletion de-repressed several genes involved in TCR signal regulation. Among them, SPRY4 and DUSP6 are known to regulate calcium flux and Erk signaling, respectively. Interestingly, Spry4/CIC double knock out mice display a partially rescued phenotype.

    The scientific approach of this paper is coherent and while the role of CIC in thymocyte selection has already been partially described by the authors in a previous study, a specific study of the CIC-deletion impact on T cell development as in this report could be useful. The techniques and mouse models used in the study are relevant and the authors answer the question they raised.

  5. eLife

    Reviewer #3 (Public Review):

    Previous studies have shown that T-cell specific capicua (CIC) deficiencies lead to lymphoproliferative autoimmune-like phenotypes. The current manuscript addresses the role of CIC in the regulation of positive and negative selection of the T cell receptor (TCR) repertoire as dysregulation of thymic selection may contribute to this autoimmune pathology. For this, the authors make use of mice in which CIC has been conditionally deleted in hematopoietic progenitor cells, prior to T cell commitment and thymic selection processes.

    While there are no overt differences in the numbers of mature CD4 and CD8 single positive thymocytes in adult Cicf/f;Vav1-Cre mice harboring a polyclonal T cell receptor repertoire, stronger evidence of defects in both positive and negative selection in the absence of CIC are unmasked with the use of TCR transgenic models. Complementary read-outs of TCR signaling and TCR signal strength suggest that immature CD4+CD8+ double positive (DP) thymocytes are refractory to TCR stimulation. This is likely due to the modulation of regulators of TCR signaling that are normally repressed by CIC; indeed, gene expression analysis of DP thymocytes from Cicf/f;Vav1-Cre and control mice identify differential expression of a number of putative CIC target genes that have been described in other cell types and systems and include Spry4, a suppressor of Ras-independent ERK activation. The authors report that over-expression of Spry4 in thymocytes is sufficient to inhibit ERK phosphorylation and suppression of calcium signaling in response to TCR stimulation, and that an Spry4-deficiency on the Cicf/f;Vav1-Cre background partially rescues defects in T cell development in this model.

    As a transcriptional repressor with known targets that act as negative regulators of the RTK/MAPK signaling pathways, major players in the signaling cascade downstream of the T cell receptor, it is perhaps not surprising that CIC may regulate thymic selection, processes that rely on finely tuned TCR signals, and the data presented are largely in line with this hypothesis. Ultimately, whether the extent of the skewing of the T cell repertoire due to differences in thymic selection is responsible for the autoimmune phenotypes in the CIC knock-out mice will need to be determined.

  6. Version published to 10.1101/2021.07.11.451936v1 on bioRxiv