Thpok‐independent repression of Runx3 by Gata3 during CD4⁺T‐cell differentiation in the thymus

Abstract

CD4+ helper T cells are essential for immune responses and differentiate in the thymus from CD4 + CD8 + "double-positive" (DP) thymocytes. The transcription factor [14,15]. However, both are expressed throughout T-cell development, raising the question how they could limit Runx3 expression to MHC-I-restricted thymocytes. Stat5 is activated in thymocytes in response to signaling by IL-7, and is therefore inactive in DP thymocytes that do not express the IL-7 receptor (IL-7R). However, IL-7R is expressed in both MHC-I-and MHC-II-selected thymocytes [16], and it is unclear how Stat5 could activate Runx3 in the former but not the latter. Reciprocally, the transcription factor Thpok, specifically expressed in MHC-II-restricted cells and required for CD4 + T-cell differentiation, represses Runx3 [10,[17][18][19][20]. However, Thpok is not expressed in DP cells and is expressed at low levels in CD4 + CD8 int "transitional" cells, the precursors of CD4 + -lineage thymocytes. Thus, the transcriptional control of Runx3 expression in early CD4 + -lineage precursor cells remains unclear. Runx3 inhibits CD4 + T-cell differentiation by repressingHere we show that a Thpok-independent mechanism represses Runx3 in MHC-II-restricted thymocytes, and we present evidence that it involves the transcription factor Gata3, previously shown to promote CD4 + -lineage differentiation [21][22][23]. These studies identify a novel, repressive function of Gata3 during CD4 + -lineage differentiation in the thymus. Results Thpok-independent Runx3 repression during CD4 + cell differentiation in the thymusTo study the kinetics of Thpok and Runx3 upregulation in the thymus, we set up an experimental system using a GFP- The latter but not the former hypothesis predicted that removing Thpok −/− CD4 SP-like thymocytes from their intrathymic environment would relieve such repression and therefore enhance, rather than reduce, Runx3 expression. Experimental evidence supported this conclusion (Fig. 1C, bottom): whereas a substantial subset of Thpok −/− CD4 SP-like thymocytes expressed little or no Runx3(tRFP) ex vivo, they all did after overnight single-cell suspension culture to disrupt thymocyte-stroma interactions. Accordingly, mutant cells also downregulated the Runx3 target CD4 (Fig. 1C). In contrast, Thpok-sufficient CD4 SP cells remained tRFP − and CD4 hi in these same conditions. These findings supported the idea that intrathymic signals, not mediated by Thpok, restrained Runx3 expression in MHC-II-signaled thymocytes. We therefore decided to explore this possibility. Gata3 represses Runx3The Thpok-deficient cells that expressed Thpok GFP but not Runx3 tRFP expressed CD69 (Fig. 1D, gray

Requirement for Ssbp2 in Hematopoietic Stem Cell Maintenance and Stress Response

“…Accumulating evidence has suggested a myeloid leukemia suppressor role for NOTCH1 (23, 25). Therefore, we queried TCGA AML dataset for SSBP2 and NOTCH1 expression as determined by RNA-Seq studies.…”

“…Further, decreased expression of Notch1 and elevated expression of E2a and its transcriptional target Cdkn1a in Ssbp2 − / − HSPCs identify a role for Ssbp2 in the HSPC-specific gene expression program. Finally, since accumulating evidence suggests a myeloid suppressor role for NOTCH1 (23–25), we evaluated the expression of NOTCH1 and SSBP2 from data obtained from The Cancer Genome Atlas (TCGA). Interestingly, NOTCH1 expression was modestly correlated with SSBP2 expression in primary AML samples.…”

GATA3: a master of many trades in immune regulation

“…Alternatively, GATA3 may also control processes relevant to expression of CD4 and CD8 in a ThPOK independent manner. Evidence exists to support the latter: Enhanced GATA3 expression increased the numbers of TCR hi CD24 lo thymocytes with a CD4SP-like phenotype in a GATA3 transgenic model [64]. GATA3 was also found to repress the expression of Runx3, a factor that suppresses CD4 expression and promotes CD8 expression (reviewed in [65]), independent of ThPOK [64].…”

What Happens in the Thymus Does Not Stay in the Thymus: How T Cells Recycle the CD4+–CD8+ Lineage Commitment Transcriptional Circuitry To Control Their Function

“…Gata3 and Tcf1 bind the Thpok gene, suggesting a direct effect on its transcription (24–26). In addition to their role in CD4 + -lineage specification, E-proteins, Gata3 and Tox are important for positive selection, whether MHC-I- or MHC II-induced (21, 24, 26, 27), and Gata3 represses Runx3 in thymocytes (28). The impact of Tcf1 on the differentiation of DP into SP thymocytes is even broader, as it contributes to Cd4 silencing in CD8 + -lineage cells (24).…”