van Hamburg Jan Piet, de Bruijn Marjolein J W, Ribeiro de Almeida Claudia, Dingjan Gemma M, Hendriks Rudi W
Department of Immunology, Erasmus MC, Rotterdam, The Netherlands.
Mol Immunol. 2009 Oct;46(16):3251-60. doi: 10.1016/j.molimm.2009.08.004. Epub 2009 Sep 2.
The zinc-finger transcription factors Gata3 and ThPOK have both been implicated in positive selection of double positive (DP) thymocytes towards the CD4 lineage. As in the absence of Gata3, expression of ThPOK is lacking, Gata3 may directly regulate ThPOK expression. As ThPOK failed to promote CD4(+) lineage differentiation of Gata3-deficient cells, ThPOK cannot be the only Gata3 target gene essential for the induction of the CD4(+) lineage program. Therefore, it is conceivable that Gata3 is essential for selected DP T cells to reach the developmental stage at which ThPOK expression is induced. Here, we show that Gata3 overexpression does not affect ThPOK expression levels in DP or CD4(+) thymocytes, providing evidence that Gata3 does not directly regulate ThPOK. To identify additional target genes that clarify Gata3 function at the DP thymocyte stage, we performed gene expression profiling assays in wild-type mice and transgenice mice with enforced expression of Gata3, in the presence or absence of the MHC class II-restricted DO11.10 TCR. We found that Gata3 expression in DP cells undergoing positive selection was associated with downregulation of the V(D)J-recombination machinery genes Rag1, Rag2 and TdT. Moreover, Gata3 overexpression was associated with downregulation of many signaling molecules and the induction of modulators of TCR signaling, including Ctla-4 and thrombospondin 2. Together with our previous finding that Gata3 reduces expression of CD5, a negative regulator of TCR signaling, and upregulates TCR expression, these findings indicate that Gata3 in DP cells mainly functions to (i) terminate TCRalpha gene rearrangement, and (ii) regulate TCR signal intensity or duration in cells undergoing positive selection towards the CD4 lineage.
锌指转录因子Gata3和ThPOK均与双阳性（DP）胸腺细胞向CD4谱系的阳性选择有关。由于在缺乏Gata3时，ThPOK的表达缺失，因此Gata3可能直接调节ThPOK的表达。由于ThPOK无法促进Gata3缺陷细胞的CD4(+)谱系分化，所以ThPOK不可能是诱导CD4(+)谱系程序所必需的唯一Gata3靶基因。因此，可以推测Gata3对于选定的DP T细胞达到诱导ThPOK表达的发育阶段至关重要。在此，我们表明Gata3的过表达不会影响DP或CD4(+)胸腺细胞中ThPOK的表达水平，这表明Gata3不会直接调节ThPOK。为了鉴定在DP胸腺细胞阶段阐明Gata3功能的其他靶基因，我们在野生型小鼠和强制表达Gata3的转基因小鼠中，在存在或不存在MHC II类限制性DO11.10 TCR的情况下，进行了基因表达谱分析。我们发现，在经历阳性选择的DP细胞中，Gata3的表达与V(D)J重组机制基因Rag1、Rag2和TdT的下调有关。此外，Gata3的过表达与许多信号分子的下调以及TCR信号调节剂的诱导有关，包括Ctla-4和血小板反应蛋白2。连同我们之前的发现，即Gata3降低TCR信号的负调节因子CD5的表达，并上调TCR的表达，这些发现表明DP细胞中的Gata3主要起到以下作用：（i）终止TCRα基因重排，以及（ii）调节向CD4谱系进行阳性选择的细胞中的TCR信号强度或持续时间。
