E蛋白（E2A/HEB）调节Treg细胞功能和稳定性的分子机制及其在炎症疾病中的意义

E proteins (E2A/HEB) are transcription factors that are essential for T cell development in thymus, our previous finding showed that E proteins regulate Treg cell development. However, whether E protein plays a role in maintaining Treg cell function and stability is still unclear. By crossing the E2Af/fHEBf/f mice with Fopx3-GFP-Cre-Rosa26-Stop-YFP mice, we propose to address the effect of E protein on Treg cells in a comprehensive fashion. First, by knocking out E protein specifically in Treg cells after Foxp3 expression, we will investigate if E protein is still has an effect on Treg cell development, then, the role of E protein in the maintaining Treg cell function and stability will be addressed by using different disease model such as EAE, transfer colitis and DSS colitis ect. Finally, the relevance mechanism of E protein that modulate Treg cell function and stability will determined by RNA-Seq and ChIP study. New target of E protein will screened from the RNA-Seq data and will be verified. We believe this project will provide novel insights into an unexplored area of E protein biology that will not only fuel a better understanding of Treg cell development, but will also facilitate the development of novel approaches to therapeutic manipulate Treg cells.

转录因子E蛋白（E2A/HEB）在调节胸腺中T细胞的发育中起着关键性的作用。我们的前期工作表明E蛋白既调控Th17细胞的分化，也参与调节nTreg细胞的发育。但E蛋白对维持Treg细胞的功能和稳定性是否重要以及E蛋白在以上过程中的机制是什么，仍不清楚。本项目通过在Foxp3表达后中敲除E蛋白，首先研究在Treg细胞中敲除E蛋白后是否仍然会影响Treg细胞的分化；利用不同的炎症疾病模型如Transfer colitis, EAE, DSS肠炎等，研究在Treg细胞中敲除E蛋白后对Treg功能及稳定性的影响； 为阐明E蛋白调节Treg细胞功能和稳定性的机制，将利用RNA-seq和染色质免疫沉淀等方法，发现E蛋白调控Treg细胞功能和稳定性的靶点，并对其靶点进行验证。 项目的研究将更深入阐明在炎症环境中Treg细胞发挥功能的机制，对通过操纵Treg细胞的免疫治疗有重要意义。

既往研究表明TCR信号对维持效应性Treg细胞的稳态及效应性Treg细胞发挥正常功能至关重要，然而其具体作用机制还不清楚。在本课题中，我们利用E蛋白条件性敲除小鼠，在表达过Foxp3的细胞中特异性的敲除E蛋白，发现E蛋白条件性敲除小鼠体内的Treg细胞明显增加；缺失E蛋白的Treg细胞具有更好的免疫抑制功能，同时变得更加稳定；此外，缺失E蛋白的Treg细胞会上调表达活化相关分子——CD103, ICOS，IRF4, KLRG-1等，使得更多的Treg细胞分化成为效应性Treg, 更易进入免疫反应高度活跃的部位（如肺脏，肠道等），或者在病理情况下（如实验性脑脊髓炎）进入炎症部位发挥免疫抑制功能。通过转录普分析发现，大部分受E蛋白调控的基因，尤其是效应性Treg细胞相关基因也会受TCR信号调控；我们对E蛋白调控Treg细胞的分子机制研究发现，E蛋白可以直接结合到效应性Treg细胞相关基因的调控区，进而调控这些基因的表达，从而对Treg细胞的活化状态、迁移以及免疫抑制功能产生影响。本课题利用E蛋白条件性敲除小鼠探究E蛋白对Treg细胞的影响，证明E蛋白作为TCR信号和效应性Treg细胞之间的桥梁，介导了TCR信号对效应性Treg细胞分化的调控。