Sequential and coordinate activation of Foxp3 enhancer elements critical for Treg development and immunological self-tolerance

The transcription factor Foxp3 plays crucial roles for Treg cell development and function. The conserved non-coding sequences (CNSs) included in the Treg-specific super-enhancer at the Foxp3 locus control Foxp3 transcription, but how they developmentally contribute to Treg lineage specification remains obscure. Here we showed that among Foxp3 CNSs, CNS0 and then CNS3 were activated at early stages of thymocyte differentiation prior to Foxp3 promoter activation, with sequential genomic looping among these regions. While deletion of either CNS0 or CNS3 partially impaired thymic, peripheral, and in vitro Treg cell differentiation, deletion of both CNS0 and CNS3 completely abrogated the generation of mature Treg cells in vivo and in vitro. As a result, CNS0/CNS3 double deficient mice spontaneously developed lethal systemic autoimmunity/inflammation. Thus, sequential and coordinate activation of Foxp3 CNS0 and CNS3 initiates and stabilizes Foxp3 gene expression, thereby crucially controlling Treg cell development and consequent establishment of immunological self-tolerance and homeostasis.

转录因子Foxp3在调节性T细胞（Treg细胞）的发育与功能中发挥关键调控作用。位于Foxp3基因座的Treg特异性超级增强子所包含的保守非编码序列（CNSs）可调控Foxp3的转录过程，但此类序列在发育层面如何参与Treg细胞谱系特化，目前仍不明确。本研究显示，在Foxp3的各类CNS中，CNS0与CNS3先后在胸腺细胞分化的早期阶段、Foxp3启动子激活前被激活，且这些调控区域间存在时序性的基因组环化相互作用。单独敲除CNS0或CNS3仅会部分削弱胸腺、外周及体外环境下Treg细胞的分化能力；而同时敲除CNS0与CNS3，则可完全阻断体内外成熟Treg细胞的生成。进一步实验发现，CNS0/CNS3双敲除小鼠会自发出现致死性全身性自身免疫/炎症反应。综上，Foxp3 CNS0与CNS3的时序协同激活可启动并稳定Foxp3基因的表达，进而关键性地调控Treg细胞的发育，以及免疫自身耐受与生理稳态的建立。