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

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.