Integrated Methods Redefine Contrasting Mechanisms Governing Foxp3 Induction and Maintenance

Abstract

Nuclear factor Foxp3 sustains immune-suppressive function of regulatory T (Treg) cells during their long lifespan, raising a fundamental question about how stochastic Foxp3 induction is transformed into stable Foxp3 expression during Treg lineage commitment. To address this issue, we examined the processes of Treg development and observed an abrupt switch of the transcriptional mechanisms regulating Foxp3 expression. By integrating genetic and pharmacological approaches with unbiased CRISPR screening, we discovered several known and novel nuclear factors spanning a diverse aspects of gene regulation that control Foxp3 induction through a super-additive effect. After Tet-dependent DNA demethylation, Foxp3 transcription was governed by a robust mechanism involving redundant nuclear programs and a shift of Foxp3 enhancers. These distinct regulatory modes transform stochastic induction to a stable state of Foxp3 transcription. Collectively, our results revealed novel regulators and redefined the contrasting mechanisms governing Foxp3 transcription for Treg selection and lineage maintenance.