Quantitative Proteomics Identifies TCF1 as a Negative Regulator of Foxp3 Expression in Conventional T Cells

Regulatory T cells are important regulators of the immune system and have versatile functions for the homeostasis and repair of tissues. They express the forkhead box transcription factor Foxp3 as a lineage-defining protein. Negative regulators of Foxp3 expression are not well understood. Here, we generated double-stranded DNA probes complementary to the Foxp3 promoter sequence and performed a pull-down with nuclear protein in vitro, followed by elution of bound proteins and quantitative mass spectrometry. Of the Foxp3-promoter-binding transcription factors identified with this approach, one was T cell factor 1 (TCF1). Using viral over-expression, we identified TCF1 as a repressor of Foxp3 expression. In TCF1-deficient animals, increased levels of Foxp3(intermediate)CD25(negative) T cells were identified. CRISPR-Cas9 knockout studies in primary human and mouse conventional CD4 T (T-conv) cells revealed that TCF1 protects T-conv cells from inadvertent Foxp3 expression. Our data implicate a role of TCF1 in suppressing Foxp3 expression in activated T cells.