Fig. 2: Foxp3 degradation induces minimal gene expression and functional changes in mature T_reg cells.

a, Experimental design of scRNA-seq and functional assays. Each genotype and time point consisted of four independent biological replicates. b, UMAP visualization of scRNA-seq data from Foxp3^AIDR26^WT and Foxp3^AIDR26^TIR1(F74G) T_reg cells before and 3 or 7 days after 5-ph-IAA-induced Foxp3 degradation. c, UMAP visualization of the same scRNA-seq data, colored by identified clusters. d, Fraction of each cluster within the total pool of Foxp3^AIDR26^WT or Foxp3^AIDR26^TIR1(F74G) T_reg cells separated by time point. Each point represents a unique mouse. e, In vitro suppression assay of T_reg cells sorted from Foxp3^AIDR26^WT and Foxp3^AIDR26^TIR1(F74G) mice after 7 days of in vivo 5-ph-IAA treatment. 5-ph-IAA was included in culture to sustain Foxp3 degradation. Line graph represents mean ± s.e.m. Data are pooled from two independent experiments and analyzed using two-sided multiple t-tests. NS, not significant. f, Experimental design of the bulk RNA-seq analysis. Each genotype and time point consisted of three independent biological replicates. g, Gating strategy for sorting resting and activated T_reg cells. h, Schematic comparison of the Foxp3^GFPKO reporter-null allele and the functional Foxp3^GFP allele. i, Scatter-plots and bar graphs showing the number of DEGs in resting or activated T_reg cells caused by Foxp3 protein degradation or genetic Foxp3 deficiency.

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