Figure 3. DCAF1 deletion leads to T cell aging in young mice.

(A) Protein expression of DCAF1 in Tregs isolated from young and aged mice, assessed by immunoblotting. Left: Representative of 3 independent experiments. Right: Statistical summary, means ± SD, *P < 0.05, by Mann-Whitney U test. (B) SA-β-gal activity in CD4⁺CD25⁺ Tregs and CD4⁺CD25^– Tconv cells in splenocytes from mice of indicated genotypes, analyzed by flow cytometry with the fluorescent β-gal substrate C₁₂FDG (gray area, no C₁₂FDG; n = 6 mice of 3 experiments; representative results are shown; means ± SD, ****P < 0.0001, by 1-way ANOVA followed by Tukey’s multiple-comparisons test). (C) Heatmap analysis of RNA-Seq data sets to compare top regulated genes in young WT, young Dcaf1-deficient (KO), and aged WT Tregs. The depicted distance was calculated based on Pearson’s correlation. (D) Upregulation of the aging program in Dcaf1-deficient (KO) Tregs (left) and Tconv cells (right), revealed by GSEA of RNA-Seq data sets. (E) Comparison of aging signature gene expression in indicated T cells by qRT-PCR analysis of indicated genes (n = 10 mice of 4 experiments; means ± SD, *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, by 1-way ANOVA followed by Tukey’s multiple-comparisons test).