Extended Data Fig. 5: Differentially expressed genes (DEGs) in cDC1s in scRNA-seq analysis and ex vivo TGFβ-dependent Ag-specific FOXP3⁺ T_reg induction by CCR7⁺ cDC1s.

a, UMAP of splenic cDC1 gene expression by sample identity. b, Dot plots of top condition-specific DEGs in Epor^flox/flox and Epor^ΔXcr1 mice (TLI/ATS vs. UNT). c, Absolute cDC1 numbers per spleen in UNT vs. TLI/ATS-treated Epor^flox/flox (n = 5/condition) and Epor^ΔXcr1 (n = 5/condition) mice. d, UMAPs of cDC1 subtypes in Epor-tdT⁺ and Epor-tdT^– cells. e-g, Dot plots of top condition-specific DEGs in Epor-tdT⁺ and Epor-tdT^– cDC1s (TLI/ATS) and in Epor^flox/flox and Epor^ΔXcr1 mice (TLI/ATS vs. UNT). Dot color = expression, size = % of indicated gene expressed cells (b,e-g). h, Bar charts showing cDC subtype (d) proportions in Epor-tdT⁺ and Epor-tdT^– cDC1s following TLI/ATS. i, Role of TGFβ in FOXP3⁺ T_reg induction by CCR7⁺ cDC1s: 12 h after apoptotic Act-mOVA injection, CCR7⁺ cDC1s (1×10⁴) were cocultured with CD45.1⁺ CTV-labeled naïve OT-II cells ± anti-TGFβ; FOXP3 expression was analyzed by flow cytometry (n = 5/group). j,k, Representative flow cytometry analysis and l,m, Absolute cell number of indicated cell populations of Fig. 3j, Itgb8^ΔXcr1 vs. littermate controls. j,l, Day 0 and k,m, Day 14 of UNT (n = 5; n = 5) or TLI/ATS-treated (n = 5; n = 5) groups post allo-BM infusion. n,o, Aldh1a2^ΔCD11c: Batf3^−/− (n = 6) vs. Aldh1a2^flox/flox: Batf3^−/− (n = 7) BM chimeric recipient mice (CD45.1⁺) were given TLI/ATS. 1 day after the last dose of TLI/ATS, 2W1S-BALB/c donor BM cells were infused i.v., and 14 days later, the percentages of donor type (H2K^d+) cells among leukocyte populations in the peripheral blood of hosts were determined (n) and 2W1S-tetramer⁺CD44⁺H-2K^b+TCRβ⁺CD4⁺ T cells from the spleens were analyzed for FOXP3 expression by flow cytometry and FOXP3⁺ T_regs were counted (o). Data are representative of at least three independent experiments with similar results (c,i) or one experiment (j-o). Statistical analysis was performed using unpaired two-tailed Student’s t-test (c,i,n,o), or two-way ANOVA followed by Tukey’s multiple-comparison test with P values adjusted (l,m), or propeller test, two-sided, no multiple-comparison correction (b), or wilcoxon rank sum test, two-sided, Bonferroni correction (h). Data are mean ± s.e.m. (c,i,l,m,n,o). The diagram in i was created in BioRender. Zhang, X. (2025) https://BioRender.com/rq2yp2e.

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