Panel A shows a UMAP plot and stacked bar graphs illustrating regulatory T-cell cluster proportions in wild-type and knockout mice. Panel B shows a heatmap of gene expression across regulatory T-cell clusters. Panel C shows a heatmap comparing regulatory T-cell clusters with previously reported regulatory T-cell states and signatures. Panel D shows stacked bar graphs comparing regulatory T-cell cluster distributions between non-obese diabetic and non-obese diabetic Tnfrsf9-deficient mice. Panel E shows a stacked bar graph showing clonotype distributions across regulatory T-cell clusters in wild-type and knockout mice. Panel F shows heatmaps displaying shared clonotype percentages among regulatory T-cell clusters in wild-type and knockout mice. Panel G shows UMAP pseudotime trajectory plots illustrating differentiation pathways of regulatory T-cell clusters in wild-type and knockout mice. O
scRNA-seq analysis reveals altered differentiation and clonal expansion of islet-infiltrating Foxp3 + Tregs in Cre + -Tnfrsf9 fl/fl mice. (A) UMAP plot of islet-infiltrating Foxp3+ T cells reclustered from CD4-2 (Treg) cells shown in Fig. 3 A. Bar plots show the proportions of the clusters. (B) Heatmap of key cluster marker genes. (C) Gene module scores of the signatures obtained from the previously defined Foxp3+ Treg (Sprouse et al., 2018; Miragaia et al., 2019) and conventional CD4 T cell (Collier et al., 2023) differentiation states. (D) Bar plots show the proportions of islet-infiltrating WT NOD and NOD.Tnfrsf9−/− Foxp3+ Tregs projected onto the clusters defined in A. (E) Clonal expansion of Cre+-Tnfrsf9+/+ (WT) and Cre+-Tnfrsf9fl/fl (KO) cells within each Treg cluster. The percentages of cells in clonotypes with 1, 2–5, 6–10, or >10 cells in the indicated cluster are shown. (F) Tile plots showing the percentage of clonotype overlap across clusters in Cre+-Tnfrsf9+/+ (WT) and Cre+-Tnfrsf9fl/fl (KO) Tregs. The number in each tile indicates the number of unique clonotypes shared by the two clusters. (G) Differentiation trajectory of Cre+-Tnfrsf9+/+ (WT) and Cre+-Tnfrsf9fl/fl (KO) Tregs.