Figure 4. Resting and activated T_reg states identified by scRNAseq.

a. Biclustering heatmap of the expression of the most characteristic genes for each T_reg cluster (down-sampled to equilibrate cluster size; each vertical line represents one cell)

b. Biclustering heatmap of expression signatures in T_reg cells belonging to different clusters (cell and clusters aligned with a).

c. Circular a posteriori projection plot (CAP) ⁴⁰. Each single cell is projected on a circular plan according to its likelihood to belong to any of the 6 clusters defined in a (n = 708). The proportion of cells in each cluster is indicated.

d. Same tSNE plots as in Fig. 2a, highlighting the location of individual T_reg cells from each of the T_reg clusters.

e. Cxcr3, Cd62l and Nr4a1 expression in single T_reg cells determined by flow cytometry (top row, representative of 3 experiments) or by scRNAseq (corrected counts, bottom row). Top right: Nr4a1-GFP expression in CXCR3⁻ and CXCR3⁺ T_regs (as gated),* p < 0.05 (t-test).

f. Proportion of cells in each T_reg cluster (defined in a) that express different effector transcripts. Proportion calculated after correction for dropout events (see Methods). *** p < 10⁻⁴ (ANOVA).