Fig. 6: Calcineurin inhibition selectively impacts the CD4 audition.

a, UMAP plots of individual thymocytes from thymic slices of postnatal (day 1) wild-type mice cultured with no drug or CsA (200 ng ml⁻¹) or U0126 (2 μg ml⁻¹ or 10 μg ml⁻¹) for 48 or 72 h. DP and SP populations are highlighted. b, UMAP plots for the 48-h time point separated into no drug, 200 ng ml⁻¹ CsA, 2 μg ml⁻¹ or 10 μg ml⁻¹ U0126 conditions as in a. c, Scatter plots showing the log fold change in cell-type proportions relative to no-drug control for DP1, DP2a, DP2b, DP2c, CD4 transitional, CD4 SP and CD8 SP cell clusters for no drug, 200 ng ml⁻¹ CsA, 2 μg ml⁻¹ or 10 μg ml⁻¹ U0126 conditions. Each dot is a separate thymic slice, and data are normalized to the mean of the no-drug condition. Error bars indicate mean ± s.e.m. d,e, Frequency (% of live cells) in CD4 SM, CD4 SP and CD8 SP populations normalized to untreated control at 48 h (d) or 72 h (e) of culture as in a, assessed by manual gating (Extended Data Fig. 7a). Error bars indicate mean ± standard error of the mean. f,g, Representative flow cytometry plots (f) and compiled data (g) showing the expression of CD5 and GATA3 in gated DP thymocytes from thymic slices cultured for 48 h with no drug, 200 ng ml⁻¹ CsA, 2 μg ml⁻¹ or 10 μg ml⁻¹ U0126 as in a. a–c, Data are from one representative experiment out of two. d,e,g, Each dot represents a thymic slice; data are compiled from three independent experiments for CsA (200 ng ml⁻¹), n = 10; one experiment for U0126 low (2 μg ml⁻¹), n = 3; two independent experiments for U0126 high (10 μg ml⁻¹), n = 6; and four independent experiments for no drug, n = 13.