Extended Data Fig. 1: Wtap depletion in T cells induces inflammation of the gut.
a, b, d Deletion efficiency of floxed Wtap alleles in thymocytes and splenic T cells showing an immunoblot (a) with Wtap-specific antibodies using Gapdh as a loading control or flow cytometry (b, d) with Wtap-specific antibodies on the indicated subpopulations, CD4+ T cells from lamina propria of the colon (d). (n = 3 in a, n = 6 in b, d three independent experiments). c, qPCR analysis of the indicated cytokine gene expression within mRNA prepared from colon tissue. Results are to Ywhaz expression and presented relative to wild-type. (n = 3, representative of three independent experiments with technical triplicates). e, f, IFNγ and IL-17a production in CD4+ T cells from the colon. Percentages of IFNγ+IL-17a+ cells are shown in e. (WT, n = 4; KO, n = 9, three independent experiments). g, h Representative images of mice, spleen and lymph nodes from 5-week-old Wtapfl/-; Foxp3-Cre and Wtapfl/fl; Foxp3-Cre mice. Body weight is shown in the bottom (g). Scale bar, 10 mm. (h) (n = 2 biological replicates) i, H&E stained colon tissue from 4−week-old Wtapfl/-; Foxp3-Cre and Wtapfl/fl; Foxp3-Cre mice. Scale bar, 10 µm, magnification x10. (n = 3 biological replicates) j-m, Stainings to detect CD4+ T and Treg cells in spleen (j, k, l) and colon (m) from 5-week-old Foxp3-Cre and Wtapfl/fl; Foxp3-Cre mice. Numbers adjacent to outlined areas indicate the percentage of cells in each gate. (WT, n = 5-7; KO, n = 4, three independent experiments). n, Stainings to detect RORγt and Helios for CD4+YFP+Foxp3+ or YFP–Foxp3+ T cells in mLNs from female Wtapfl/fl; Foxp3-Crehet mice. (n = 3, two independent experiments). o, IFNγ and IL-17a production in Treg cells in lamina propria of the colon. Percentages of IFNγ+ or IL17a+ cells are shown in the right panel. (WT, n = 4; KO, n = 3 biological replicates). All data are presented as mean values ± s.e.m. Statistical analyses were performed using unpaired two-tailed Student’s t-test.
