Figure S5. Loss of Mll1 abolishes proliferation and promotes Paneth/goblet cell differentiation in intestinal organoids and crypts with concomitant activation of Wnt and Mapk signalling.

(A) Immunofluorescence staining for Ki67 (red) on sections of non-induced control and tamoxifen-induced β-cat^GOF; Mek1^GOF; Mll1^+/− and β-cat^GOF; Mek1^GOF; Mll1^−/− organoids. E-cadherin (yellow) stains cell borders, nuclei in blue (DAPI), scale bars 20 μm. Right: Quantification of Ki67⁺ cells per organoid in control, β-cat^GOF; Mek1^GOF; Mll1^+/− and β-cat^GOF; Mek1^GOF; Mll1^−/− organoids, n = 4 independent experiments, Mann–Whitney U test, ****P < 0.0001. Box plot indicates median (middle line) and 25^th, 75^th percentile (box) with Tukey whiskers. (B) H&E staining on sections of small intestines of β-cat^GOF; Mek1^GOF; Mll1^+/− and β-cat^GOF; Mek1^GOF; Mll1^−/− at 20 d after induction with tamoxifen and wild-type control mice, scale bars 100 μm. Stainings were performed in n = 3 independent wild-type and n = 5 independent mutant mice per genotype. (C) Quantification of villus length in n = 5 independent β-cat^GOF; Mek1^GOF; Mll1^+/− and β-cat^GOF; Mek1^GOF; Mll1^−/− mice each at 20 d after induction relative to n = 3 independent control mice, at least 10 villi per mouse measured, Mann–Whitney U test, ****P < 0.0001. Box plot indicates median (middle line) and 25^th, 75^th percentile (box) with Tukey whiskers. (D) mRNA expression of secretory lineage markers Spdef, Mmp7, Itf, Klf4, Muc2, and Gob5 in intestines of control (grey, n = 3 independent mice), β-cat^GOF (green, n = 3 independent mice), β-cat^GOF; Mek1^GOF; Mll1^+/− (blue, n = 5 independent mice), and β-cat^GOF; Mek1^GOF; Mll1^−/− mice (light blue, n = 5 independent mice) at 20 d after the induction of mutagenesis, two-tailed unpaired t test, Spdef: **P = 0.009 (control/β-cat^GOF), **P = 0.0046 (β-cat^GOF/Mll1^+/−), Mmp7: **P = 0.0012 (control/β-cat^GOF), **P = 0.0026 (β-cat^GOF/Mll1^+/−), Itf: **P = 0.0014 (β-cat^GOF/Mll1^+/−), **P = 0.008 (Mll1^+/−/Mll1^−/−), Klf4: **P = 0.009, Muc2: **P = 0.002 (β-cat^GOF/Mll1^+/−), **P = 0.007 (Mll1^+/−/Mll1^−/−), Gob5: *P = 0.04, **P = 0.003, ***P = 0.00015. Data are presented as mean values ± SEM.