Extended Data Figure 7: Stimulation of Yap-independent growth via stromally derived Ereg expression.
From: Yap-dependent reprogramming of Lgr5+ stem cells drives intestinal regeneration and cancer
a, Yap+/Δ and YapΔ/Δ organoids were grown for 3 days in standard growth media and supplemented with 0.5 μg ml−1 of recombinant Ereg. Panels in the top row show Edu incorporation and active caspase 3 stainings in organoid cultures. Proliferation and apoptosis status in Ereg-stimulated YapΔ/Δ organoids are comparable to control organoids. Arrowhead points to apoptotic cells in Yap-deficient organoids. Panels in the bottom row show endogenous Yap expression and confirm that Ereg-stimulated YapΔ/Δ organoids are Yap deficient. Green signal in YapΔ/Δ organoids is non-specific staining of cellular debris in lumen. b, ISH to monitor Ereg expression in untreated and irradiated (2 dpi, 12 Gy) Yap+/Δ and YapΔ/Δ mice. In iii and vi, dotted lines demarcate crypt boundaries. c, Ereg was detected by fluorescence ISH (red) and epithelia highlighted by counterstaining for β-catenin protein (green). Open arrowheads point to examples of Ereg expression in the epithelium, and filled arrowheads indicate stromal cells. Note Ereg expression in certain cells of the regenerating epithelium. All images are representative of three stainings performed on tissues derived from separate mice. Scale bars (a–c), 70 μm. d, qPCR analysis of stromally derived factors Ereg, Sfrp1, Bmp4, Wnt2b, Wnt4, Wnt5a from samples of whole intestines of irradiated (2 dpi; 10 Gy) Yap+/Δ and YapΔ/Δ mice. Error bars indicate s.e.m.; n = 7 (n represents number of independent mice analysed per genotype).
