Figure 4. ERK5 signalling is required for the proliferation of Erk1/2^ΔIEC organoids.

(a,b) Small intestinal organoids generated from naive ΔIEC mice were treated with EtOH or tamoxifen (0.5 μM) for two consecutive days. Organoids were stained for ERK1/2 with counter staining for F-actin (phalloidin-AF546), on day 5 after initiation of treatment, and analysed by confocal fluorescent imaging (a). Immunoblotting with intestinal organoid lysates for indicated proteins (b). (c–f) Intestinal organoids were treated with EtOH or tamoxifen (0.5 μM) in combination with DMSO or XMD8-92 (10 μM), respectively, and analysed on day 5. Ki67 staining and confocal fluorescent imaging (c). Analysis of RNA samples by Q-PCR (n=2–3 per group) for Mki67 (d), c-Fos (e), Lgr5 and Mmp7 (f), normalized to Gapdh. Data are mean±s.e.m. *P<0.05, **P<0.01 versus control (EtOH) or as indicated, by analysis of variance. Scale bars, 100 μm (a and c). All data are representative of at least two independent experiments.