Fig. 4: ACC1 deletion interferes with PPARδ/β-catenin activation in intestinal organoids.

a Transcription levels of genes associated with intracellular de novo fatty acid synthesis. CPM values were derived from RNA-seq of organoids from ACC1^∆/∆IEC and ACC1^lox/lox mice at day 4 after 4-OHT treatment. RNA-seq was performed in triplicates from 3 independent experiments. b ¹³C labeled glucose was added to organoids derived from ACC1^∆/∆IEC and ACC1^lox/lox mice directly after 4-OHT treatment. Organoids were harvested at indicated time points and incorporation (δ) of ¹³C into de novo synthesized fatty acids was measured by mass-spec. Data pooled from N = 3 independent experiments. c Accumulation of neutral lipids in organoids. LipidTox green was added 30 min prior to analysis of mean fluorescence intensity (MFI) by flow cytometry. Data is representative for 4 independent experiments. d Western blot analysis of nuclear PPARδ and β-catenin protein was performed at indicated time points in organoids derived from ACC1^lox/lox and ACC1^∆/∆IEC mice. Anti-histone H3 was used as loading control. Representative data is shown from one out of 2 independent experiments with similar results. e ACC1^Δ/ΔIEC and ACC1^lox/lox control mice were injected with tamoxifen and treated daily (i.p.) with PPARδ agonist GW501516 or vehicle (veh) until analysis on day 14. H&E stainings of ileum sections are representative of 2 independent experiments with n = 3–5 mice per group. Scale bar represents 100 µm. f–i Organoids from ACC1^∆/∆IEC mice were cultured in the absence or presence of 4-OHT for 24 h to induce ACC1 deletion. PPARδ agonist GW501516 or vehicle control (DMSO) were added for the whole culture period of 5 days. f Representative pictures of organoids and quantification of crypt domains. More than 30 organoids per group were analyzed. GW = GW501516. Bar = 200 µm. g qPCR analysis of Lgr5 expression. h Western blot analysis of nuclear PPARδ and β-catenin protein. GW = GW501516. Representative data is shown from one out of 2 independent experiments with similar results. i Number of secondary organoids per dissociated crypt-derived primary organoids. Primary organoids from ACC1^∆/∆IEC mice were cultured +/− 4-OHT for 24 h. GW501516 or DMSO were added directly after plating. Primary organoids were subcloned on day 4. Data was pooled (N = 3) g, i or is representative (f) for 3 independent experiments with similar results. Statistical significance was analyzed using Bonferroni-Dunn method (b), unpaired two-tailed Student’s t-test (c, Benjamini–Hochberg procedure (a) or One-way ANOVA with Tukey’s multiple comparison test (f, g, i) with *p < 0.05, **p < 0.01, ***p < 0.001. ****p < 0.0001. Exact p values provided as Source Data. Bar graphs represent mean and error bars indicate SD. Boxes in boxplots denote 25th to 75th percentiles with whiskers representing min-max, and the central line the median. Source data are provided as a Source Data file.