Fig. 4: Yap/Yki signaling is reduced in Pex2^−/− and Mex > Pex5-i intestines and affects intestinal epithelial structure and function.

A The heatmap represents the ratio of log2 fold change expression of genes encoding for proteins of the Hippo pathway in Mex > Pex5-i versus control flies as found in the RNA-seq screen on Drosophila intestines dissected from flies raised on regular cornmeal or high-fat diet (HFD). B The bar graph reports the amount of Yap transcript in WT and Pex2^−/− small intestines. n = 6 mice per each genotype. C Imaging of Yap in intestinal cryosections. The bar graph represents the MFI of Yap staining per ROI. The values reported were calculated on 10 images per experiment, n = 3 mice. Scalebar = 10 µm. D Western blot analyses of total protein extracts from MIO-derived monolayers to quantify p-YAP protein. The bar graph represents ratiometric analyses of the mean intensity value between p-Yap and α-Tubulin in western blots experiments. n = 3 mice. E IF image of p-Yap in small intestine cryosections. The bar graphs represent the MFI of p-Yap staining per ROI and the ratiometric analyses of the MFI between the p-Yap fluorescent signal and the DAPI fluorescent signal, respectively. The values reported were calculated on 10 images per experiment, n = 3 mice. Scalebar = 10 µm. F Imaging of p-Yap in MIO-derived monolayers. The bar graphs show the MFI of total p-Yap staining per ROI and the ratiometric analyses of the MFI between the p-Yap fluorescent signal and the DAPI fluorescent signal, respectively. The values reported were calculated on 10 images per experiment, n = 3 mice. Scalebar = 10 µm. G Western blot analyses of total protein extracts from MIO-derived monolayers to quantify p-MST1/2. The bar graph represents ratiometric analyses of the mean intensity value between p-Mst1/2 and a-Tubulin in western blots experiments. n = 3 extracts from MIO-derived monolayers from three mice. H Western blotting analyses of total protein extracts from Drosophila intestines of the reported genotypes. The bar graph represents ratiometric analyses of the mean intensity value between p-Hpo and α−Tubulin in western blots experiments. n = 25 guts. I IF image of Armadillo protein to detect progenitor cells (bright cells) and enterocyte boundaries (dim cells) in adult Drosophila guts; n = 12 guts. The dotted lines in the lower panels are enterocyte boundary traces of the above panels to illustrate enterocyte size. The bar graphs show the average cell area of enterocytes in μm²; n = 20 intestines per genotype Scalebar = 10 µm. J Representative picture of 20-days-old female flies of the reported genotypes fed with blue-colored food. The dotted lines in the right panels are enterocyte boundary traces of the above panels to illustrate enterocyte size. The bar graph indicates the percentage of dark blue area relative to the total abdomen area; scale bar = 500 µm, n = 20. In all histograms, the error bars represent standard deviations. Significance was determined using a one-way ANOVA test in I and J and Student’s t-test for all the other graphs. ****p < 0.0001; **p < 0.01; *p < 0.05; ns not significant.