Extended Data Fig. 8: Downstream mTOR activity influences progenitor expansion.

a) Heat map showing scaled expression of differentially expressed genes in DHT-treated compared to control organoids, detected by scRNA-seq. Cell identities were assigned to their treatment group and differential expression analysis performed in Seurat as performed for comparison across clusters (see Methods). b) Dot plot showing the relative expression of bRG markers, overlapping markers of RG1 cells and DE transcripts in DHT, and genes of the mTOR pathway in cells from control, DHT and E treated organoids, detected by scRNA-seq. c) Protein interaction network of upregulated DEG directly interacting genes, obtained by bulk RNA seq. Translation/ribosome biogenesis genes are in red. d) Immunostaining for phosphorylated S6 (PS6) (fire look-up table), an indicator of mTOR activity, in 35d old XX organoids, treated with DHT and E. White arrowheads indicate PS6+ cell bodies in the ventricular zone (VZ). e) Quantification of PS6+ cells in the VZ of control, DHT- and E-treated 35d old XX organoids. f) Immunostaining for TBR2 (white) in control and MHY-1485-treated 35d XX organoids. Co-stained with DAPI (blue). g) Quantification of TBR2+ cells per mm² progenitor layer in control and MHY-1485-treated 35d XX organoids. Scale bars: d), f) 50µm. See Methods for details of statistics and Supplementary Table 5 for details of n numbers.