Fig. 4: Snail overexpression promotes stemness with minimal gene expression changes.

a Primary (left) and secondary (middle) sphere forming capacity of mOSE cells overexpressing Snail (n = 3). Data points represent the average number of spheres per 4 fields of view for each replicate. Boxplots show median value (horizontal black line), estimated 25th and 75th percentiles, and whiskers represent 1.5 times the interquartile range. Right: Phase contrast images of control (inducible GFP) and Snail-overexpressing spheroids. Spheroids were cultured for 14 days. Scale bar = 100 μm. b Plot showing the distribution of differentially expressed genes following Snail overexpression. Each point corresponds to a single gene. Selected genes related to stemness and/or the EMT are highlighted on the plot. Dashed lines correspond to significance criteria (absolute log fold change >0.5, p < 0.05). c Inferred pathway activity in control and Snail-overexpressing mOSE cells. P-values were computed from the t statistic of a linear regression model and were adjusted using the Benjamini–Hochberg FDR method. No pathway is significantly different between conditions. d GSEA results for selected gene sets enriched in differentially expressed genes following Snail overexpression. All gene sets are significantly enriched (p < 0.05) and NES values are shown. e UpSet plot showing overlaps in differentially expressed genes between all conditions assessed ranked by the condition/overlap with the largest number of genes. The top chart shows the intersection size for the conditions highlighted in the middle grid. A single, unconnected point corresponds to genes unique to only that condition. The total number of differentially expressed genes in each condition is shown in the left chart.