Fig. 2: SNAI1 knockout generates an intermediate mesenchymal to epithelial phenotype.

A Volcano plot depicting the up- and down-regulated genes in MDA-MB-231 SNAI1-KO cells relative to WT cells. The horizontal dashed line represents the statistical significance threshold (FDR < 0.05) and the vertical lines indicate the expression fold-change threshold (−2 ≤ log₂ ≥ 2). B Venn diagram showing the number of differentially expressed genes in SNAI1-KO CS16 and CS19 clones relative to WT cells. C Heatmap indicating the correlation coefficients among MDA-MB-231-WT, CS16 and CS19 SNAI1-KOs based on whole transcriptome profiling. D Manhattan plot (upper panel) illustrating the pathway-enrichment analysis of the 5957 differentially expressed genes in SNAI KOs retrieved from different databases. Twenty-four GO terms are circled and numbered in the plot and listed in detail in the table. GO:BP, gene ontology biological processes; REAC, Reactome; TF, transcription factors. The p-values are adjusted for multiple testing and are also color-coded. E Phase-contrast images showing cell morphology in MDA-MB-231-WT and SNAI1-KO cells. Scale bars = 100 μm. F Representative immunofluorescence staining pictures of CDH1, ZO1, and F-actin (green) in MDA-MB-231-WT, SNAI1-KOs and ZR-75-1. Nuclei are stained in blue with DAPI. Scale bars = 50 μm. G Protein expression levels of the EMT transcription factors ZEB1, SNAI2, the mesenchymal proteins FN1, VIM and the epithelial protein CDH1 in MDA-MB-231-WT and SNAI1-KO cells. H Protein expression levels of the aforementioned proteins in MDA-MB-231-WT and SNAI1-KO cells stimulated with 5 ng/ml of TGFβ1 for the indicated time periods. HP95 serves as loading control. E and F show representative photomicrographs from experiments performed in three independent biological replicates. G and H show representative immunoblots of three independent biological replicates along with molecular mass markers in kDa.