Extended Data Figure 2: Inhibition of LATS promotes a luminal phenotype.

a, Heat map derived from hierarchical clustering of the 182 genes that were differentially expressed between shNT-treated PHBECs (n = 5) and shLATS-treated PHBECs (n = 6). Cut-off: adjusted P < 0.01, fold change > 2.0, expression values > 4. Arrows indicate canonical luminal genes as well as LATS1. b, Principal component analysis (PCA) of microarrays performed on shNT- and shLATS-treated PHBECs. shNT-treated samples cluster separately from both shLATS1B- and shLATS1 + 2-treated samples. c, Venn diagram analysis of differentially regulated genes at a low stringency cut-off of P < 0.05, 2.0-fold change, expression values > 4.0. Both LATS shRNAs show a high degree of overlap when compared to shNT-treated samples and no genes were significantly differentially regulated between cells treated with the two shRNAs. d, shLATS-treated PHBEC expression profiles were significantly enriched in genes that are downregulated in normal basal breast cells. GSEA with shLATS-specific genes and a gene set of normal basal/stem breast cells. shNT-treated PHBECs (n = 5), shLATS-treated PHBECs (n = 6). NES = 2.5; FDR < 0.0001; P < 0.0001. e, shLATS-treated PHBECs express low levels of basal genes. Heatmaps with average normalized expression values of a subset of genes associated with normal basal breast cells. f, Removal of LATS imposes a luminal breast cancer signature on PHBECs. Left, GSEA with shLATS-specific genes derived from PHBECs and gene sets of ERα-positive (left graph) and luminal breast cancer samples (right graph). shNT-treated PHBECs (n = 5), shLATS-treated PHBECs (n = 6). NES = 2.29 (left) and 1.98 (right); FDR <0.0001; P < 0.0001. Right, clustered heatmap showing correlation coefficients between human breast cancer profiles and shNT- or shLATS-treated PHBECs. g, Graphs of ISMARA transcription factor activity analysis of shLATS- (n = 6) versus shNT-treated (n = 5) PHBECs showing high activity upon removal of LATS for CEBPβ (Z = 1.92; P < 0.0001) and STAT5A,B (Z = 2.184, P < 0.01). h, qPCR analysis of shNT- and shLATS-treated PHBECs. mRNA levels of ERα and c-KIT as well as several canonical ERα-target genes are higher in shLATS-treated PHBECs (n = 6 experimental and 2 technical replicates), i, FACS analysis of PHBECs with LATS knockdown. Markers of luminal progenitors (c-KIT, ALDH activity) and luminal epithelial (EPCAM) cells increased and expression of the basal marker CD10 was reduced upon removal of LATS (n = 6 experimental replicates). Data are mean ± s.d. (h, i); *P < 0.05, **P < 0.01; NS, not significant.