Fig. 4: Breast cancer cells depleted of Kaiso are enriched for autophagy terms and show a defect in LC3A/B maturation.

a Differential gene expression analysis of wild type TNBC cell line MDA-MB-231 versus MDA-MB-231 cells depleted of Kaiso by RNAi shows a significant overlap of differentially expressed genes with an autophagy-related gene list (also see Supplementary Fig. 6, and Supplementary Table 3) (p-value for the significance of the overlap is provided by the hypergeometric test). b Gene set enrichment analysis shows significant enrichment for autophagy terms in MDA-MB-231 cells that differentially express Kaiso (Supplementary Data 4). c Analysis of autophagocytic flux in WT versus MDA-MB-231 depleted of Kaiso by RNAi reveal a significant defect in autophagy demonstrated by decreased autophagocytic puncta formation in cells depleted of Kaiso. d Quantitative GFP-LC3 immunofluorescent analysis of LC3 puncta formation (average puncta per cell before (left) and after (right) addition of chloroquine in MDA-MB-231 cells expressing GFP-LC3). e Immunoblot analysis of LC3A/B lipidation (GFP-LC3ii formation) in MDA-MB-231 transfected with scramble control RNAi versus 3 different short hairpins against Kaiso. f Quantitative densitometer scan (N = 4 biological replicates each with N = 3 technical replicates). (*) = p-value < 0.05, (***) = p-value < 0.001 (t-test) for each unique Kaiso RNAi hairpin. g Immunofluorescent colocalization of Kaiso (green) and LC3A/B (red) in MCF-7 and MDA-MB-231 cells. h Quantitative profiling of Kaiso and LC3A/B colocalization in both the cytoplasm and nucleus (overall) of MCF-7 and MDA-MB-231 cells.