Fig. 4

Demethylating agent 5-azacytidine (AZA) can partially revert DNA hypermethylation of CpG islands in the hCIMP ACC cell line H295R restoring activity of immune response genes. a Overlap between genes significantly up- or down-regulated upon AZA treatment (FDR < 0.05) in H295R (5242 genes), MUC-1 (125 genes), and HUVEC (254 genes) cell lines. b Unsupervised hierarchical clustering of 3 cell lines (H295R, MUC-1 and HUVEC) treated or not with AZA 5 μM was based on their promoter CpG islands methylation profile. The clustering was obtained using the CpG with the most variable methylation level (s.d. > 0.4). c Global percentage of methylated CpG among the CpG covered by RRBS reads. AZA induces a decrease in CpG methylation in all three cell lines, but this effect is less pronounced in the MUC-1 cell line that has the lowest basal level of DNA methylation. d Pathway enrichment analysis on genes induced by AZA in H295R through promoter demethylation. Pathways related to the immune response are highlighted in red. e Examples of differentially methylated region (DMRs) upon AZA treatment in H295R located in promoters of two immune response genes