Fig. 4

KDM6B promotes C/EBPβ transcription via modulating H3K27 methylation. a RNA-seq results showed the gene expression differentiation between the sh-NC group and the sh-KDM6B-1 group through heat map analysis. Compared with the sh-NC group, the number of down-regulated genes was 1371 and the number of up-regulated genes was 542 in the sh-KDM6B-1 group (fold change≥2,p < 0.05). Red means up, blue means down. The data are representative of 2 biological repeats. b GSEA analysis of the RNA-seq data showed the enriched pathway of TNFA_SIGNALING_VIA_NFκB, INTERFERON_ GAMMA_RESPONSE, P53, and IL6_JAK_STAT3_SIGNALING pathway in KYSE150 cell that KDM6B was knockdown as compared with Ctrl cells. ES, enrichment score; FDR, false discovery rate. c,d RT-qPCR verified the expression of target genes after KDM6B was knocked down in KYSE150. e RT-qPCR verified the expression of target genes after over-expressing KDM6B in Eca9706. f,g Correlation between the expression levels of KDM6B and C/EBPβ, a key gene of TNFA_SIGNALING_VIA_NFκB signaling pathways in ESCC patient datasets from TCGA(f) and our ESCC simple(g). All expression data were log (2) transformed. The correlation coefficient (r) and p values were detected by “Pearson Correlation”. h Western blot was used to detect the expression of H3K27me3 in KYSE150 and TE10 cells after the KDM6B knockdown. i Western blot detected the expression of H3K27me3 in Eca9706, TE11 after over-expressing KDM6B for 72 h. j Genomic visualization analysis shows a representative snapshot of the H3K27me3 binding site at the C/EBPβ locus in KYSE150 cells in the control and KDM6B knockdown groups. The long red line is the promoter region, and the short black line is the primer design site. The arrow is the direction of gene transcription. k ChIP-qPCR analysis of H3K27me3 ChIP products from the control and shRNA or GSK-J4 treated KYSE150 cells (n = 3, values are expressed as mean ± s.d.). * p < 0.05, ** p < 0.01, *** p < 0.001