Fig. 1 | Signal Transduction and Targeted Therapy

Fig. 1

From: The H2BG53D oncohistone directly upregulates ANXA3 transcription and enhances cell migration in pancreatic ductal adenocarcinoma

Fig. 1

a Heat map showing the differential expression of genes in two wild-type and two H2BG53D mutant clones with two repeats. b Enrichment map of gene sets significantly overrepresented for differentially expressed genes between H2BG53D mutant and wild-type clones (Benjamini–Hochberg adjusted p < 0.05, hypergeometric tests). c Heat map showing the gene loci with differential Pol II occupancy. d Scatter plot illustrating the significant genome-wide correlation between RNA-seq and PRO-seq (Pearson correlation coefficient: 0.37, p < 2.2e−16). Ninety-nine genes with significantly elevated signals in both RNA-seq and PRO-seq are highlighted in red, among which 31 genes with significant FLAG enrichment in mutants are labeled with gene symbols and sized by the level of FLAG enrichment. e Genome viewer representations of normalized FLAG CUT&RUN reads for wild type and G53D mutant H2B (brown), H2A (yellow), and ATAC-seq reads (purple) in two wild-type (WT#7 & #16) and two H2BG53D mutant clones (G53D#3 & #32). Gray box represents area with ATAC-seq signal but without G53D-specific peak. Red boxes illustrate the overlapping of ATAC-seq and G53D-specific peaks. f Genomic distributions of the FLAG CUT&RUN H2BG53D-specific peaks (982) and randomly shuffled peaks (p < 2.2e−16, χ2 test). g Co-localization of H2BG53D-specific peaks with ATAC-seq peaks. A large proportion of promoter enriched H2BG53D peaks are co-localized with ATAC-seq peaks. h Gene set enrichment plot illustrating that the 99 genes with both significantly elevated RNA-seq and PRO-seq signals show significantly higher FLAG signal in G53D mutant than wild-type clones (p = 0.0170, permutation test, n = 10,000). The upper panel illustrates the running sum scores of GSEA (gene set enrichment analysis) random walks, the middle and lower panels show the positions of the 99 genes in the gene list ranked by FLAG log2-fold enrichment. i Genome viewer representations of normalized FLAG CUT&RUN reads for wild-type and G53D mutant H2B (brown), H2A (yellow), and ATAC-seq reads (purple) at the ANXA3 gene locus in two wild type (WT#7 & #16) and two H2BG53D mutant clones (G53D#3 & #32). j Elevated transcription of ANXA3 but not SNAP47 in H2BG53D mutant cells. Levels of transcription of the indicated genes were detected by RT-qPCR using primers at indicated intro–exon boundaries (*p < 0.05, **p < 0.01). k shRNA knockdown of ANXA3 reduced gap closure property. One representative set of data including the gap closure assay. RT-qPCR, western blotting (Supplementary Fig. 4) is shown from at least four independent repeats. Scale bar, 200 µm. l Quantification of four independent gap closure assays by Image J (*p < 0.05, **p < 0.01), error bar showing standard deviation. m A model for the effect of H2BG53D mutation on transcriptional regulation of ANXA3

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