Fig. 3: Deletion of a CTCF site near the MYC promoter leads to re-organization of CTCF looping.

a The scatter plot showing the fold changes of CTCF binding affinities at CTCF sites looping to gene promoters and the expression fold changes of corresponding genes. The interactions between CTCF sites and gene promoters were determined by CTCF HiChIP loops. VCaP and 22Rv1 CTCF ChIP-Seq data were obtained from ENCODE and gene expression data were retrieved from GSE25183. The CTCF-gene pairs with consistent binding/expression fold changes and both absolute fold changes > 1.5 are labelled red, and opposite fold changes are labelled blue. The dots and triangles indicate CTCF sites with and without H3K27ac overlapping, respectively. Genes in representative CTCF-gene pairs were annotated. n = 2088. P-value was calculated by Chi-square test. b Upper: The highlighted CTCF site was connected to MYC promoter by a CTCF loop. The CTCF binding affinities at this site (−10 Kb from MYC promoter) were negatively correlated with MYC expression in prostate cancer cell lines. The CTCF site −10 Kb from MYC promoter was deleted by CRISPR/Cas9-mediated knock-out in 22RV1 cells. The control (sgCtrl) and CTCF deletion (sgDele-10Kb) cells were then used for H3K27ac and CTCF HiChIP experiments. Bottom: Significantly changed H3K27ac loops at MYC region by the “−10 Kb CTCF site” deletion. For each group of H3K27ac HiChIP, two biological replicates were performed. c Number of significantly changed CTCF loops at each chromosome before and after “−10Kb CTCF site” deletion. d Enrichment analysis of MYC binding at the anchors of dysregulated CTCF or H3K27ac loops. Orange points represent the actual ratio of dysregulated loop anchors with MYC binding. Each box represents 500-time random sampling from all CTCF or H3K27ac loop anchors. Same number of loop anchors as in the sgCtrl-specific or sgDele-10Kb-specific anchor set was used for random sampling, respectively. O/E (observed vs. expected) was calculated by comparing the overlap percentage of actual dysregulated loop anchors with that of the average of randomly sampled anchors. Box plots indicating the mean (middle line), 25th and 75th percentile (box), and 10th and 90th percentile (whiskers). Points were highlighted by red if P < 0.05. P-values were determined by Student’s t-test. n = 215, 297, 2896, 2322 from left to right, respectively. e Motifs enriched in the CTCF peaks at CTCF anchors of indicated loops. f Motif distribution in CTCF peaks at CTCF anchors of indicated loops. For each CTCF peak, the ___location of the best-ranked CTCF motif was used as the center for the motif density plot. g The aggregated CTCF ChIP-Seq signal in 22Rv1 cells at indicated peaks.