Fig. 2: MyoD is an anchor protein for chromatin loop formation in muscle cells.

a Aggregate Peak Analysis (APA) plots showing the aggregated Hi-C contacts around chromatin loops identified in WT-GM and WT-DM cells. n represents the total number of chromatin loops called by HiCCUPS (see Methods). The APA score at the upper left corner of each aggregation plot was calculated as the ratio of Hi-C contacts at the central pixel to the mean Hi-C contacts of the lower-left pixels. The Hi-C contacts at the central pixel are shown at upper right corner of each aggregation plot. b Enriched motifs in accessible regions at the anchors of the chromatin loops identified in panel a. p-values of motif enrichment were determined using cumulative binomial distributions, as applied by Homer. c Percentage of chromatin loops with anchors bound by MyoD in WT-GM and WT-DM cells. MyoD is indicated as being bound at both anchors (Both, red), one of the two anchors (Xor, blue), or neither of the two anchors (Neither, green). Pseudo-peaks of MyoD binding derived from combined ChIP-seq data of public databases and our study were used for this analysis (Methods). d Classification of the four types of chromatin loops identified in the study. MyoD-MyoD(noCTCF) loops had both anchors bound only by MyoD. MyoD-MyoD(CTCF) loops had both anchors bound by MyoD and either anchor concomitantly bound by CTCF. MyoD-CTCF loops had only one anchor bound by MyoD and either anchor bound with CTCF. CTCF-CTCF loops had both anchors bound only by CTCF. e Representative examples of the MyoD-bound chromatin loops. ChIP-seq data of SMC3 in C2C12 cells were obtained from the public database (GSE113248). f dSTORM image showing colocalization of MyoD (green) and CTCF (red) from immunofluorescent staining of C2C12 cells grown in DM for 24 h and stained with antibodies against MyoD and CTCF. The image is representative of three independent experiments. g APA score dynamics for the four types of chromatin loops among WT and MKO cells grown in GM and DM.