a, High DCC occupancy correlates with TAD boundaries lost or reduced upon DCC depletion. Top, ChIP-seq profiles of DCC subunit SDC-3 in wild-type (red) and DC mutant (green) embryos. Y-axis, reads per million (RPM) normalized to IgG control. Middle, insulation profiles of wild-type (red) and DC mutant (green) embryos. Bottom, insulation difference plot for wild-type insulation profile subtracted from DC mutant profile. Black lines, TAD boundary locations. Blue dots, boundaries with insulation changes > 0.1 between wild-type and DC mutant embryos. Red lines, locations of 25 highest DCC-occupied rex sites. Cyan bars, sites with the largest insulation loss. b, Confocal images of embryonic nuclei of various genotypes stained with a DNA intercalating dye (blue) and 500 kb FISH probes around the rex-47 TAD boundary. c, d, e, Quantification of FISH probe colocalization confirms DCC-dependent and DCC-independent boundaries found by Hi-C. Box plots, distribution of Pearson’s correlation coefficients between pairwise combinations of FISH probes within (blue) or across (orange) TADs. Boxes, middle 50% of coefficients. Center bars, median (M) coefficients. N, total number of nuclei. Asterisks of same color indicate values compared with one-tailed Mann-Whitney U test. ns, not significant.