Fig. 5: Heterochromatin domains become smaller upon WAPL-depletion.

a Schematic representation of chromatin loop extrusion. WAPL-depletion results in increased cohesin loading and excessive transcription-driven chromatin loop extrusion. Note that nucleosomes, despite being present, are not shown to improve clarity. b Numerical prediction of distribution of heterochromatin domains in the interior and the LADs along the periphery (all domains in red) in a nucleus without (Cas9) and with (WAPLΔ) cohesin unloading disruption. c Heatmap density of DNA super-resolution images in d control (Cas9, left panel) and WAPL knock-out (WAPLΔ) treated HeLa nuclei. All scale bars - 3 μm. d Left: Zoomed in views of Cas9 and WAPLΔ treated nuclei focusing on the interior heterochromatin domains. White solid boxes shown in c are zoomed into. All scale bars - 1 μm. Right: Zoomed in views of Cas9 and WAPLΔ treated nuclei along the nuclear periphery. White dashed boxes shown in c are zoomed into. All scale bars - 1 μm. e Quantification of heterochromatin domain radius in the interior of Cas9- and WAPLΔ - treated nuclei. (n = 2386 loci in 6 nuclei for Cas9-treatment and 2416 loci in 7 nuclei for WAPLΔ treatment). WAPLΔ treated nuclei exhibit a significantly lower (\(\sim\) 0.86 times) mean heterochromatin radius (unpaired two-tailed t-test, p = 6e−10). Quantification of LAD thickness along the periphery of Cas9- and WAPLΔ - treated nuclei. (n = 219 loci in 6 nuclei for Cas9-treatment and 169 loci in 7 nuclei for WAPLΔ treatment). WAPLΔ treated nuclei exhibit a significantly lower (\(\sim\) 0.43 times) mean LAD thickness (unpaired two-tailed t-test, p = 1e−13). f Boxplot in left panel shows the distribution of domain radii predicted numerically. WAPLΔ nuclei have a mean domain radius 0.8 times that of Cas9-treated nuclei (unpaired two-tailed t-test, p = 0). Boxplot in right panels shows the distribution of LAD thicknesses predicted numerically. WAPLΔ nuclei have a mean LAD thickness 0.82 times that of Cas9-treated nuclei. All boxplots show the mean (cross), median (horizontal line), upper and bottom quartiles (box outlines) and the maximum and minimum non-outlier data points (whiskers) of the plotted distribution. All source data are provided as a source data file.