(
A) Wpl1p and Pds5p are efficiently depleted upon auxin addition. The depletion of Wpl1p and Pds5p tagged with AID upon auxin addition to the media is shown by western-blot analysis. Tubulin is used for loading control. (
B) Wpl1p-depleted cells present more loops and more loop expansion. Contact maps from mitotically arrested cells produced with Micro-C on wild-type (
WT), cohesin Wpl1p-depleted (
WPL1-AID). Contact maps show contacts between the whole genome (first column), followed by contacts from chromosomes VIII to XI at 6.4 kb resolution (second column). The following contact maps show chromosome X and a zoomed-in region (from the full 745 kb to a 250 kb region), with 1.6 kb and 800 bp resolution respectively. Standard colormap scheme that uses the shades of red from white (no detectable interactions) to black (maximum interactions detected) in log10 scale. (
C) The loop signal at wild-type loops is still present upon Wpl1p-depletion. Heatmaps were plotted with 200 bp data resolution for wild-type (
WT) and Wpl1p-depleted (
Wpl1-AID) cells in ±5 kb regions surrounding the wild-type loop anchors. (
D) Distribution of contacts upon Wpl1p-depletion and Pds5p-depletion. Interactions-versus-distance decaying curve shows the normalized contact density (y-axis) against the distance between the pair of crosslinked nucleosomes from 100 bp to 1 Mb (x-axis) for wild-type (
WT in blue), Wpl1p-depleted (
WPL1-AID in green), and Pds5p-depleted cells (PDS5-AID in purple) (left). Plot of the slopes derived from the interactions-versus-distance decaying curves for the same datasets along with Mcd1p-depleted cells (
MCD1-AID in red) (right). The dotted line indicates the size of high contact enrichment (or the slowest contact decay with distance) for each strain. (
E) Loop signal is detected till the +10 CARs interval upon Wpl1p-depletion. Quantification of the heatmap analysis in
Figure 5D for Wpl1p-depleted (
WPL1-AID in green) cells compared to wild-type (
WT in blue) and cohesin-depleted (
MCD1-AID in red). The chart shows the loop enrichment (Log2 of the ration of the loop signal in the center of the heatmap divided for the background signal in the corner) on the y-axis present at each CAR interval (x-axis). (
F) Pds5p-depleted cells present no loops on chromosome arms. Contact maps from mitotically arrested cells produced with Micro-C on wild-type (
WT), Pds5p-depleted (
PDS5-AID). Contact maps show contacts between chromosomes VII to X at 6.4 kb resolution (first column). The following contact maps show chromosome VII and zoomed-in regions (from the full 1090 kb to a 125 kb region), from 1.6 kb to 400 bp resolution. Standard colormap scheme that uses the shades of red from white (no detectable interactions) to black (maximum interactions detected) in log10 scale.