Figure 4. YAP/TAZ and BRD4 regulate Pol II loading and H3K122 acetylation on TSSs.

a) Box-and whiskers plots showing the distribution of RNA-Pol II ChIP-seq signal (expressed as normalized read density, RPKM) comparing promoters of genes not activated by YAP/TAZ (n=8026) or of YAP/TAZ target genes (n=616) in control (DMSO) or YAP/TAZ depleted cells. The box includes values within the 25^th and 75^th percentile (with the median highlighted by the line in the middle) and whiskers extend from the 5^th to the 95^th percentile. **** p<10-10 (one-tailed Mann-Whitney U test)

b) Heatmap showing RNA-Pol II loading on the promoters of YAP/TAZ targets in MDA-MB-231 cells, in a window of ±1.5kb centered on the transcription start site (TSS).

c) Linear correlation between BRD4 and RNA-Pol II occupancy (both expressed in RPKM) on the TSS of YAP/TAZ target genes (n=616). r² was calculated using linear regression analysis (F-test p-value<0.0001).

d) Box-and whiskers plots (defined as in a) showing the change in RNA-Pol II promoter occupancy in JQ1-treated cells vs. control cells (DMSO), comparing promoters of genes not activated by YAP/TAZ (n=8026) with promoters of YAP/TAZ target genes (n=616). **** p<10-10 (one-tailed Mann-Whitney U test)

e) Genome browser view of RNA-Pol II binding profiles at representative promoters of YAP/TAZ target genes or not-YAP/TAZ targets. Pol II binding is reduced upon JQ1 treatment or YAP/TAZ depletion on the TSS of YAP/TAZ targets.

f) ChIP-qPCR verifying RNA-Pol II binding to promoters of established YAP/TAZ targets upon depletion of BET proteins. GAPDH promoter represents a non-YAP/TAZ target. ChIP with pre-immune IgG displayed background signal (which was comparable in all samples). DNA enrichment was calculated as fraction of input and is presented as % of RNA-Pol II binding in control cells (siCO).

g) Box-and whiskers plots of H3K122ac ChIP-seq signal (RPKM) showing its enrichment on YAP/TAZ target genes (n=616) in comparison with inactive promoters (n=4618) and not-YAP/TAZ targets (n=8026). Data are presented as in a. **** p<10^-10 (one-tailed Mann-Whitney U test)

h) Box-and whiskers plots showing the change in H3K122ac promoter levels in YAP/TAZ-depleted (left) or JQ1-treated cells (right), both compared to control cells (DMSO), showing preferential loss of H3K122ac at promoters of YAP/TAZ target genes (n=616) vs. promoters of not YAP/TAZ targets (n=8026). Data are presented as in a. **** p<10^-10 (one-tailed Mann-Whitney U test)

i) Heat map showing acetylation of H3K122 on the promoters of YAP/TAZ targets in MDA-MB-231 cells, in a window of ±1.5kb centered on the transcription start site (TSS).

j) Average ChIP-seq profile of H3K122ac on the promoters of YAP/TAZ target genes (n=616) in MDA-MB-231 cells, in a window of ±1.5 kb centered on TSS.

k) Genome browser view of H3K122ac levels at representative promoters of YAP/TAZ target genes vs. not-YAP/TAZ targets.