Figure 7.

ERG (ETS-related gene) regulates distinct endothelial super-enhancers (SEs). A, SE identification on siCtl H3K27ac using H3K27ac enrichment from siCtl and siERG-treated human umbilical vein endothelial cell (HUVEC). Selected genes relevant to endothelial function are indicated in red (see also Online Table VIII). B, Volcano plot depicting the log₂FC vs –log (P value) of differential SEs at 1015 H3K27ac SE identified in siCtl-treated cells. Significantly up or downregulated SEs are selected according to −0.58≥log₂FC≥0.58 (−1.5≥FC≥1.5); −log (P value)≥4. C, Boxplot representing the log₂FC of MED (Mediator complex subunit)-1 occupancy in siERG-treated HUVEC in decreased SE and constant SE. ****P<0.0001, Wilcoxon rank-sum test. D, Chromatin immunoprecipitation-quantitative polymerase chain reaction (ChIP-qPCR) of p300 enrichment in siCtl or siERG-treated HUVEC at selected SE constituent enhancers (E) associated with decreased SE (CLDN5 [claudin-5], DLL4 [Delta-like protein 4] or constant SE (IL6, PXN). Data are represented as fold change over IgG, n=4. *P<0.05, paired 2-tailed t test. E, Genomic occupancy of ERG and collaborative TFs: GATA-2 (GATA-binding protein 2), cFOS, and cJUN at decreased SE compared with constant SE, measured by the chromatin immunoprecipitation with deep sequencing signal in HUVEC. ****P<0.0001; ***P<0.001; *P<0.05, Wilcoxon rank-sum test. F, Model of ERG-dependent SE assembly in EC. Cooperative SE activation is associated with a strong transcriptional complex (ERG, AP-1 [activator protein 1], GATA-2) at constant SE (left). In a subset of SE, activation of SE is strongly dependent on ERG due to less abundance of transcription factor network partners with reduced cooperativity (right).