MMSET alters EZH2 binding in t(4;14)+ myeloma cells.

<p>(A) Venn diagram showing overlap of genes bound at their promoters by EZH2 in NTKO (blue) and TKO (yellow) cells. (B) UCSC genome browser display of H3K27me3 (top, gray) and EZH2 binding (bottom, red) in NTKO cells. (C) UCSC genome browser display of EZH2 ChIP-seq tracks in NTKO (top, red) and TKO (bottom, green) cells associated with MMSET-repressed genes, <i>CDCA7</i> (left) and <i>DLL4</i> (right). (D) Heat map of over-represented gene categories among genes bound by EZH2 in either NTKO cells, TKO cells or both cell types. Enrichment was measured using iPAGE analysis <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004566#pgen.1004566-Goodarzi1" target="_blank">[74]</a>. (E) Motif analysis using HOMER <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004566#pgen.1004566-Heinz1" target="_blank">[73]</a> identified conserved sequences bound by EZH2 in NTKO cells, TKO cells or both cell types. (F) Relative cell number of MMSET-high and MMSET-low cells treated with indicated doses of the EZH2 small molecule inhibitor (GSK343). Inactive compound, GSK669, was used as a control. Graph represents four independent experiments +/− standard deviation. (G) Quantitative RT-PCR measurement of miR-126* expression in KMS11 cells treated with GSK669 or GSK 343. Graph represents average expression from three independent experiments +/− standard deviation (* p<0.05). (H) Representative immunoblot of nuclear extracts from MMSET-high (KMS11) and MMSET-low (TKO) cells treated for seven days with 2 µM EZH2 inhibitor (GSK343) or inactive control (GSK669). This experiment was performed in biological triplicate.