title: Epigenetic regulation of cell adhesion and communication by enhancer of zeste homolog 2 in human endothelial cells.
abstract: The histone methyltransferase enhancer of zeste homolog 2 (Ezh2) mediates trimethylation of lysine 27 in histone 3, which acts as a repressive epigenetic mark. Ezh2 is essential for maintaining pluripotency of stem cells, but information on its role in differentiated cells is sparse. Whole-genome mRNA expression arrays identified 964 genes that were regulated by >2-fold 72 hours after small interfering RNA-mediated silencing of Ezh2 in human umbilical vein endothelial cells. Among them, genes associated with the gene ontology terms cell communication and cell adhesion were significantly overrepresented, suggesting a functional role for Ezh2 in the regulation of angiogenesis. Indeed, adhesion, migration, and tube formation assays revealed significantly altered angiogenic properties of human umbilical vein endothelial cells after silencing of Ezh2. To identify direct target genes of Ezh2, we performed chromatin immunoprecipitation experiments followed by whole-genome promoter arrays (chromatin immunoprecipitation-on-chip) and identified 5585 genes associated with trimethylation of lysine 27 in histone 3. Comparative analysis with our mRNA expression data identified 276 genes that met our criteria for putative Ezh2 target genes, upregulation by >2-fold after Ezh2 silencing and association with trimethylation of lysine 27 in histone 3. Notably, we observed a striking overrepresentation of genes involved in wingless-type mouse mammary tumor virus integration site (WNT) signaling pathways. Epigenetic regulation of several of these genes by Ezh2 was specifically confirmed by polymerase chain reaction analysis of DNA enrichment after chromatin immunoprecipitation using an antibody specific for trimethylation of lysine 27 in histone 3. Combining mRNA expression arrays and chromatin immunoprecipitation-on-chip analysis, we identified 276 Ezh2 target genes in endothelial cells. Ezh2-dependent repression of genes involved in cell adhesion and communication contributes to the regulation of angiogenesis.
authors.affiliation: Medizinische Klinik für Kardiologie und Angiologie, Charité - Universitätsmedizin Berlin, Charitéplatz 1, D-10117 Berlin, Germany. henryk.dreger@charite.de
authors.forename: Henryk
authors.initials: H
authors.lastname: Dreger
authors.forename: Antje
authors.initials: A
authors.lastname: Ludwig
authors.forename: Andrea
authors.initials: A
authors.lastname: Weller
authors.forename: Verena
authors.initials: V
authors.lastname: Stangl
authors.forename: Gert
authors.initials: G
authors.lastname: Baumann
authors.forename: Silke
authors.initials: S
authors.lastname: Meiners
authors.forename: Karl
authors.initials: K
authors.lastname: Stangl
mesh_terms: D002448:Cell Adhesion / Q000235:genetics; D002450:Cell Communication / Q000235:genetics*; D002465:Cell Movement / Q000235:genetics; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation; D000071221:Enhancer of Zeste Homolog 2 Protein; D057890:Epigenomics; D020869:Gene Expression Profiling; D000076983:Histone Methyltransferases; D011495:Histone-Lysine N-Methyltransferase / Q000235:genetics / Q000378:metabolism; D006657:Histones / Q000235:genetics / Q000378:metabolism; D061307:Human Umbilical Vein Endothelial Cells / Q000378:metabolism*; D006801:Humans; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D008745:Methylation; D018919:Neovascularization, Physiologic / Q000235:genetics; D020411:Oligonucleotide Array Sequence Analysis; D063151:Polycomb Repressive Complex 2 / Q000235:genetics* / Q000378:metabolism; D034622:RNA Interference
chemical_list: D006657:Histones; D000076983:Histone Methyltransferases; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D011495:Histone-Lysine N-Methyltransferase; D063151:Polycomb Repressive Complex 2; D008239:Lysine