title: Down-regulation of miR-101 in endothelial cells promotes blood vessel formation through reduced repression of EZH2.
abstract: Angiogenesis is a balanced process controlled by pro- and anti-angiogenic molecules of which the regulation is not fully understood. Besides classical gene regulation, miRNAs have emerged as post-transcriptional regulators of angiogenesis. Furthermore, epigenetic changes caused by histone-modifying enzymes were shown to modulate angiogenesis as well. However, a possible interplay between miRNAs and histone-modulating enzymes during angiogenesis has not been described. Here we show that VEGF-mediated down-regulation of miR-101 caused pro-angiogenic effects. We found that the pro-angiogenic effects are partly mediated through reduced repression by miR-101 of the histone-methyltransferase EZH2, a member of the Polycomb group family, thereby increasing methylation of histone H3 at lysine 27 and transcriptome alterations. In vitro, the sprouting and migratory properties of primary endothelial cell cultures were reduced by inhibiting EZH2 through up-regulation of miR-101, siRNA-mediated knockdown of EZH2, or treatment with 3-Deazaneplanocin-A (DZNep), a small molecule inhibitor of EZH2 methyltransferase activity. In addition, we found that systemic DZNep administration reduced the number of blood vessels in a subcutaneous glioblastoma mouse model, without showing adverse toxicities. Altogether, by identifying a pro-angiogenic VEGF/miR-101/EZH2 axis in endothelial cells we provide evidence for a functional link between growth factor-mediated signaling, post-transcriptional silencing, and histone-methylation in the angiogenesis process. Inhibition of EZH2 may prove therapeutic in diseases in which aberrant vascularization plays a role.
authors.affiliation: Neuro-oncology Research Group, Department of Neurosurgery, VU University Medical Center, Amsterdam, The Netherlands.
authors.forename: Michiel
authors.initials: M
authors.lastname: Smits
authors.forename: Shahryar E
authors.initials: SE
authors.lastname: Mir
authors.forename: R Jonas A
authors.initials: RJ
authors.lastname: Nilsson
authors.forename: Petra M
authors.initials: PM
authors.lastname: van der Stoop
authors.forename: Johanna M
authors.initials: JM
authors.lastname: Niers
authors.forename: Victor E
authors.initials: VE
authors.lastname: Marquez
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Cloos
authors.forename: Xandra O
authors.initials: XO
authors.lastname: Breakefield
authors.forename: Anna M
authors.initials: AM
authors.lastname: Krichevsky
authors.forename: David P
authors.initials: DP
authors.lastname: Noske
authors.forename: Bakhos A
authors.initials: BA
authors.lastname: Tannous
authors.forename: Thomas
authors.initials: T
authors.lastname: Würdinger
mesh_terms: D042501:Angiogenic Proteins / Q000502:physiology; D000818:Animals; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D015536:Down-Regulation / Q000235:genetics; D042783:Endothelial Cells / Q000378:metabolism / Q000502:physiology*; D000071221:Enhancer of Zeste Homolog 2 Protein; D006657:Histones / Q000378:metabolism; D006801:Humans; D008745:Methylation; D051379:Mice; D035683:MicroRNAs / Q000235:genetics*; D018919:Neovascularization, Physiologic; D063151:Polycomb Repressive Complex 2; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A / Q000502:physiology
chemical_list: D042501:Angiogenic Proteins; D004268:DNA-Binding Proteins; D006657:Histones; C534072:MIRN101 microRNA, human; D035683:MicroRNAs; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; C498409:EZH2 protein, human; D000071221:Enhancer of Zeste Homolog 2 Protein; D063151:Polycomb Repressive Complex 2