title: Epigenetic engineering reveals a balance between histone modifications and transcription in kinetochore maintenance.
abstract: Centromeres consist of specialized centrochromatin containing CENP-A nucleosomes intermingled with H3 nucleosomes carrying transcription-associated modifications. We have designed a novel synthetic biology 'in situ epistasis' analysis in which H3 dimethylated on lysine 4 (H3K4me2) demethylase LSD2 plus synthetic modules with competing activities are simultaneously targeted to a synthetic alphoidtetOHAC centromere. This allows us to uncouple transcription from histone modifications at the centromere. Here, we report that H3K4me2 loss decreases centromeric transcription, CENP-A assembly and stability and causes spreading of H3K9me3 across the HAC, ultimately inactivating the centromere. Surprisingly, CENP-28/Eaf6-induced transcription of the alphoidtetOarray associated with H4K12 acetylation does not rescue the phenotype, whereas p65-induced transcription associated with H3K9 acetylation does rescue. Thus mitotic transcription plus histone modifications including H3K9ac constitute the 'epigenetic landscape' allowing CENP-A assembly and centrochromatin maintenance. H3K4me2 is required for the transcription and H3K9ac may form a barrier to prevent heterochromatin spreading and kinetochore inactivation at human centromeres.
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, EH9 3QR, UK.
authors.forename: Oscar
authors.initials: O
authors.lastname: Molina
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, EH9 3QR, UK.
authors.forename: Giulia
authors.initials: G
authors.lastname: Vargiu
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, EH9 3QR, UK.
authors.forename: Maria Alba
authors.initials: MA
authors.lastname: Abad
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, EH9 3QR, UK.
authors.forename: Alisa
authors.initials: A
authors.lastname: Zhiteneva
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, EH9 3QR, UK.
authors.forename: A Arockia
authors.initials: AA
authors.lastname: Jeyaprakash
authors.affiliation: Department of Frontier Research, Laboratory of Cell Engineering, Kazusa DNA research Institute, Kisarazu 292-0818, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Masumoto
authors.affiliation: Genome Structure and Function Group, Developmental Therapeutics Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Natalay
authors.initials: N
authors.lastname: Kouprina
authors.affiliation: Genome Structure and Function Group, Developmental Therapeutics Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Larionov
authors.affiliation: Wellcome Trust Centre for Cell Biology, University of Edinburgh, EH9 3QR, UK.
authors.forename: William C
authors.initials: WC
authors.lastname: Earnshaw
mesh_terms: D000107:Acetylation; D045744:Cell Line, Tumor; D002503:Centromere / Q000235:genetics / Q000378:metabolism; D000076247:Centromere Protein A / Q000235:genetics / Q000378:metabolism; D044127:Epigenesis, Genetic; D006367:HeLa Cells; D006570:Heterochromatin / Q000235:genetics / Q000378:metabolism; D042421:Histone Code; D006657:Histones / Q000378:metabolism; D006801:Humans; D018386:Kinetochores / Q000378:metabolism*; D008239:Lysine / Q000378:metabolism; D008745:Methylation; D009707:Nucleosomes / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D000076247:Centromere Protein A; D006570:Heterochromatin; D006657:Histones; D009707:Nucleosomes; D008239:Lysine