title: Direct screening for chromatin status on DNA barcodes in yeast delineates the regulome of H3K79 methylation by Dot1.
abstract: Given the frequent misregulation of chromatin in cancer, it is important to understand the cellular mechanisms that regulate chromatin structure. However, systematic screening for epigenetic regulators is challenging and often relies on laborious assays or indirect reporter read-outs. Here we describe a strategy, Epi-ID, to directly assess chromatin status in thousands of mutants. In Epi-ID, chromatin status on DNA barcodes is interrogated by chromatin immunoprecipitation followed by deep sequencing, allowing for quantitative comparison of many mutants in parallel. Screening of a barcoded yeast knock-out collection for regulators of histone H3K79 methylation by Dot1 identified all known regulators as well as novel players and processes. These include histone deposition, homologous recombination, and adenosine kinase, which influences the methionine cycle. Gcn5, the acetyltransferase within the SAGA complex, was found to regulate histone methylation and H2B ubiquitination. The concept of Epi-ID is widely applicable and can be readily applied to other chromatin features.
authors.affiliation: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Hanneke
authors.identifier: 0000-0003-1743-6428
authors.initials: H
authors.lastname: Vlaming
authors.affiliation: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Thom M
authors.initials: TM
authors.lastname: Molenaar
authors.affiliation: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Tibor
authors.initials: T
authors.lastname: van Welsem
authors.affiliation: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Deepani W
authors.initials: DW
authors.lastname: Poramba-Liyanage
authors.affiliation: Department of Clinical Chemistry, Metabolic Laboratory, VU University Medical Center, Amsterdam, Netherlands.
authors.forename: Desiree E
authors.initials: DE
authors.lastname: Smith
authors.affiliation: Central Genomics Facility, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Arno
authors.initials: A
authors.lastname: Velds
authors.affiliation: Mass Spectrometry/Proteomics Facility, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Liesbeth
authors.initials: L
authors.lastname: Hoekman
authors.affiliation: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Tessy
authors.initials: T
authors.lastname: Korthout
authors.affiliation: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Sjoerd
authors.initials: S
authors.lastname: Hendriks
authors.affiliation: Mass Spectrometry/Proteomics Facility, Netherlands Cancer Institute, Amsterdam, Netherlands.|Biomolecular Mass Spectrometry and Proteomics, Utrecht Institute for Pharmaceutical Sciences, University of Utrecht, Utrecht, Netherlands.
authors.forename: A F Maarten
authors.initials: AFM
authors.lastname: Altelaar
authors.affiliation: Division of Gene Regulation, Netherlands Cancer Institute, Amsterdam, Netherlands.
authors.forename: Fred
authors.identifier: 0000-0002-7267-7251
authors.initials: F
authors.lastname: van Leeuwen
mesh_terms: D002843:Chromatin / Q000737:chemistry*; D047369:Chromatin Immunoprecipitation; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D005820:Genetic Testing; D005827:Genetics, Microbial / Q000379:methods; D011495:Histone-Lysine N-Methyltransferase / Q000378:metabolism*; D006657:Histones / Q000378:metabolism*; D008745:Methylation; D008967:Molecular Biology / Q000379:methods; D009687:Nuclear Proteins / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D017422:Sequence Analysis, DNA
chemical_list: D002843:Chromatin; D004271:DNA, Fungal; D006657:Histones; D009687:Nuclear Proteins; D029701:Saccharomyces cerevisiae Proteins; C416235:Dot1 protein, S cerevisiae; D011495:Histone-Lysine N-Methyltransferase
keywords: DNA repair; Dot1; H3K79 methylation; S. cerevisiae; SAGA; adenosine kinase; chromosomes; genes; histone modifications