title: The bone-specific Runx2-P1 promoter displays conserved three-dimensional chromatin structure with the syntenic Supt3h promoter.
abstract: Three-dimensional organization of chromatin is fundamental for transcriptional regulation. Tissue-specific transcriptional programs are orchestrated by transcription factors and epigenetic regulators. The RUNX2 transcription factor is required for differentiation of precursor cells into mature osteoblasts. Although organization and control of the bone-specific Runx2-P1 promoter have been studied extensively, long-range regulation has not been explored. In this study, we investigated higher-order organization of the Runx2-P1 promoter during osteoblast differentiation. Mining the ENCODE database revealed interactions between Runx2-P1 and Supt3h promoters in several non-mesenchymal human cell lines. Supt3h is a ubiquitously expressed gene located within the first intron of Runx2. These two genes show shared synteny across species from humans to sponges. Chromosome conformation capture analysis in the murine pre-osteoblastic MC3T3-E1 cell line revealed increased contact frequency between Runx2-P1 and Supt3h promoters during differentiation. This increase was accompanied by enhanced DNaseI hypersensitivity along with RUNX2 and CTCF binding at the Supt3h promoter. Furthermore, interplasmid-3C and luciferase reporter assays showed that the Supt3h promoter can modulate Runx2-P1 activity via direct association. Taken together, our data demonstrate physical proximity between Runx2-P1 and Supt3h promoters, consistent with their syntenic nature. Importantly, we identify the Supt3h promoter as a potential regulator of the bone-specific Runx2-P1 promoter.
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA.
authors.forename: A Rasim
authors.initials: AR
authors.lastname: Barutcu
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington, VT 05405, USA.
authors.forename: Phillip W L
authors.initials: PW
authors.lastname: Tai
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington, VT 05405, USA.
authors.forename: Hai
authors.initials: H
authors.lastname: Wu
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington, VT 05405, USA.
authors.forename: Jonathan A R
authors.initials: JA
authors.lastname: Gordon
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA Program in Bioinformatics and Integrative Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA.
authors.forename: Troy W
authors.initials: TW
authors.lastname: Whitfield
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA.
authors.forename: Jason R
authors.initials: JR
authors.lastname: Dobson
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA.
authors.forename: Anthony N
authors.initials: AN
authors.lastname: Imbalzano
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington, VT 05405, USA.
authors.forename: Jane B
authors.initials: JB
authors.lastname: Lian
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA.
authors.forename: André J
authors.initials: AJ
authors.lastname: van Wijnen
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington, VT 05405, USA.
authors.forename: Janet L
authors.initials: JL
authors.lastname: Stein
authors.affiliation: Department of Cell and Developmental Biology, University of Massachusetts Medical School, 55 Lake Avenue North, Worcester, MA 01655, USA Department of Biochemistry and Vermont Cancer Center, University of Vermont College of Medicine, 89 Beaumont Avenue, Burlington, VT 05405, USA gary.stein@uvm.edu.
authors.forename: Gary S
authors.initials: GS
authors.lastname: Stein
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D002460:Cell Line; D002843:Chromatin / Q000737:chemistry*; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics*; D006801:Humans; D051379:Mice; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism; D011401:Promoter Regions, Genetic; D026801:Synteny; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism
chemical_list: D002843:Chromatin; D050659:Core Binding Factor Alpha 1 Subunit; C493709:RUNX2 protein, human; C493716:Runx2 protein, mouse; C115819:SUPT3H protein, human; D014157:Transcription Factors