title: Nucleosome exclusion from the interspecies-conserved central AT-rich region of the Ars insulator.
abstract: The Ars insulator is a boundary element identified in the upstream region of the arylsulfatase (HpArs) gene in the sea urchin, Hemicentrotus pulcherrimus, and possesses the ability to both block enhancer-promoter communications and protect transgenes from silent chromatin. To understand the molecular mechanism of the Ars insulator, we investigated the correlation between chromatin structure, DNA structure and insulator activity. Nuclease digestion of nuclei isolated from sea urchin embryos revealed the presence of a nuclease-hypersensitive site within the Ars insulator. Analysis of micrococcal nuclease-sensitive sites in the Ars insulator, reconstituted with nucleosomes, showed the exclusion of nucleosomes from the central AT-rich region. Furthermore, the central AT-rich region in naked DNA was sensitive to nucleotide base modification by diethylpyrocarbonate (DEPC). These observations suggest that non-B-DNA structures in the central AT-rich region may inhibit nucleosomal formation, which leads to nuclease hypersensitivity. Furthermore, comparison of nucleotide sequences between the HpArs gene and its ortholog in Strongylocentrotus purpuratus revealed that the central AT-rich region of the Ars insulator is conserved, and this conserved region showed significant enhancer blocking activity. These results suggest that the central AT-rich nucleosome-free region plays an important role in the function of the Ars insulator.
authors.affiliation: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Higashi-Hiroshima 739-8526, Japan.
authors.forename: Haruna
authors.initials: H
authors.lastname: Takagi
authors.forename: Yuta
authors.initials: Y
authors.lastname: Inai
authors.forename: Shun-ichiro
authors.initials: S
authors.lastname: Watanabe
authors.forename: Sayuri
authors.initials: S
authors.lastname: Tatemoto
authors.forename: Mamiko
authors.initials: M
authors.lastname: Yajima
authors.forename: Koji
authors.initials: K
authors.lastname: Akasaka
authors.forename: Takashi
authors.initials: T
authors.lastname: Yamamoto
authors.forename: Naoaki
authors.initials: N
authors.lastname: Sakamoto
mesh_terms: D020856:AT Rich Sequence / Q000235:genetics*; D000818:Animals; D001192:Arylsulfatases / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D017124:Conserved Sequence / Q000235:genetics*; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D003851:Deoxyribonucleases / Q000378:metabolism; D004625:Embryo, Nonmammalian / Q000196:embryology / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D047330:Hemicentrotus / Q000196:embryology / Q000235:genetics; D006657:Histones / Q000235:genetics / Q000378:metabolism; D038101:Insulator Elements / Q000235:genetics*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D009707:Nucleosomes / Q000235:genetics* / Q000378:metabolism; D016133:Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid; D013045:Species Specificity; D047270:Strongylocentrotus purpuratus / Q000235:genetics
chemical_list: D002843:Chromatin; D006657:Histones; D009707:Nucleosomes; D004247:DNA; D004259:DNA-Directed DNA Polymerase; D003851:Deoxyribonucleases; D001192:Arylsulfatases