title: N-terminal phosphorylation of HP1α increases its nucleosome-binding specificity.
abstract: Heterochromatin protein 1 (HP1) is an evolutionarily conserved chromosomal protein that binds to lysine 9-methylated histone H3 (H3K9me), a hallmark of heterochromatin. Although HP1 phosphorylation has been described in several organisms, the biological implications of this modification remain largely elusive. Here we show that HP1's phosphorylation has a critical effect on its nucleosome binding properties. By in vitro phosphorylation assays and conventional chromatography, we demonstrated that casein kinase II (CK2) is the kinase primarily responsible for phosphorylating the N-terminus of human HP1α. Pull-down assays using in vitro-reconstituted nucleosomes showed that unmodified HP1α bound H3K9-methylated and H3K9-unmethylated nucleosomes with comparable affinity, whereas CK2-phosphorylated HP1α showed a high specificity for H3K9me3-modified nucleosomes. Electrophoretic mobility shift assays showed that CK2-mediated phosphorylation diminished HP1α's intrinsic DNA binding, which contributed to its H3K9me-independent nucleosome binding. CK2-mediated phosphorylation had a similar effect on the nucleosome-binding specificity of fly HP1a and S. pombe Swi6. These results suggested that HP1 phosphorylation has an evolutionarily conserved role in HP1's recognition of H3K9me-marked nucleosomes.
authors.affiliation: Graduate School of Natural Sciences, Nagoya City University, Nagoya 467-8501, Japan.
authors.forename: Gohei
authors.initials: G
authors.lastname: Nishibuchi
authors.affiliation: Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Machida
authors.affiliation: Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Akihisa
authors.initials: A
authors.lastname: Osakabe
authors.affiliation: Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Hiromu
authors.initials: H
authors.lastname: Murakoshi
authors.affiliation: Laboratory of Chromatin Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
authors.forename: Kyoko
authors.initials: K
authors.lastname: Hiragami-Hamada
authors.affiliation: Proteomics Support Unit, RIKEN Center for Developmental Biology, Kobe 650-0047, Japan.
authors.forename: Reiko
authors.initials: R
authors.lastname: Nakagawa
authors.affiliation: Laboratory of Chromatin Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Fischle
authors.affiliation: Graduate School of Medical Life Science, Yokohama City University, 1-7-29, Suehiro-cho, Tsurumi-ku, Yokohama 230-0045, Japan.
authors.forename: Yoshifumi
authors.initials: Y
authors.lastname: Nishimura
authors.affiliation: Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kurumizaka
authors.affiliation: Graduate School of Natural Sciences, Nagoya City University, Nagoya 467-8501, Japan.
authors.forename: Hideaki
authors.initials: H
authors.lastname: Tagami
authors.affiliation: Graduate School of Natural Sciences, Nagoya City University, Nagoya 467-8501, Japan jnakayam@nsc.nagoya-cu.ac.jp.
authors.forename: Jun-ichi
authors.initials: J
authors.lastname: Nakayama
mesh_terms: D047390:Casein Kinase II / Q000378:metabolism; D002460:Cell Line; D000090266:Chromobox Protein Homolog 5; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D006801:Humans; D009707:Nucleosomes / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D012694:Serine / Q000378:metabolism
chemical_list: C000715594:CBX5 protein, human; D002868:Chromosomal Proteins, Non-Histone; D006657:Histones; D009707:Nucleosomes; D000090266:Chromobox Protein Homolog 5; D012694:Serine; D004247:DNA; D047390:Casein Kinase II