title: Cdt1-binding protein GRWD1 is a novel histone-binding protein that facilitates MCM loading through its influence on chromatin architecture.
abstract: Efficient pre-replication complex (pre-RC) formation on chromatin templates is crucial for the maintenance of genome integrity. However, the regulation of chromatin dynamics during this process has remained elusive. We found that a conserved protein, GRWD1 (glutamate-rich WD40 repeat containing 1), binds to two representative replication origins specifically during G1 phase in a CDC6- and Cdt1-dependent manner, and that depletion of GRWD1 reduces loading of MCM but not CDC6 and Cdt1. Furthermore, chromatin immunoprecipitation coupled with high-throughput sequencing (Seq) revealed significant genome-wide co-localization of GRWD1 with CDC6. We found that GRWD1 has histone-binding activity. To investigate the effect of GRWD1 on chromatin architecture, we used formaldehyde-assisted isolation of regulatory elements (FAIRE)-seq or FAIRE-quantitative PCR analyses, and the results suggest that GRWD1 regulates chromatin openness at specific chromatin locations. Taken together, these findings suggest that GRWD1 may be a novel histone-binding protein that regulates chromatin dynamics and MCM loading at replication origins.
authors.affiliation: Department of Cellular Biochemistry, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Nozomi
authors.initials: N
authors.lastname: Sugimoto
authors.affiliation: Faculty of Medicine, Division of Epigenetics, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Kazumitsu
authors.initials: K
authors.lastname: Maehara
authors.affiliation: Department of Cellular Biochemistry, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Kazumasa
authors.initials: K
authors.lastname: Yoshida
authors.affiliation: Department of Cellular Biochemistry, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Shuhei
authors.initials: S
authors.lastname: Yasukouchi
authors.affiliation: Division of Virology, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan.
authors.forename: Satoko
authors.initials: S
authors.lastname: Osano
authors.affiliation: Department of Cellular Biochemistry, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Shinya
authors.initials: S
authors.lastname: Watanabe
authors.affiliation: Department of Cellular Biochemistry, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Aizawa
authors.affiliation: Division of Virology, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Yugawa
authors.affiliation: Division of Virology, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan.
authors.forename: Tohru
authors.initials: T
authors.lastname: Kiyono
authors.affiliation: Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kurumizaka
authors.affiliation: Faculty of Medicine, Division of Epigenetics, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan.
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Ohkawa
authors.affiliation: Department of Cellular Biochemistry, Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan mfujita@phar.kyushu-u.ac.jp.
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Fujita
mesh_terms: D002352:Carrier Proteins / Q000032:analysis / Q000378:metabolism* / Q000502:physiology; D002453:Cell Cycle; D018797:Cell Cycle Proteins / Q000378:metabolism; D002467:Cell Nucleus / Q000737:chemistry; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D004261:DNA Replication; D057809:HEK293 Cells; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism*; D006801:Humans; D034921:Lamin Type B / Q000235:genetics; D064148:Minichromosome Maintenance Complex Component 4 / Q000235:genetics; D064110:Minichromosome Maintenance Proteins / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism; D018741:Replication Origin
chemical_list: C103706:CDC6 protein, human; C467199:CDT1 protein, human; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002843:Chromatin; C504316:GRWD1 protein, human; D006657:Histones; D034921:Lamin Type B; D009687:Nuclear Proteins; C099243:MCM4 protein, human; D064148:Minichromosome Maintenance Complex Component 4; D064110:Minichromosome Maintenance Proteins