title: Human Orc2 localizes to centrosomes, centromeres and heterochromatin during chromosome inheritance.
abstract: The initiation of DNA replication in S phase requires the prior assembly of an origin recognition complex (ORC)-dependent pre-replicative complex on chromatin during G1 phase of the cell division cycle. In human cells, the Orc2 subunit localized to the nucleus as expected, but it also localized to centrosomes throughout the entire cell cycle. Furthermore, Orc2 was tightly bound to heterochromatin and heterochromatin protein 1alpha (HP1alpha) and HP1beta in G1 and early S phase, but during late S, G2 and M phases tight chromatin association was restricted to centromeres. Depletion of Orc2 by siRNA caused multiple phenotypes. A population of cells showed an S-phase defect with little proliferating cell nuclear antigen (PCNA) on chromatin, although MCM proteins remained. Orc2 depletion also disrupted HP1 localization, but not histone-H3-lysine-9 methylation at prominent heterochromatic foci. Another subset of Orc2-depleted cells containing replicated DNA arrested with abnormally condensed chromosomes, failed chromosome congression and multiple centrosomes. These results implicate Orc2 protein in chromosome duplication, chromosome structure and centrosome copy number control, suggesting that it coordinates all stages of the chromosome inheritance cycle.
authors.affiliation: Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11724, USA.
authors.forename: Supriya G
authors.initials: SG
authors.lastname: Prasanth
authors.forename: Kannanganattu V
authors.initials: KV
authors.lastname: Prasanth
authors.forename: Khalid
authors.initials: K
authors.lastname: Siddiqui
authors.forename: David L
authors.initials: DL
authors.lastname: Spector
authors.forename: Bruce
authors.initials: B
authors.lastname: Stillman
mesh_terms: D017209:Apoptosis; D002453:Cell Cycle; D002503:Centromere / Q000378:metabolism*; D018385:Centrosome / Q000378:metabolism*; D002843:Chromatin / Q000378:metabolism; D000090266:Chromobox Protein Homolog 5; D002868:Chromosomal Proteins, Non-Histone / Q000378:metabolism; D002875:Chromosomes / Q000235:genetics*; D003001:Cloning, Molecular; D004261:DNA Replication; D004268:DNA-Binding Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006570:Heterochromatin / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008677:Metaphase; D051738:Origin Recognition Complex; D018809:Proliferating Cell Nuclear Antigen / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology
chemical_list: C000715593:CBX1 protein, human; C000715594:CBX5 protein, human; D002843:Chromatin; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D006570:Heterochromatin; C497224:ORC2 protein, human; D051738:Origin Recognition Complex; D018809:Proliferating Cell Nuclear Antigen; D034741:RNA, Small Interfering; D000090266:Chromobox Protein Homolog 5