title: Octameric CENP-A nucleosomes are present at human centromeres throughout the cell cycle.
abstract: The presence of a single centromere on each chromosome that signals formation of a mitotic kinetochore is central to accurate chromosome segregation. The histone H3 variant centromere protein-A (CENP-A) is critical for centromere identity and function; CENP-A chromatin acts as an epigenetic mark to direct both centromere and kinetochore assembly. Interpreting the centromere epigenetic mark ensures propagation of a single centromere per chromosome to maintain ploidy. Thus, understanding the nature of CENP-A chromatin is crucial for all cell divisions. However, there are ongoing debates over the fundamental composition of centromeric chromatin. Here we show that natively assembled human CENP-A nucleosomes are octameric throughout the cell cycle. Using total internal reflection fluorescence (TIRF)-coupled photobleaching-assisted copy-number counting of single nucleosomes obtained from cultured cells, we find that the majority of CENP-A nucleosomes contain CENP-A dimers. In addition, we detect the presence of H2B and H4 in these nucleosomes. Surprisingly, CENP-A associated with the chaperone HJURP can exist as either monomer or dimer, indicating possible assembly intermediates. Thus, our findings indicate that octameric CENP-A nucleosomes mark the centromeric region to ensure proper epigenetic inheritance and kinetochore assembly.
authors.affiliation: Institute for Research in Immunology and Cancer (IRIC), Faculty of Medicine, University of Montreal, Montreal, QC H3T 1J4, Canada.
authors.forename: Abbas
authors.initials: A
authors.lastname: Padeganeh
authors.forename: Joël
authors.initials: J
authors.lastname: Ryan
authors.forename: Jacques
authors.initials: J
authors.lastname: Boisvert
authors.forename: Anne-Marie
authors.initials: AM
authors.lastname: Ladouceur
authors.forename: Jonas F
authors.initials: JF
authors.lastname: Dorn
authors.forename: Paul S
authors.initials: PS
authors.lastname: Maddox
mesh_terms: D001324:Autoantigens / Q000235:genetics / Q000378:metabolism*; D002453:Cell Cycle; D002503:Centromere / Q000378:metabolism*; D000076247:Centromere Protein A; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D044127:Epigenesis, Genetic; D018628:Gene Dosage; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000737:chemistry / Q000378:metabolism; D009707:Nucleosomes / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization
chemical_list: D001324:Autoantigens; C000619700:CENPA protein, human; D000076247:Centromere Protein A; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; C524371:HJURP protein, human; D009707:Nucleosomes