title: The CCAN recruits CENP-A to the centromere and forms the structural core for kinetochore assembly.
abstract: CENP-A acts as an important epigenetic marker for kinetochore specification. However, the mechanisms by which CENP-A is incorporated into centromeres and the structural basis for kinetochore formation downstream of CENP-A remain unclear. Here, we used a unique chromosome-engineering system in which kinetochore proteins are targeted to a noncentromeric site after the endogenous centromere is conditionally removed. Using this system, we created two distinct types of engineered kinetochores, both of which were stably maintained in chicken DT40 cells. Ectopic targeting of full-length HJURP, CENP-C, CENP-I, or the CENP-C C terminus generated engineered kinetochores containing major kinetochore components, including CENP-A. In contrast, ectopic targeting of the CENP-T or CENP-C N terminus generated functional kinetochores that recruit the microtubule-binding Ndc80 complex and chromosome passenger complex (CPC), but lack CENP-A and most constitutive centromere-associated network (CCAN) proteins. Based on the analysis of these different engineered kinetochores, we conclude that the CCAN has two distinct roles: recruiting CENP-A to establish the kinetochore and serving as a structural core to directly recruit kinetochore proteins.
authors.affiliation: Department of Molecular Genetics, National Institute of Genetics, Mishima, Shizuoka 411-8540, Japan.
authors.forename: Tetsuya
authors.initials: T
authors.lastname: Hori
authors.forename: Wei-Hao
authors.initials: WH
authors.lastname: Shang
authors.forename: Kozo
authors.initials: K
authors.lastname: Takeuchi
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Fukagawa
mesh_terms: D000818:Animals; D001324:Autoantigens / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D000076247:Centromere Protein A; D002645:Chickens; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D018386:Kinetochores / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary
chemical_list: D001324:Autoantigens; D000076247:Centromere Protein A; D002868:Chromosomal Proteins, Non-Histone; D004268:DNA-Binding Proteins; D046912:Multiprotein Complexes; C060905:centromere protein C