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J Cell Sci. 2015 Dec 15;128(24):4572-87. doi: 10.1242/jcs.180786. Epub 2015 Nov 2.

CENP-C and CENP-I are key connecting factors for kinetochore and CENP-A assembly.

Author information

1
Laboratory of Cell Engineering, Department of Frontier Research, Kazusa DNA Research Institute, 2-6-7 Kazusa-Kamatari, Kisarazu, Chiba 292-0818, Japan Division of Biological Science, Graduate School of Science, Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi 464-8602, Japan.
2
Laboratory of Cell Engineering, Department of Frontier Research, Kazusa DNA Research Institute, 2-6-7 Kazusa-Kamatari, Kisarazu, Chiba 292-0818, Japan.
3
Wellcome Trust Centre for Cell Biology University of Edinburgh, King's Buildings, Mayfield Road, Edinburgh EH9 3JR, UK.
4
Public Relations Team, Kazusa DNA Research Institute, 2-6-7 Kazusa-Kamatari, Kisarazu, Chiba 292-0818, Japan.
5
Department of Biological Sciences, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama 226-8501, Japan.
6
Developmental Therapeutic Branch, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
7
Laboratory of Cell Engineering, Department of Frontier Research, Kazusa DNA Research Institute, 2-6-7 Kazusa-Kamatari, Kisarazu, Chiba 292-0818, Japan masumoto@kazusa.or.jp.

Abstract

Although it is generally accepted that chromatin containing the histone H3 variant CENP-A is an epigenetic mark maintaining centromere identity, the pathways leading to the formation and maintenance of centromere chromatin remain unclear. We previously generated human artificial chromosomes (HACs) whose centromeres contain a synthetic alpha-satellite (alphoid) DNA array containing the tetracycline operator (alphoid(tetO)). We also obtained cell lines bearing the alphoid(tetO) array at ectopic integration sites on chromosomal arms. Here, we have examined the regulation of CENP-A assembly at centromeres as well as de novo assembly on the ectopic arrays by tethering tetracycline repressor (tetR) fusions of substantial centromeric factors and chromatin modifiers. This analysis revealed four classes of factors that influence CENP-A assembly. Interestingly, many kinetochore structural components induced de novo CENP-A assembly at the ectopic site. We showed that these components work by recruiting CENP-C and subsequently recruiting M18BP1. Furthermore, we found that CENP-I can also recruit M18BP1 and, as a consequence, enhances M18BP1 assembly on centromeres in the downstream of CENP-C. Thus, we suggest that CENP-C and CENP-I are key factors connecting kinetochore to CENP-A assembly.

KEYWORDS:

CENP-A; CENP-C; CENP-I; Centromere; HAC; Human artificial chromosome; Kinetochore

PMID:
26527398
PMCID:
PMC4696500
DOI:
10.1242/jcs.180786
[Indexed for MEDLINE]
Free PMC Article

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