Human CENP-H multimers colocalize with CENP-A and CENP-C at active centromere-kinetochore complexes

• Published in: Human molecular genetics Vol. 9; no. 19; pp. 2919 - 2926
• Main Authors: SUGATA, Naoko, SHULAN LI, EARNSHAW, William C, YEN, Tim J, YODA, Kinya, MASUMOTO, Hiroshi, MUNEKATA, Eisuke, WARBURTON, Peter E, TODOKORO, Kazuo
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 22.11.2000; Oxford Publishing Limited (England)
• Subjects: Amino Acid Sequence; Autoantigens; Biological and medical sciences; CENP-A protein; CENP-C protein; CENP-H protein; Centromere - metabolism; Centromere Protein A; Chromatin. Chromosome; Chromosomal Proteins, Non-Histone - chemistry; Chromosomal Proteins, Non-Histone - genetics; Chromosomal Proteins, Non-Histone - metabolism; Cloning, Molecular; Fluorescent Antibody Technique; Fundamental and applied biological sciences. Psychology; HeLa Cells; Humans; In Situ Hybridization, Fluorescence; kinetochores; Kinetochores - metabolism; Macromolecular Substances; MCAK protein; Microscopy, Fluorescence; Mitosis; Molecular and cellular biology; Molecular genetics; Molecular Sequence Data; Protein Binding; Recombinant Fusion Proteins - metabolism; Sequence Alignment; Human; Mitosis; Rodentia; Chromosome; Homology; Hela cell line; Vertebrata; Cell line; Mammalia; Kinetochore; Centromere; Mouse; DNA; Comparative study; Nuclear protein; Aminoacid sequence
• ISSN: 0964-6906; 1460-2083; 1460-2083
• DOI: 10.1093/hmg/9.19.2919

Centromere and kinetochore proteins have a pivotal role in centromere structure, kinetochore formation and sister chromatid separation. However, the molecular architecture and the precise dynamic function of the centromere-kinetochore complex during mitosis remain poorly understood. Here we report the isolation and characterization of human CENP-H. Confocal microscopic analyses of HeLa cells with anti-human CENP-H-specific antibody demonstrated that CENP-H colocalizes with inner kinetochore plate proteins CENP-A and CENP-C in both interphase and metaphase. CENP-H was present outside centromeric heterochromatin, where CENP-B is localized, and inside the kinetochore corona, where CENP-E is localized during prometaphase. Furthermore, CENP-H was detected at neocentromeres, but not at inactive centromeres in stable dicentric chromosomes. In vitro binding assays of human CENP-H with centromere-kinetochore proteins suggest that the CENP-H binds to itself and MCAK, but not to CENP-A, CENP-B or CENP-C. CENP-H multimers were observed in cells in which both FLAG-tagged CENP-H and hemagglutinin-tagged CENP-H were expressed. These results suggest that CENP-H multimers localize constitutively to the inner kinetochore plate and play an important fundamental role in organization and function of the active human centromere-kinetochore complex.