Characterization of a CENP-C homolog in Arabidopsis thaliana

Ogura, Yutaka; Shibata, Fumiaki; Sato, Hiroshi; Murata, Minoru

doi:10.1266/ggs.79.139

Cited by 46 publications

(45 citation statements)

References 28 publications

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“…We generated a polyclonal antibody to an 89-amino acid fragment in the amino terminus of XlCENP-C ( Figure 1B). We stained cultured X. laevis S3 cells with this antibody and observed that XlCENP-C localizes to centromeres in both interphase and mitosis ( Figure 1C), consistent with the constitutive cell cycle localization of CENP-C in other eukaryotes (Saitoh et al, 1992;Knehr et al, 1996;Kalitsis et al, 1998b;Dawe et al, 1999;Ogura et al, 2004;Heeger et al, 2005).…”

Section: Characterization Of X Laevis Cenp-csupporting

confidence: 76%

Dissection of CENP-C–directed Centromere and Kinetochore Assembly

Milks

Moree

Straight

2009

MBoC

118

145

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Eukaryotic cells ensure accurate chromosome segregation in mitosis by assembling a microtubule-binding site on each chromosome called the kinetochore that attaches to the mitotic spindle. The kinetochore is assembled specifically during mitosis on a specialized region of each chromosome called the centromere, which is constitutively bound by >15 centromere-specific proteins. These proteins, including centromere proteins A and C (CENP-A and -C), are essential for kinetochore assembly and proper chromosome segregation. How the centromere is assembled and how the centromere promotes mitotic kinetochore formation are poorly understood. We have used Xenopus egg extracts as an in vitro system to study the role of CENP-C in centromere and kinetochore assembly. We show that, unlike the histone variant CENP-A, CENP-C is not maintained at centromeres through spermatogenesis but is assembled at the sperm centromere from the egg cytoplasm. Immunodepletion of CENP-C from metaphase egg extract prevents kinetochore formation on sperm chromatin, and depleted extracts can be complemented with in vitro-translated CENP-C. Using this complementation assay, we have identified CENP-C mutants that localized to centromeres but failed to support kinetochore assembly. We find that the amino terminus of CENP-C promotes kinetochore assembly by ensuring proper targeting of the Mis12/MIND complex and CENP-K. INTRODUCTIONCell proliferation requires the equal segregation of the genome between daughter cells during division. Eukaryotic chromosome segregation is accomplished by attaching each replicated chromosome to opposite poles of the mitotic spindle so that chromosomes are equally distributed in anaphase. The interaction site between chromosomes and the mitotic spindle is the kinetochore, a multiprotein complex that assembles in mitosis to bind spindle microtubules. Kinetochores also monitor improper attachment to the spindle through the mitotic checkpoint and directly couple the chromosomes to spindle forces during anaphase segregation (Inoue and Salmon, 1995;Nicklas, 1997;Rieder and Salmon, 1998;Cleveland et al., 2003).Although kinetochores only assemble in mitosis, the site of kinetochore assembly, the centromere, persists throughout the cell cycle. The first identification of centromerespecific binding proteins came from analysis of scleroderma patient sera and led to the identification of three proteins that constitutively localize to centromeres (Moroi et al., 1980;Earnshaw and Rothfield, 1985). Two of these proteins, CENP-A and -C, are essential for centromere formation in all eukaryotes. CENP-A is a histone H3 variant that replaces histone H3 in centromeric nucleosomes and is required for kinetochore formation (Palmer et al., 1991;Cleveland et al., 2003;Carroll and Straight, 2006). CENP-A chromatin directs the assembly of at least 19 additional centromere proteins (CENP-C, -H, -I, K-U, and -W and the Mis12/MIND proteins: Mis12, Nsl1, Nnf1, and Dsn1) that are required for formation of the mitotic kinetochore Hori et al., 2008).Inhibi...

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Section: Characterization Of X Laevis Cenp-csupporting

confidence: 76%

Dissection of CENP-C–directed Centromere and Kinetochore Assembly

Milks

Moree

Straight

2009

MBoC

118

145

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“…This sort of close relationship between the localization of proteins and centromere activities has hitherto not been clearly demonstrated in plants. However, since the centromeric proteins homologous to human CENP-A and -C have been found or predicted in maize (Dawe et al, 1999;Zhong et al, 2002) and A. thaliana (Yu et al, 2000;Talbert et al, 2002;Ogura et al, 2004), it should be also possible to detect centromere activity in plants by localization of these homologues. Our sequential detection of HTR12 (a centromeric histone H3 variant of A. thaliana), AtCENP-C (Arabidopsis CENP-C homologue) and the 180 bp repeats revealed that these two centromeric proteins colocalize with the 180 bp repeats.…”

Section: Discussionmentioning

confidence: 99%

“…In addition, the anti-AtCENP-C antibodies were purified from a rabbit immunized with a synthetic peptide (SKVKSFVSDYKKLVD) corresponding to the C-terminal amino acids of A. thaliana CENP-C homologue (accession nos. AB128986 and AB128987) (Ogura et al, 2004). The antibodies were diluted 1:200, 1:2000 and 1:200, respectively for application.…”

Section: Fish and Immunofluorescence Labelingmentioning

confidence: 99%

Differential localization of the centromere-specific proteins in the major centromeric satellite ofArabidopsis thaliana

Shibata

Murata

2004

Journal of Cell Science

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The 180 bp family of tandem repetitive sequences, which constitutes the major centromeric satellite in Arabidopsis thaliana, is thought to play important roles in kinetochore assembly. To assess the centromere activities of the 180 bp repeats, we performed indirect fluorescence immunolabeling with antibodies against phosphorylated histone H3 at Ser10, HTR12 (Arabidopsis centromeric histone H3 variant) and AtCENP-C (Arabidopsis CENP-C homologue) for the A. thaliana cell cultures. The immunosignals from all three antibodies appeared on all sites of the 180 bp repeats detected by fluorescence in situ hybridization. However, some of the 180 bp repeat clusters, particularly those that were long or stretched at interphase, were not fully covered with the signals from anti-HTR12 or AtCENP-C. Chromatin fiber immunolabeling clearly revealed that the centromeric proteins examined in this study, localize only at the knobs on the extended chromatin fibers, which form a limited part of the 180 bp clusters. Furthermore, outer HTR12 and inner phosphohistone H3 (Ser10) localization at the kinetochores of metaphase chromosomes suggests that two kinds of histone H3 (a centromere variant and a phosphorylated form) might be linked to different roles in centromere functionality; the former for spindle-fiber attachment, and the latter for chromatid cohesion.

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“…In particular, investigations concerning centromeric proteins and their functions have been intensely pursued in yeasts and humans (Amor et al, 2004). Recently, centromere analyses with centromeric proteins have been pursued in plants (Dawe et al, 1999;Talbert et al, 2002Talbert et al, , 2004Zhong et al, 2002;Nagaki et al, 2003Nagaki et al, , 2004Ogura et al, 2004;Shibata and Murata, 2004). These investigations revealed that centromeric proteins are conserved among plants, whereas the sequences of centromeric DNAs are variable (Murata, 2002;Jiang et al, 2003;Talbert et al, 2004).…”

Section: Introductionmentioning

confidence: 99%

Visualization of Diffuse Centromeres with Centromere-Specific Histone H3 in the Holocentric PlantLuzula nivea

2005

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Although holocentric species are scattered throughout the plant and animal kingdoms, only holocentric chromosomes of the nematode worm Caenorhabditis elegans have been analyzed with centromeric protein markers. In an effort to determine the holocentric structure in plants, we investigated the snowy woodrush Luzula nivea. From the young roots, a cDNA encoding a putative centromere-specific histone H3 (LnCENH3) was successfully isolated based on sequence similarity among plant CENH3s. The deduced amino acid sequence was then used to raise an anti-LnCENH3 antibody. Immunostaining clearly revealed the diffuse centromere-like structure that appears in the linear shape at prophase to telophase. Furthermore, it was shown that the amount of LnCENH3 decreased significantly at interphase. The polar side positioning on each chromatid at metaphase to anaphase also confirmed that LnCENH3 represents one of the centromere-specific proteins in L. nivea. These data from L. nivea are compared with those from C. elegans, and common features of holocentric chromosomes are discussed.

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Characterization of a CENP-C homolog in Arabidopsis thaliana

Cited by 46 publications

References 28 publications

Dissection of CENP-C–directed Centromere and Kinetochore Assembly

Dissection of CENP-C–directed Centromere and Kinetochore Assembly

Differential localization of the centromere-specific proteins in the major centromeric satellite ofArabidopsis thaliana

Visualization of Diffuse Centromeres with Centromere-Specific Histone H3 in the Holocentric PlantLuzula nivea

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