A putative protein complex consisting of Ctf19, Mcm21, and Okp1 represents a missing link in the budding yeast kinetochore

Ortiz, Jennifer; Stemmann, Olaf; Rank, Simone; Lechner, Johannes

doi:10.1101/gad.13.9.1140

Cited by 195 publications

(276 citation statements)

References 53 publications

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“…Despite the variation at the inactive cCEN, all of the proteins tested localized to the active cCEN at levels comparable with the eCEN. Although we detected full occupancy of Cse4 at the inactive cCEN, it is not clear why we detected only partial localization of Ndc10, because Ndc10 is required for the localization of Cse4 (Ortiz et al, 1999). One possibility is that Cse4 and Ndc10 transiently bind to the inactive cCEN, but Cse4 is easier to detect because it has a stronger binding affinity.…”

Section: A Conditional Centromere Provides An Assay To Study Yeast Kimentioning

confidence: 55%

De Novo Kinetochore Assembly Requires the Centromeric Histone H3 Variant

Collins

Castillo

Tatsutani

et al. 2005

MBoC

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Kinetochores mediate chromosome attachment to the mitotic spindle to ensure accurate chromosome segregation. Budding yeast is an excellent organism for kinetochore assembly studies because it has a simple defined centromere sequence responsible for the localization of >65 proteins. In addition, yeast is the only organism where a conditional centromere is available to allow studies of de novo kinetochore assembly. Using a conditional centromere, we found that yeast kinetochore assembly is not temporally restricted and can occur in both G 1 phase and prometaphase. We performed the first investigation of kinetochore assembly in the absence of the centromeric histone H3 variant Cse4 and found that all proteins tested depend on Cse4 to localize. Consistent with this observation, Cse4-depleted cells had severe chromosome segregation defects. We therefore propose that yeast kinetochore assembly requires both centromeric DNA specificity and centromeric chromatin. INTRODUCTIONAccurate chromosome segregation in mitosis and meiosis is essential for the maintenance of genomic stability. Chromosomes attach to the mitotic spindle at the kinetochore, the protein complex that assembles onto centromeric DNA. Although kinetochore function is conserved, the underlying centromeric DNA is highly variable. Budding yeast contain a 125-base pair sequence-specific centromere that is sufficient for kinetochore formation (Fitzgerald-Hayes et al., 1982). In contrast, centromeres in multicellular eukaryotes are composed of megabases of highly repetitive DNA that lack sequence specificity (for review, see Sullivan et al., 2001). In these organisms, kinetochore assembly seems to be propagated by unidentified epigenetic component(s) (Karpen and Allshire, 1997;Sullivan et al., 2001).The best-characterized kinetochore is in budding yeast where Ͼ65 components have been identified that constitutively localize to the kinetochore (for reviews, see Biggins and Walczak, 2003;McAinsh et al., 2003). Most of the yeast kinetochore proteins are found in biochemically distinct complexes known as the CBF3, CTF19/COMA, MTW1, NDC80, and DAM1 complexes that seem to assemble on a single centromeric nucleosome (Meluh et al., 1998). Although the exact architecture of the kinetochore is not known, dependency relationships subdivide the kinetochore into inner, central, and outer domains. The inner kinetochore contains the CBF3 complex (Ndc10, Cep3, Skp1, and Ctf13) as well as the DNA binding proteins Mif2, Cbf1, and the yeast centromeric histone H3 variant (CenH3) Cse4. CBF3 binds directly to the centromeric DNA and is thought to nucleate kinetochore assembly because all kinetochore proteins require it for localization (Russell et al., 1999;Goshima and Yanagida, 2000;He et al., 2001;Janke et al., 2001Janke et al., , 2002. The central kinetochore contains the MTW1 (Mtw1, Dsn1, Nnf1, and Nsl1) and CTF19/COMA (Ctf19, Mcm16, Mcm19, Mcm21, Mcm22, Ctf3, Chl4, Okp1, Ame1, Iml3, Nkp1, and Nkp2) complexes. CTF19/COMA can be further divided into two subcomplexes, with Ame1...

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Section: A Conditional Centromere Provides An Assay To Study Yeast Kimentioning

confidence: 55%

De Novo Kinetochore Assembly Requires the Centromeric Histone H3 Variant

Collins

Castillo

Tatsutani

et al. 2005

MBoC

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“…This model predicts that the youngest satellites found in centromeric regions would represent the extant centromere, a prediction that has recently been confirmed by high-resolution mapping of the human X centromere (36). Under this model, fungal CenH3s are not expected to evolve adaptively both because they are not subject to meiotic asymmetries and because they rely on other factors for their centromeric localization (8,37).…”

Section: Discussionmentioning

confidence: 86%

Recurrent evolution of DNA-binding motifs in the Drosophila centromeric histone

Malik

Vermaak

Henikoff

2002

Proc. Natl. Acad. Sci. U.S.A.

113

110

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All eukaryotes contain centromere-specific histone H3 variants (CenH3s), which replace H3 in centromeric chromatin. We have previously documented the adaptive evolution of the Drosophila CenH3 (Cid) in comparisons of Drosophila melanogaster and Drosophila simulans, a divergence of Ϸ2.5 million years. We have proposed that rapidly changing centromeric DNA may be driving CenH3's altered DNA-binding specificity. Here, we compare Cid sequences from a phylogenetically broader group of Drosophila species to suggest that Cid has been evolving adaptively for at least 25 million years. Our analysis also reveals conserved blocks not only in the histone-fold domain but also in the N-terminal tail. In several lineages, the N-terminal tail of Cid is characterized by subgroup-specific oligopeptide expansions. These expansions resemble minor groove DNA binding motifs found in various histone tails. Remarkably, similar oligopeptides are also found in N-terminal tails of human and mouse CenH3 (Cenp-A). The recurrent evolution of these motifs in CenH3 suggests a packaging function for the N-terminal tail, which results in a unique chromatin organization at the primary constriction, the cytological marker of centromeres.

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“…7E). At centromeres, multiple factors would favor incorporation of a single stable tetramer, including exclusion of H3 octamers by Cbf1 (15,21), the 90% AT-richness of CDEII, which resists assembly of Cse4 octamers (14,40), and the recruitment of Cse4 by the adjacent CBF3 complex (41). In multicellular eukaryotes, heterochromatin condensation would help to stabilize CenH3 tetramers by preventing nucleosome turnover (42).…”

Section: Discussionmentioning

confidence: 99%

Tripartite organization of centromeric chromatin in budding yeast

Krassovsky

Henikoff

2011

Proc. Natl. Acad. Sci. U.S.A.

166

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The centromere is the genetic locus that organizes the proteinaceous kinetochore and is responsible for attachment of the chromosome to the spindle at mitosis and meiosis. In most eukaryotes, the centromere consists of highly repetitive DNA sequences that are occupied by nucleosomes containing the CenH3 histone variant, whereas in budding yeast, a ∼120-bp centromere DNA element (CDE) that is sufficient for centromere function is occupied by a single right-handed histone variant CenH3 (Cse4) nucleosome. However, these in vivo observations are inconsistent with in vitro evidence for left-handed octameric CenH3 nucleosomes. To help resolve these inconsistencies, we characterized yeast centromeric chromatin at single base-pair resolution. Intact particles containing both Cse4 and H2A are precisely protected from micrococcal nuclease over the entire CDE of all 16 yeast centromeres in both solubilized chromatin and the insoluble kinetochore. Small DNAbinding proteins protect CDEI and CDEIII and delimit the centromeric nucleosome to the ∼80-bp CDEII, only enough for a single DNA wrap. As expected for a tripartite organization of centromeric chromatin, loss of Cbf1 protein, which binds to CDEI, both reduces the size of the centromere-protected region and shifts its location toward CDEIII. Surprisingly, Cse4 overproduction caused genome-wide misincorporation of nonfunctional CenH3-containing nucleosomes that protect ∼135 base pairs and are preferentially enriched at sites of high nucleosome turnover. Our detection of two forms of CenH3 nucleosomes in the yeast genome, a singly wrapped particle at the functional centromere and octamer-sized particles on chromosome arms, reconcile seemingly conflicting in vivo and in vitro observations. Saccharomyces cerevisiae | chromatin immunoprecipitation | chromosome segregation

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A putative protein complex consisting of Ctf19, Mcm21, and Okp1 represents a missing link in the budding yeast kinetochore

Cited by 195 publications

References 53 publications

De Novo Kinetochore Assembly Requires the Centromeric Histone H3 Variant

De Novo Kinetochore Assembly Requires the Centromeric Histone H3 Variant

Recurrent evolution of DNA-binding motifs in the Drosophila centromeric histone

Tripartite organization of centromeric chromatin in budding yeast

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