Tetrameric Structure of Centromeric Nucleosomes in Interphase Drosophila Cells

Dalal, Yamini; Wang, Hongda; Lindsay, Stuart; Henikoff, Steven

doi:10.1371/journal.pbio.0050218

Cited by 218 publications

(265 citation statements)

References 53 publications

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“…This finding confirms that DNA wraps only once around a Cse4-containing core (33), as implied by our previous study showing that Cse4 nucleosomes induce positive supercoiling at functional centromeres in vivo (6). Our findings are consistent with the observation that singly wrapped CenH3 particles also occupy Drosophila and human centromeres (8,9), suggesting that this organization is a universal feature of centromeric nucleosomes. Singly wrapped tetrameric nucleosomes are universal for archaea (34), as if centromeres have retained the ancestral nucleosomal organization.…”

Section: Discussionsupporting

confidence: 92%

“…Most important is the histone variant, CenH3 (CENP-A in mammals and Cse4 in yeast), which replaces histone H3 in centromeric nucleosomes and is essential for recruitment of the other structural components of the kinetochore (5). It has been previously shown that the Cse4 nucleosome wraps DNA in a right-handed orientation (6,7), consistent with in vivo observations of heterotypic tetrameric nucleosomes in Drosophila (8) and humans (9). However, several studies have shown that CenH3 nucleosomes are left-handed octamers in vitro (10)(11)(12)(13)(14).…”

supporting

confidence: 77%

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Tripartite organization of centromeric chromatin in budding yeast

Krassovsky

Henikoff

2011

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

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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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Section: Discussionsupporting

confidence: 92%

supporting

confidence: 77%

Tripartite organization of centromeric chromatin in budding yeast

Krassovsky

Henikoff

2011

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

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166

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“…Experimental evidence in favor of a histone tetramer [hemisome; H2A/H2B/H4/(CenH3 = Cse4)] or a histone octamer (H2A/H2B/H4/CenH3/CenH3/H4/H2B/ H2A) has been reported (22,25,(28)(29)(30). The specter of an unusual hexameric nucleosome core, bereft of H2A and H2B, but including the Cse4-associated nonhistone protein Scm3 [H4/ Cse4/Cse4/H4-(Scm3)2], has also been raised (31).…”

Section: Discussionmentioning

confidence: 99%

Histone H3-variant Cse4-induced positive DNA supercoiling in the yeast plasmid has implications for a plasmid origin of a chromosome centromere

Huang

Chang

Cui

et al. 2011

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

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The Saccharomyces cerevisiae 2-μm plasmid is a multicopy selfish genome that resides in the nucleus. The genetic organization of the plasmid is optimized for stable, high-copy propagation in hostcell populations. The plasmid's partitioning system poaches host factors, including the centromere-specific histone H3-variant Cse4 and the cohesin complex, enabling replicated plasmid copies to segregate equally in a chromosome-coupled fashion. We have characterized the in vivo chromatin topology of the plasmid partitioning locus STB in its Cse4-associated and Cse4-nonassociated states. We find that the occupancy of Cse4 at STB induces positive DNA supercoiling, with a linking difference (ΔLk) contribution estimated between +1 and +2 units. One plausible explanation for this contrary topology is the presence of a specialized Cse4-containing nucleosome with a right-handed DNA writhe at a functional STB, contrasted by a standard histone H3-containing nucleosome with a left-handed DNA writhe at a nonfunctional STB. The similarities between STB and centromere in their nucleosome signature and DNA topology would be consistent with the potential origin of the unusual point centromere of budding yeast chromosomes from the partitioning locus of an ancestral plasmid.CEN evolution | nucleosome topology | reversome T he 2-μm plasmid of Saccharomyces cerevisiae resides in the nucleus at 40 to 60 copies per cell, and propagates itself with nearly the same stability as chromosomes (1, 2). The plasmid is a benign selfish DNA element that seems to provide no advantage to its host, but poses, at its normal copy number, no serious disadvantage either. In haploid cells the plasmid is organized into a cluster of three to five foci, and segregates as a cluster (3). This effective reduction in copy number necessitates an active partitioning system, comprised of two plasmid-coded proteins, Rep1 and Rep2, and a cis-acting partitioning locus STB, to ensure equal or nearly equal plasmid segregation. A decline in plasmid copy number because of rare missegregation events is corrected via DNA amplification triggered by the Flp sitespecific recombination system harbored by the plasmid (4, 5). Positive and negative regulatory circuits implemented through the plasmid-coded Raf1 protein and the Rep proteins ensure quick amplification response without the danger of runaway increase in copy number (6-8).The Rep-STB system channels several host factors involved in chromosome segregation toward the execution of plasmid segregation. These factors include the mitotic spindle, the spindleassociated motor Kip1, the RSC2 chromatin-remodeling complex, the centromere-specific histone H3-variant Cse4 (CenH3), and the yeast cohesin complex (9-15). The de novo assembly of the plasmid-partitioning complex at STB during the G1-S window of each cell cycle (9,12,14,16) is reminiscent of the assembly of the kinetochore complex at centromeres. However, kinetochore components have not been detected at STB by ChIP (13).The assembly of the plasmid-partitioning complex culmin...

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“…Specifically, histones H3 and H4 were found as dimers together with histone chaperones within predeposition complexes in human cells . Moreover, the investigation of CenH3 (centromeric-specific H3) nucleosomes identified hemisomes containing one copy each of CenH3, H2A, H2B, and H4 (Dalal et al 2007). Together these data suggest that histones H3 -H4 or CenH3 -H4 can be first provided as dimers.…”

Section: Stoichiometry Of Histone Depositionmentioning

confidence: 99%