Genome-wide maps of CENP-A nucleosomes in three neocentromere-containing cell lines
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ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Ben Black
PROVIDER: E-GEOD-44724 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:We used native ChIP-seq of CENP-A-containing particles from normal centromeres on alpha-satellite DNA and three naturally-occurring neocentromeres to test the proposed models for the major form of the fundamental repeating unit of centromeric chromatin. We found that the predominant form of the CENP-A particle at the centromere is an octameric nucleosome with loose terminal DNA. Additionally, we found CENP-A nucleosomes are strongly phased on the 171 bp alpha-satellite monomers of normal centromeres, and also display strong positioning and neocentromeres.
2013-05-04 | GSE44724 | GEO
Project description:We use high-resolution chemical cleavage mapping and both native and cross-linked chromatin immunoprecipitation with paired-end sequencing to elucidate the profile of nuceleosomes containing the centromere-specific variant of H3 (cenH3), known as CENP-A or Cnp1 in fission yeast. We find that in the central domain of fission yeast centromeres H3 nucleosomes are nearly absent and CENP-A nucleosomes are more widely spaced that nucleosomes elsewere. CENP-A (Cnp1), CENP-C (Cnp3), CENP-T (Cnp20) and CENP-I (Mis6) are highly enriched at every position in the central domain except at tRNA genes, with weak enrichment in the flanking heterochromatin where these proteins show no evidence of the positioning that has been seen in point centromeres and in the satellite-rich centromeres of plants and animals. Our findings suggest that classical regional centromeres are distinguished from other centromere classes by the absence of cenH3 nucleosome positioning.
2015-08-17 | GSE64294 | GEO
Project description:Epigenetic mechanisms are thought to specify the identity of centromeres, yet the degree to which DNA sequence contributes to centromere determination remains unresolved. For example, the conserved CENP-B protein binds the 17-bp âCENP-B boxâ motif found in α-satellite arrays at most human centromeres. Although CENP-B is required for artificial centromere function, it is non-essential and some species lack CENP-B boxes entirely. To address this âCENP-B paradox,â we determined the distribution of CENP-B boxes in primates and directly demonstrated the absence of CENP-B at Old World Monkey centromeres. We found extensive interspecific variation in α-satellite arrays including abundance of <10-bp dyad symmetries and of non-B-form DNA structures. We confirmed the presence of non-canonical DNA at human centromeres and neocentromeres and detected similar structures at mouse, chicken and yeast centromeres. To resolve the CENP-B paradox, we propose that CENP-B enhances non-canonical DNA formation, thus providing a general basis for centromere specification.
2018-01-28 | GSE102111 | GEO
Project description:The focal attachment of the kinetochore to the centromere core is essential for genome maintenance, yet the highly repetitive nature of human centromeres limits our understanding of their chromatin organization. We demonstrate that single-molecule chromatin fiber sequencing can uniquely resolve chromatin organization within centromeres at single-molecule and single-nucleotide resolution. We find that the centromere core contains a dichotomous chromatin organization not found elsewhere in the genome, which is characterized by highly accessible chromatin patches heterogeneously punctuated amongst tightly compacted nucleosome arrays. These highly accessible chromatin patches correspond to sites of kinetochore attachment, and clustered CENP-B occupancy within these patches phase nucleosome arrays to the alpha-satellite repeat. This dichotomous chromatin organization is conserved between humans despite the marked divergence of the underlying alpha-satellite organization and is similarly conserved in primate centromeres that lack alpha-satellite repeats, indicating that functional conservation within centromeres is mediated at the level of chromatin, not DNA.
2023-08-02 | GSE226394 | GEO
Project description:Chromatin assembled with histone H3 variant CENP-A is the heritable epigenetic determinant of human centromere identity. Using genome-wide mapping and reference models for 23 human centromeres, CENP-A is shown in early G1 to be assembled into nucleosomes within megabase, repetitive a-satellite DNAs at each centromere and onto 11,390 sites on the chromosome arms. Centromere-bound CENP-A is found to be quantitatively maintained during DNA replication by coordinated action of the MCM2 helicase, CAF1, HJURP, and the CCAN network of constitutive centromere components. CCAN serves to tether CENP-A removed by MCM2, thereby enabling local reassembly onto both daughter centromeres with identical DNA sequence preferences and nucleosome phasing as the loading in G1 and independent of CENP-B. Conversely, without CCAN-mediated tethering, DNA replication removes CENP-A from sites on the chromosome arms. Our data identify an MCM2/CAF1/HJURP- and CCAN-dependent error correction mechanism that acts in S-phase to maintain CENP-A-dependent centromere identity.
2019-03-20 | GSE111381 | GEO
Project description:We find that CENP-T acts as a bridge between two well-positioned CENP-A nucleosomes that are present on young alpha-satellite dimers that dominate functional human centromeres. CENP-T is centered over the CENP-B box, where it interacts with the CENP-B/CENP-C complex. Upon cross-linking, the entire CENP-A/CENP-C/CENP-T-containing complex is recovered as a nuclease-protected particle over an alpha-satellite dimer that comprises the fundamental unit of kinetochore chromatin. Our work reveals that CENP-A/CENP-C and CENP-T branches of kinetochore assembly are physically integrated.
2016-05-24 | GSE69839 | GEO
Project description:CENP-A is a centromere-specific histone 3 variant essential for centromere specification. CENP-A partially replaces canonical histone H3 at the centromeres. How the particular CENP-A/H3 ratio at centromeres is precisely maintained is unknown. It also remains unclear how CENP-A is excluded from non-centromeric chromatin. Here we identify Ccp1, an uncharacterized NAP family protein in fission yeast that antagonizes CENP-A loading at both centromeric and non-centromeric regions. Like the CENP-A loading factor HJURP, Ccp1 interacts with CENP-A, and is recruited to centromeres at the end of mitosis in a Mis16-dependent manner. These data indicate that factors with opposing CENP-A loading activities are recruited to centromeres. Furthermore, Ccp1 also cooperates with H2A.Z to evict CENP-A assembled in euchromatin. Structural analyses indicate that Ccp1 forms a homodimer that is required for its anti-CENP-A loading activity. Our study establishes mechanisms for maintenance of CENP-A homeostasis at centromeres and the prevention of ectopic assembly of centromeres. Examination of cnp1 distribution in one wild type (wt) and two ccp1 mutants.
2016-08-04 | E-GEOD-85166 | biostudies-arrayexpress
Project description:CENP-A chromatin the foundation for kinetochore assembly in all eukaryotes. CENP-A is incorporated outside of S phase and depends on its chaperone HJURPScm3, and Mis18 in vertebrates and fission yeast. The recruitment of Mis18 and HJURPScm3 to centromeres is cell cycle regulated. Vertebrate Mis18 associates with Mis18BP1KNL2 and RbAp46/48Mis16/Hat2. Mis18BP1KNL2 is critical for the recruitment of Mis18 and HJURPScm3 to vertebrate centromeres. However, no ortholog of Mis18BP1KNL2 has been identified in fission yeast, consequently it remains unknown how the key Cnp1CENP-A loading factor Mis18 is recruited. We identify two novel Mis18-interacting proteins (Eic1 and Eic2) in fission yeast. Our analyses show that Eic1 and Eic2 are components of a Mis18 complex. Eic1 is essential to maintain normal Cnp1CENP-A levels at centromeres and therefore is crucial for kinetochore integrity whereas Eic2 is dispensable. Eic1 also associates with components of the large constitutive CCAN/Sim4/Ctf19 complex. Our findings suggest that Eic1 is the functional counterpart of Mis18BP1KNL2 and acts as a link with constitutive kinetochore components to allow the temporally regulated recruitment of the Mis18 Cnp1CENP-A loader. 100bp paired end ChIP-Seq analysis of GFP tagged Eic1, Eic2, Scm3, and Mis18 with sonicated whole cell extract input DNA in asynchornous Schizosaccharomyces pombe cells.
2014-05-19 | E-GEOD-54685 | biostudies-arrayexpress
Project description:Centromeres, the sites of spindle attachment during mitosis and meiosis, are located in specific positions in the human genome, normally coincident with diverse subsets of alpha satellite DNA. While there is strong evidence supporting the association of some subfamilies of alpha satellite with centromere function, the basis for establishing whether a given alpha satellite sequence is or is not designated a functional centromere is unknown, and attempts to understand the role of particular sequence features in establishing centromere identity have been limited by the near identity and repetitive nature of satellite sequences. Utilizing a broadly applicable experimental approach to test sequence competency for centromere specification, we have carried out a genomic and epigenetic functional analysis of endogenous human centromere sequences available in the current human genome assembly. The data support a model in which functionally competent sequences confer an opportunity for centromere specification, integrating genomic and epigenetic signals and promoting the concept of context-dependent centromere inheritance. Sequences bound to CENP-A in HuRef cell line
2013-03-29 | E-GEOD-45497 | biostudies-arrayexpress
Project description:Centromeres of most eukaryotes are multi-megabase arrays of satellite DNA that assemble proteinaceous kinetochores to facilitate faithful chromosome segregation. However, the nature of the chromatin landscape at centromeres remains unclear, in part due to the difficulty of isolating intact centromeric chromatin rendered insoluble by megadalton-size kinetochore protein complexes. To address this challenge we combined classical salt fractionation with chromatin immunoprecipitation to recover human centromeric chromatin under native conditions. We found that >85% of the total centromeric chromatin is insoluble under conditions typically used for native chromatin extraction. To map both soluble and insoluble chromatin in situ, we combined CUT&RUN, a targeted nuclease method, with salt fractionation. Using this approach, we found that the strength of Centromere Protein B (CENP-B) binding and the density of CENP-B motifs corresponds to the occupancy of Constitutive Centromere-Assocated Network (CCAN) complexes bound to α-satellite arrays. We also observed unexpected structural variations of CENP-A-containing complexes on different α-satellite dimeric units within highly homogenous arrays. Our results suggest that slight α-satellite sequence differences controls the structure and occupancy of the associated centromeric chromatin complex.
2018-01-31 | GSE104805 | GEO