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- EMDB-0768: H3-CENP-A-H3 tri-nucleosome with the 22 base-pair linker DNA -

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Basic information

Entry
Database: EMDB / ID: EMD-0768
TitleH3-CENP-A-H3 tri-nucleosome with the 22 base-pair linker DNA
Map data
SampleH3-CENP-A-H3 tri-nucleosome with the 22 base-pair linker DNA
Function / homology
Function and homology information


condensed chromosome inner kinetochore / kinetochore assembly / protein localization to chromosome, centromeric region / pericentric heterochromatin => GO:0005721 / condensed nuclear chromosome kinetochore / condensed chromosome, centromeric region => GO:0000779 / negative regulation of megakaryocyte differentiation / establishment of mitotic spindle orientation / CENP-A containing nucleosome assembly / negative regulation of tumor necrosis factor-mediated signaling pathway ...condensed chromosome inner kinetochore / kinetochore assembly / protein localization to chromosome, centromeric region / pericentric heterochromatin => GO:0005721 / condensed nuclear chromosome kinetochore / condensed chromosome, centromeric region => GO:0000779 / negative regulation of megakaryocyte differentiation / establishment of mitotic spindle orientation / CENP-A containing nucleosome assembly / negative regulation of tumor necrosis factor-mediated signaling pathway / chromosome, centromeric region / DNA replication-independent nucleosome assembly / telomere capping / interleukin-7-mediated signaling pathway / mitotic cytokinesis / chromatin silencing / telomere organization / DNA replication-dependent nucleosome assembly / nucleosome => GO:0000786 / innate immune response in mucosa / rDNA heterochromatin assembly / negative regulation of gene expression, epigenetic / regulation of gene silencing / regulation of gene silencing by miRNA / nuclear chromosome / DNA-templated transcription, initiation / regulation of megakaryocyte differentiation / nucleosome assembly / nucleosome / lipopolysaccharide binding / double-strand break repair via nonhomologous end joining / chromatin organization / chromosome, telomeric region => GO:0000781 / killing of cells of other organism / antibacterial humoral response / antimicrobial humoral immune response mediated by antimicrobial peptide / blood coagulation / protein ubiquitination / cadherin binding / defense response to Gram-negative bacterium / protein heterodimerization activity / defense response to Gram-positive bacterium / negative regulation of cell population proliferation / chromatin => GO:0000785 / protein domain specific binding / cellular protein metabolic process / chromatin binding / host cell nucleus / viral process / protein-containing complex / RNA binding / DNA binding / extracellular space / extracellular exosome / membrane / extracellular region / nucleoplasm / nucleus / cytosol
Histone H4, conserved site / Histone-fold / Histone H2A, C-terminal domain / Histone H2A conserved site / CENP-T/Histone H4, histone fold / Histone H2A/H2B/H3 / Histone H3/CENP-A / TATA box binding protein associated factor (TAF) / Histone H2A / Histone H4 / Histone H2B
Histone H2A type 1-B/E / Histone H2B type 1-J / Histone H3-like centromeric protein A / Histone H4 / Histone H3.1
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 18.7 Å
AuthorsTakizawa Y / Ho C-H / Tachiwana H / Ohi M / Wolf M / Kurumizaka H
Funding support Japan, 6 items
OrganizationGrant numberCountry
Japan Society for the Promotion of ScienceJP18H05534 Japan
Japan Society for the Promotion of ScienceJP17H05013 Japan
Japan Society for the Promotion of ScienceJP17H01408 Japan
Japan Society for the Promotion of Science15H05972 Japan
Japan Society for the Promotion of Science17H06167 Japan
Japan Society for the Promotion of ScienceJP19K06522 Japan
CitationJournal: Structure / Year: 2020
Title: Cryo-EM Structures of Centromeric Tri-nucleosomes Containing a Central CENP-A Nucleosome.
Authors: Yoshimasa Takizawa / Cheng-Han Ho / Hiroaki Tachiwana / Hideyuki Matsunami / Wataru Kobayashi / Midori Suzuki / Yasuhiro Arimura / Tetsuya Hori / Tatsuo Fukagawa / Melanie D Ohi / Matthias ...Authors: Yoshimasa Takizawa / Cheng-Han Ho / Hiroaki Tachiwana / Hideyuki Matsunami / Wataru Kobayashi / Midori Suzuki / Yasuhiro Arimura / Tetsuya Hori / Tatsuo Fukagawa / Melanie D Ohi / Matthias Wolf / Hitoshi Kurumizaka /
Abstract: The histone H3 variant CENP-A is a crucial epigenetic marker for centromere specification. CENP-A forms a characteristic nucleosome and dictates the higher-order configuration of centromeric ...The histone H3 variant CENP-A is a crucial epigenetic marker for centromere specification. CENP-A forms a characteristic nucleosome and dictates the higher-order configuration of centromeric chromatin. However, little is known about how the CENP-A nucleosome affects the architecture of centromeric chromatin. In this study, we reconstituted tri-nucleosomes mimicking a centromeric nucleosome arrangement containing the CENP-A nucleosome, and determined their 3D structures by cryoelectron microscopy. The H3-CENP-A-H3 tri-nucleosomes adopt an untwisted architecture, with an outward-facing linker DNA path between nucleosomes. This is distinct from the H3-H3-H3 tri-nucleosome architecture, with an inward-facing DNA path. Intriguingly, the untwisted architecture may allow the CENP-A nucleosome to be exposed to the solvent in the condensed chromatin model. These results provide a structural basis for understanding the 3D configuration of CENP-A-containing chromatin, and may explain how centromeric proteins can specifically target the CENP-A nucleosomes buried in robust amounts of H3 nucleosomes in centromeres.
Validation ReportPDB-ID: 6l49

SummaryFull reportAbout validation report
History
DepositionAug 23, 2019-
Header (metadata) releaseDec 4, 2019-
Map releaseDec 4, 2019-
UpdateDec 4, 2019-
Current statusDec 4, 2019Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 4
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6l49
  • Surface level: 4
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_0768.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.4 Å/pix.
x 280 pix.
= 392. Å
1.4 Å/pix.
x 280 pix.
= 392. Å
1.4 Å/pix.
x 280 pix.
= 392. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.4 Å
Density
Contour LevelBy AUTHOR: 4 / Movie #1: 4
Minimum - Maximum-3.761383 - 13.348018
Average (Standard dev.)-0.00000000181 (±0.99999994)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 392.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.41.41.4
M x/y/z280280280
origin x/y/z0.0000.0000.000
length x/y/z392.000392.000392.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS280280280
D min/max/mean-3.76113.348-0.000

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Supplemental data

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Sample components

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Entire H3-CENP-A-H3 tri-nucleosome with the 22 base-pair linker DNA

EntireName: H3-CENP-A-H3 tri-nucleosome with the 22 base-pair linker DNA
Number of components: 1

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Component #1: protein, H3-CENP-A-H3 tri-nucleosome with the 22 base-pair linker DNA

ProteinName: H3-CENP-A-H3 tri-nucleosome with the 22 base-pair linker DNA
Recombinant expression: No
MassTheoretical: 600 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.03 mg/mL / pH: 7.8
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
ImagingMicroscope: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 25 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.7 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: FEI FALCON II (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 4515

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 4999
3D reconstructionSoftware: RELION / Resolution: 18.7 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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Atomic model buiding

Modeling #1Refinement protocol: rigid body
Output model

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