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- PDB-8ybk: Cryo-EM structure of the human nucleosome containing the H3.1 E97... -

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Entry
Database: PDB / ID: 8ybk
TitleCryo-EM structure of the human nucleosome containing the H3.1 E97K mutant
Components
  • (DNA (145-MER)) x 2
  • Histone H2A type 1-B/E
  • Histone H2B type 1-J
  • Histone H3.1
  • Histone H4
KeywordsDNA BINDING PROTEIN/DNA / chromatin / nucleosome / gene regulation / DNA BINDING PROTEIN-DNA COMPLEX
Function / homology
Function and homology information


negative regulation of tumor necrosis factor-mediated signaling pathway / negative regulation of megakaryocyte differentiation / protein localization to CENP-A containing chromatin / Chromatin modifying enzymes / Replacement of protamines by nucleosomes in the male pronucleus / CENP-A containing nucleosome / Packaging Of Telomere Ends / Recognition and association of DNA glycosylase with site containing an affected purine / Cleavage of the damaged purine / Deposition of new CENPA-containing nucleosomes at the centromere ...negative regulation of tumor necrosis factor-mediated signaling pathway / negative regulation of megakaryocyte differentiation / protein localization to CENP-A containing chromatin / Chromatin modifying enzymes / Replacement of protamines by nucleosomes in the male pronucleus / CENP-A containing nucleosome / Packaging Of Telomere Ends / Recognition and association of DNA glycosylase with site containing an affected purine / Cleavage of the damaged purine / Deposition of new CENPA-containing nucleosomes at the centromere / Recognition and association of DNA glycosylase with site containing an affected pyrimidine / Cleavage of the damaged pyrimidine / telomere organization / Inhibition of DNA recombination at telomere / Meiotic synapsis / Interleukin-7 signaling / RNA Polymerase I Promoter Opening / Assembly of the ORC complex at the origin of replication / Regulation of endogenous retroelements by the Human Silencing Hub (HUSH) complex / innate immune response in mucosa / SUMOylation of chromatin organization proteins / DNA methylation / Condensation of Prophase Chromosomes / Chromatin modifications during the maternal to zygotic transition (MZT) / HCMV Late Events / SIRT1 negatively regulates rRNA expression / epigenetic regulation of gene expression / ERCC6 (CSB) and EHMT2 (G9a) positively regulate rRNA expression / PRC2 methylates histones and DNA / Regulation of endogenous retroelements by KRAB-ZFP proteins / Defective pyroptosis / Regulation of endogenous retroelements by Piwi-interacting RNAs (piRNAs) / HDACs deacetylate histones / Nonhomologous End-Joining (NHEJ) / RNA Polymerase I Promoter Escape / lipopolysaccharide binding / Transcriptional regulation by small RNAs / Formation of the beta-catenin:TCF transactivating complex / RUNX1 regulates genes involved in megakaryocyte differentiation and platelet function / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / G2/M DNA damage checkpoint / HDMs demethylate histones / NoRC negatively regulates rRNA expression / DNA Damage/Telomere Stress Induced Senescence / B-WICH complex positively regulates rRNA expression / PKMTs methylate histone lysines / Meiotic recombination / Pre-NOTCH Transcription and Translation / Metalloprotease DUBs / RMTs methylate histone arginines / Activation of anterior HOX genes in hindbrain development during early embryogenesis / Transcriptional regulation of granulopoiesis / HCMV Early Events / antimicrobial humoral immune response mediated by antimicrobial peptide / structural constituent of chromatin / antibacterial humoral response / UCH proteinases / nucleosome / heterochromatin formation / E3 ubiquitin ligases ubiquitinate target proteins / nucleosome assembly / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / chromatin organization / HATs acetylate histones / RUNX1 regulates transcription of genes involved in differentiation of HSCs / Factors involved in megakaryocyte development and platelet production / MLL4 and MLL3 complexes regulate expression of PPARG target genes in adipogenesis and hepatic steatosis / Processing of DNA double-strand break ends / Senescence-Associated Secretory Phenotype (SASP) / defense response to Gram-negative bacterium / Oxidative Stress Induced Senescence / gene expression / killing of cells of another organism / Estrogen-dependent gene expression / chromosome, telomeric region / defense response to Gram-positive bacterium / Ub-specific processing proteases / cadherin binding / protein heterodimerization activity / Amyloid fiber formation / negative regulation of cell population proliferation / protein-containing complex / DNA binding / extracellular space / RNA binding / extracellular exosome / extracellular region / nucleoplasm / nucleus / membrane / cytosol
Similarity search - Function
: / Histone H2B signature. / Histone H2B / Histone H2B / Histone H2A conserved site / Histone H2A signature. / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone H2A / Histone 2A ...: / Histone H2B signature. / Histone H2B / Histone H2B / Histone H2A conserved site / Histone H2A signature. / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone H2A / Histone 2A / TATA box binding protein associated factor / TATA box binding protein associated factor (TAF), histone-like fold domain / Histone H4, conserved site / Histone H4 signature. / Histone H4 / Histone H4 / CENP-T/Histone H4, histone fold / Centromere kinetochore component CENP-T histone fold / Histone H3 signature 1. / Histone H3 signature 2. / Histone H3 / Histone H3/CENP-A / Histone H2A/H2B/H3 / Core histone H2A/H2B/H3/H4 / Histone-fold
Similarity search - Domain/homology
DNA / DNA (> 10) / DNA (> 100) / Histone H2A type 1-B/E / Histone H2B type 1-J / Histone H4 / Histone H3.1
Similarity search - Component
Biological speciesHomo sapiens (human)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.69 Å
AuthorsKimura, T. / Hirai, S. / Kujirai, T. / Ogasawara, M. / Takizawa, Y. / Kurumizaka, H.
Funding support Japan, 9items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP23H05475 Japan
Japan Society for the Promotion of Science (JSPS)JP22K06098 Japan
Japan Society for the Promotion of Science (JSPS)JP22K15033 Japan
Japan Society for the Promotion of Science (JSPS)JP20H03201 Japan
Japan Society for the Promotion of Science (JSPS)JP20H05690 Japan
Japan Society for the Promotion of Science (JSPS)JP22KJ0858 Japan
Japan Science and TechnologyJPMJER1901 Japan
Japan Agency for Medical Research and Development (AMED)JP23ama121009 Japan
Japan Society for the Promotion of Science (JSPS)JP23K17392 Japan
CitationJournal: Genes Cells / Year: 2024
Title: Cryo-EM structure and biochemical analyses of the nucleosome containing the cancer-associated histone H3 mutation E97K.
Authors: Tomoaki Kimura / Seiya Hirai / Tomoya Kujirai / Risa Fujita / Mitsuo Ogasawara / Haruhiko Ehara / Shun-Ichi Sekine / Yoshimasa Takizawa / Hitoshi Kurumizaka /
Abstract: The Lys mutation of the canonical histone H3.1 Glu97 residue (H3E97K) is found in cancer cells. Previous biochemical analyses revealed that the nucleosome containing the H3E97K mutation is extremely ...The Lys mutation of the canonical histone H3.1 Glu97 residue (H3E97K) is found in cancer cells. Previous biochemical analyses revealed that the nucleosome containing the H3E97K mutation is extremely unstable as compared to the wild-type nucleosome. However, the mechanism by which the H3E97K mutation causes nucleosome instability has not been clarified yet. In the present study, the cryo-electron microscopy structure of the nucleosome containing the H3E97K mutation revealed that the entry/exit DNA regions of the H3E97K nucleosome are disordered, probably by detachment of the nucleosomal DNA from the H3 N-terminal regions. This may change the intra-molecular amino acid interactions with the replaced H3 Lys97 residue, inducing structural distortion around the mutated position in the nucleosome. Consistent with the nucleosomal DNA end flexibility and the nucleosome instability, the H3E97K mutation exhibited reduced binding of linker histone H1 to the nucleosome, defective activation of PRC2 (the essential methyltransferase for facultative heterochromatin formation) with a poly-nucleosome, and enhanced nucleosome transcription by RNA polymerase II.
History
DepositionFeb 14, 2024Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jul 24, 2024Provider: repository / Type: Initial release
Revision 1.1Sep 11, 2024Group: Data collection / Database references / Category: citation / em_admin
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Histone H3.1
B: Histone H4
C: Histone H2A type 1-B/E
D: Histone H2B type 1-J
E: Histone H3.1
F: Histone H4
G: Histone H2A type 1-B/E
H: Histone H2B type 1-J
I: DNA (145-MER)
J: DNA (145-MER)


Theoretical massNumber of molelcules
Total (without water)201,63510
Polymers201,63510
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: native gel electrophoresis, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Protein , 4 types, 8 molecules AEBFCGDH

#1: Protein Histone H3.1 / Histone H3/a / Histone H3/b / Histone H3/c / Histone H3/d / Histone H3/f / Histone H3/h / Histone ...Histone H3/a / Histone H3/b / Histone H3/c / Histone H3/d / Histone H3/f / Histone H3/h / Histone H3/i / Histone H3/j / Histone H3/k / Histone H3/l


Mass: 15719.512 Da / Num. of mol.: 2 / Mutation: E97K
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: H3C1 / Production host: Escherichia coli (E. coli) / References: UniProt: P68431
#2: Protein Histone H4


Mass: 11676.703 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: H4C1 / Production host: Escherichia coli (E. coli) / References: UniProt: P62805
#3: Protein Histone H2A type 1-B/E / Histone H2A.2 / Histone H2A/a / Histone H2A/m


Mass: 14447.825 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: H2AC4 / Production host: Escherichia coli (E. coli) / References: UniProt: P04908
#4: Protein Histone H2B type 1-J / Histone H2B.1 / Histone H2B.r / H2B/r


Mass: 14217.516 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: H2BC11 / Production host: Escherichia coli (E. coli) / References: UniProt: P06899

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DNA chain , 2 types, 2 molecules IJ

#5: DNA chain DNA (145-MER)


Mass: 44520.383 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)
#6: DNA chain DNA (145-MER)


Mass: 44991.660 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Escherichia coli (E. coli)

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: The human nucleosome containing H3.1 E97K mutant / Type: COMPLEX
Details: Human nucleosome with 145 bp Widom 601 DNA and H3.1 E97K mutant
Entity ID: all / Source: RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Details: Human nucleosome with 145 bp Widom 601 DNA sequence and H3.1 E97K mutant
Specimen supportGrid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE-PROPANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm
Specimen holderCryogen: NITROGEN
Image recordingElectron dose: 58.935 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

EM software
IDNameVersionCategory
10RELION4initial Euler assignment
11RELION4final Euler assignment
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.69 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 1569054 / Symmetry type: POINT

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