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- EMDB-60592: RNA polymerase II elongation complex stalled at SHL(-5) of the nu... -

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Basic information

Entry
Database: EMDB / ID: EMD-60592
TitleRNA polymerase II elongation complex stalled at SHL(-5) of the nucleosome containing histone variant H2A.B
Map data
Sample
  • Complex: Spt4/5-Elf1-RNAPII-H2A.B nucleosome complex
    • Complex: RNA polymerase II
    • Complex: Histone complex
      • Complex: H2A.B
      • Complex: H2B
      • Complex: H3.3
      • Complex: H4
    • Complex: DNA
    • Complex: RNA
    • Complex: TFIIS
    • Complex: Spt4
    • Complex: Spt5
    • Complex: Elf1
KeywordsTranscription-DNA-RNA COMPLEX / RNAPII / TRANSCRIPTION
Biological speciesKomagataella phaffii (fungus) / Homo sapiens (human) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.7 Å
AuthorsAkatsu M / Kujirai T / Hirano R / Ehara H / Takizawa Y / Sekine S / Kurumizaka H
Funding support Japan, 10 items
OrganizationGrant numberCountry
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)JP22K15033 Japan
Japan Society for the Promotion of Science (JSPS)JP22K06098 Japan
Japan Society for the Promotion of Science (JSPS)JP23H05475 Japan
Japan Society for the Promotion of Science (JSPS)JP23K17392 Japan
Japan Society for the Promotion of Science (JSPS)JP24H02328 Japan
Japan Science and TechnologyJPMJER1901 Japan
Japan Science and TechnologyJPMJSP2108 Japan
Japan Agency for Medical Research and Development (AMED)JP24ama121009 Japan
CitationJournal: EMBO J / Year: 2025
Title: Structural basis of RNAPII transcription on the nucleosome containing histone variant H2A.B.
Authors: Munetaka Akatsu / Rina Hirano / Tomoya Kujirai / Mitsuo Ogasawara / Haruhiko Ehara / Shun-Ichi Sekine / Yoshimasa Takizawa / Hitoshi Kurumizaka /
Abstract: H2A.B is a distant histone H2A variant associated with actively transcribed regions of the genome, suggesting its positive role in promoting transcription. In the present study, we demonstrate that ...H2A.B is a distant histone H2A variant associated with actively transcribed regions of the genome, suggesting its positive role in promoting transcription. In the present study, we demonstrate that the RNA polymerase II elongation complex (EC) transcribes the nucleosome containing H2A.B more efficiently than that with canonical H2A in vitro. Our cryo-electron microscopy analysis of the H2A.B nucleosome during transcription revealed that the proximal H2A.B-H2B dimer is released from the nucleosome as the EC transcribes the proximal half of the nucleosomal DNA. This dissociation, which is not observed in the canonical H2A nucleosome, likely enhances the EC elongation efficiency in the H2A.B nucleosome. Mutational analyses suggested that the unique short C-terminal region of H2A.B alone enhances EC elongation efficiency when substituted for its counterpart in canonical H2A. Additionally, other regions of H2A.B contribute to this enhancement. These structural and biochemical findings provide new insights into the role of H2A.B in regulating gene expression.
History
DepositionJun 19, 2024-
Header (metadata) releaseJun 11, 2025-
Map releaseJun 11, 2025-
UpdateJun 11, 2025-
Current statusJun 11, 2025Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_60592.png

Downloads & links

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Map

FileDownload / File: emd_60592.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.06 Å/pix.
x 360 pix.
= 381.6 Å		1.06 Å/pix.
x 360 pix.
= 381.6 Å		1.06 Å/pix.
x 360 pix.
= 381.6 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.00464
Minimum - Maximum-0.008368411 - 0.025199698
Average (Standard dev.)0.00021278049 (±0.0012599676)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 381.59998 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: #3

Fileemd_60592_additional_1.map
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Additional map: #2

Fileemd_60592_additional_2.map
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Additional map: #1

Fileemd_60592_additional_3.map
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Additional map: #4

Fileemd_60592_additional_4.map
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Half map: #2

Fileemd_60592_half_map_1.map
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Half map: #1

Fileemd_60592_half_map_2.map
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Sample components

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Entire : Spt4/5-Elf1-RNAPII-H2A.B nucleosome complex

EntireName: Spt4/5-Elf1-RNAPII-H2A.B nucleosome complex
Components
  • Complex: Spt4/5-Elf1-RNAPII-H2A.B nucleosome complex
    • Complex: RNA polymerase II
    • Complex: Histone complex
      • Complex: H2A.B
      • Complex: H2B
      • Complex: H3.3
      • Complex: H4
    • Complex: DNA
    • Complex: RNA
    • Complex: TFIIS
    • Complex: Spt4
    • Complex: Spt5
    • Complex: Elf1

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Supramolecule #1: Spt4/5-Elf1-RNAPII-H2A.B nucleosome complex

SupramoleculeName: Spt4/5-Elf1-RNAPII-H2A.B nucleosome complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#22

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Supramolecule #2: RNA polymerase II

SupramoleculeName: RNA polymerase II / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#12
Source (natural)Organism: Komagataella phaffii (fungus)

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Supramolecule #3: Histone complex

SupramoleculeName: Histone complex / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #17-#20
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #4: DNA

SupramoleculeName: DNA / type: complex / ID: 4 / Parent: 1 / Macromolecule list: #21-#22
Source (natural)Organism: synthetic construct (others)

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Supramolecule #5: RNA

SupramoleculeName: RNA / type: complex / ID: 5 / Parent: 1 / Macromolecule list: #14
Source (natural)Organism: synthetic construct (others) / Synthetically produced: Yes

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Supramolecule #6: TFIIS

SupramoleculeName: TFIIS / type: complex / ID: 6 / Parent: 1 / Macromolecule list: #13
Source (natural)Organism: Komagataella phaffii (fungus)

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Supramolecule #7: Spt4

SupramoleculeName: Spt4 / type: complex / ID: 7 / Parent: 1 / Macromolecule list: #15
Source (natural)Organism: Komagataella phaffii (fungus)

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Supramolecule #8: Spt5

SupramoleculeName: Spt5 / type: complex / ID: 8 / Parent: 1 / Macromolecule list: #16
Source (natural)Organism: Komagataella phaffii (fungus)

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Supramolecule #9: Elf1

SupramoleculeName: Elf1 / type: complex / ID: 9 / Parent: 1 / Macromolecule list: #13
Source (natural)Organism: Komagataella phaffii (fungus)

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Supramolecule #10: H2A.B

SupramoleculeName: H2A.B / type: complex / ID: 10 / Parent: 3 / Macromolecule list: #17
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #11: H2B

SupramoleculeName: H2B / type: complex / ID: 11 / Parent: 3 / Macromolecule list: #18
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #12: H3.3

SupramoleculeName: H3.3 / type: complex / ID: 12 / Parent: 3 / Macromolecule list: #19
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #13: H4

SupramoleculeName: H4 / type: complex / ID: 13 / Parent: 3 / Macromolecule list: #20
Source (natural)Organism: Homo sapiens (human)

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.09144 mg/mL
BufferpH: 7.5
Component:
ConcentrationNameFormula
20.0 mMHEPES-KOH(pH7.5)
50.0 mMPotassium acetateCH3COOK
200.0 nMZinc acetate(CH3COO)2Zn
0.1 mMTCEP-HCl

Details: 20 mM HEPES-KOH(pH7.5), 50 mM Potassium acetate, 200 nM Zinc acetate, 0.1 mM TCEP-HCl
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK III
DetailsdsDNA concentration is 0.09144 mg/mL. This sample contains 0.005% Tween20.

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 57.24 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Startup modelType of model: EMDB MAP
EMDB ID:

9713

Final reconstructionResolution.type: BY AUTHOR / Resolution: 4.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 62759
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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