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- EMDB-19724: RNA polymerase II early elongation complex bound to TFIIE, TFIIF ... -

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Basic information

Entry
Database: EMDB / ID: EMD-19724
TitleRNA polymerase II early elongation complex bound to TFIIE, TFIIF and TFIIH - state a
Map data
Sample
  • Complex: mammalian RNA polymerase II general transcription factor IIE general transcription factor IIF general transcription factor IIH DNA
    • Complex: RNA polymerase II
KeywordsRNA polymerase II / promoter escape / de novo transcription / TRANSCRIPTION
Biological speciesSus scrofa (pig)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.0 Å
AuthorsZhan Y / Grabbe F / Oberbeckmann E / Dienemann C / Cramer P
Funding support Germany, 1 items
OrganizationGrant numberCountry
Max Planck Society Germany
CitationJournal: Mol Cell / Year: 2024
Title: Three-step mechanism of promoter escape by RNA polymerase II.
Authors: Yumeng Zhan / Frauke Grabbe / Elisa Oberbeckmann / Christian Dienemann / Patrick Cramer /
Abstract: The transition from transcription initiation to elongation is highly regulated in human cells but remains incompletely understood at the structural level. In particular, it is unclear how ...The transition from transcription initiation to elongation is highly regulated in human cells but remains incompletely understood at the structural level. In particular, it is unclear how interactions between RNA polymerase II (RNA Pol II) and initiation factors are broken to enable promoter escape. Here, we reconstitute RNA Pol II promoter escape in vitro and determine high-resolution structures of initially transcribing complexes containing 8-, 10-, and 12-nt ordered RNAs and two elongation complexes containing 14-nt RNAs. We suggest that promoter escape occurs in three major steps. First, the growing RNA displaces the B-reader element of the initiation factor TFIIB without evicting TFIIB. Second, the rewinding of the transcription bubble coincides with the eviction of TFIIA, TFIIB, and TBP. Third, the binding of DSIF and NELF facilitates TFIIE and TFIIH dissociation, establishing the paused elongation complex. This three-step model for promoter escape fills a gap in our understanding of the initiation-elongation transition of RNA Pol II transcription.
History
DepositionFeb 22, 2024-
Header (metadata) releaseApr 10, 2024-
Map releaseApr 10, 2024-
UpdateMay 15, 2024-
Current statusMay 15, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19724.png

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Map

FileDownload / File: emd_19724.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
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AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 400 pix.
= 420. Å		1.05 Å/pix.
x 400 pix.
= 420. Å		1.05 Å/pix.
x 400 pix.
= 420. Å

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Images are generated by Spider.

Voxel sizeX=Y=Z: 1.05 Å
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Contour LevelBy AUTHOR: 0.0275
Minimum - Maximum-0.02363689 - 0.12055555
Average (Standard dev.)0.0007459169 (±0.0059590745)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 419.99997 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19724_msk_1.map
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Half map: #1

Fileemd_19724_half_map_1.map
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Half map: #2

Fileemd_19724_half_map_2.map
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Sample components

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Entire : mammalian RNA polymerase II general transcription factor IIE gene...

EntireName: mammalian RNA polymerase II general transcription factor IIE general transcription factor IIF general transcription factor IIH DNA
Components
  • Complex: mammalian RNA polymerase II general transcription factor IIE general transcription factor IIF general transcription factor IIH DNA
    • Complex: RNA polymerase II

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Supramolecule #1: mammalian RNA polymerase II general transcription factor IIE gene...

SupramoleculeName: mammalian RNA polymerase II general transcription factor IIE general transcription factor IIF general transcription factor IIH DNA
type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Sus scrofa (pig)

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Supramolecule #2: RNA polymerase II

SupramoleculeName: RNA polymerase II / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: Sus scrofa (pig)

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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

BufferpH: 7.5
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 40.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.7 µm / Nominal defocus min: 0.7000000000000001 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:

12613

Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.0 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 12628
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD

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