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- EMDB-11975: Cryo-EM map of the transcribing Pol II-U1 snRNP complex with 155-... -

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Basic information

Entry
Database: EMDB / ID: EMD-11975
TitleCryo-EM map of the transcribing Pol II-U1 snRNP complex with 155-nt RNA
Map data
SampleThe transcribing RNA polymerase II-U1 snRNP complex with 155-nt RNA:
RNA polymerase II / Nucleic acidsNucleic acid / U1 snRNP
Biological speciesSus scrofa domesticus (domestic pig) / synthetic construct (others) / Homo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.9 Å
AuthorsZhang S / Aibara S / Vos SM / Agafonov DE / Luehrmann R / Cramer P
Funding support4 items
OrganizationGrant numberCountry
European Molecular Biology Organization (EMBO)ALTF 830-2018
European Research Council (ERC)882357
H2020 Marie Curie Actions of the European Commission894862
German Research Foundation (DFG)EXC 2067/1-390729940
CitationJournal: Science / Year: 2021
Title: Structure of a transcribing RNA polymerase II-U1 snRNP complex.
Authors: Suyang Zhang / Shintaro Aibara / Seychelle M Vos / Dmitry E Agafonov / Reinhard Lührmann / Patrick Cramer /
Abstract: To initiate cotranscriptional splicing, RNA polymerase II (Pol II) recruits the U1 small nuclear ribonucleoprotein particle (U1 snRNP) to nascent precursor messenger RNA (pre-mRNA). Here, we report ...To initiate cotranscriptional splicing, RNA polymerase II (Pol II) recruits the U1 small nuclear ribonucleoprotein particle (U1 snRNP) to nascent precursor messenger RNA (pre-mRNA). Here, we report the cryo-electron microscopy structure of a mammalian transcribing Pol II-U1 snRNP complex. The structure reveals that Pol II and U1 snRNP interact directly. This interaction positions the pre-mRNA 5' splice site near the RNA exit site of Pol II. Extension of pre-mRNA retains the 5' splice site, leading to the formation of a "growing intron loop." Loop formation may facilitate scanning of nascent pre-mRNA for the 3' splice site, functional pairing of distant intron ends, and prespliceosome assembly. Our results provide a starting point for a mechanistic analysis of cotranscriptional spliceosome assembly and the biogenesis of mRNA isoforms by alternative splicing.
Validation ReportSummary, Full report, XML, About validation report
History
DepositionNov 23, 2020-
Header (metadata) releaseJan 20, 2021-
Map releaseJan 20, 2021-
UpdateJan 27, 2021-
Current statusJan 27, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 4
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 4
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_11975.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesX (Sec.)Y (Row.)Z (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

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.05 Å
Density
Contour LevelBy AUTHOR: 4.0 / Movie #1: 4
Minimum - Maximum-16.949575 - 30.356455
Average (Standard dev.)1.5985361e-12 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

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

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.051.051.05
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z420.000420.000420.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ400400400
MAP C/R/S321
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-16.95030.3560.000

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Supplemental data

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Segmentation: #1

Fileemd_11975_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Unsharpened map

Fileemd_11975_additional_1.map
AnnotationUnsharpened map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map 2

Fileemd_11975_half_map_1.map
AnnotationHalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: Half map 1

Fileemd_11975_half_map_2.map
AnnotationHalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire The transcribing RNA polymerase II-U1 snRNP complex with 155-nt RNA

EntireName: The transcribing RNA polymerase II-U1 snRNP complex with 155-nt RNA
Number of components: 4

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Component #1: protein, The transcribing RNA polymerase II-U1 snRNP complex with...

ProteinName: The transcribing RNA polymerase II-U1 snRNP complex with 155-nt RNA
Recombinant expression: No

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Component #2: protein, RNA polymerase II

ProteinName: RNA polymerase II / Recombinant expression: No
SourceSpecies: Sus scrofa domesticus (domestic pig)

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Component #3: protein, Nucleic acids

ProteinName: Nucleic acidsNucleic acid / Recombinant expression: No
SourceSpecies: synthetic construct (others)
Source (engineered)Expression System: synthetic construct (others)

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Component #4: protein, U1 snRNP

ProteinName: U1 snRNP / Recombinant expression: No
SourceSpecies: Homo sapiens (human)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionpH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277.15 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 1.01 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: OTHER

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 14497
3D reconstructionSoftware: RELION / Resolution: 3.9 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

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