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- EMDB-8479: Cryo-EM map of the transcriptional Mediator -

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Basic information

Entry
Database: EMDB / ID: 8479
TitleCryo-EM map of the transcriptional Mediator
Map dataSp Mediator
SampleMediator complexMediator (coactivator)
  • (Mediator complex subunit ...Mediator (coactivator)) x 16
Function/homologyMediator complex, subunit Med27 / Mediator complex subunit 27 / Mediator complex, subunit Med14 / Mediator complex, subunit Med31 / Mediator complex subunit MED14 / Mediator complex, subunit Med21 / Mediator complex, subunit Med7 / Mediator complex, subunit Med7/Med21-like / Mediator complex, subunit Med6, fungi / MED7 protein ...Mediator complex, subunit Med27 / Mediator complex subunit 27 / Mediator complex, subunit Med14 / Mediator complex, subunit Med31 / Mediator complex subunit MED14 / Mediator complex, subunit Med21 / Mediator complex, subunit Med7 / Mediator complex, subunit Med7/Med21-like / Mediator complex, subunit Med6, fungi / MED7 protein / Mediator complex, subunit Med17 / SOH1 / Mediator complex, subunit Med6 / Subunit 21 of Mediator complex / Mediator complex, subunit Med20 / Mediator complex, subunit Med8, fungi / Mediator of RNA polymerase II transcription subunit 22 / Mediator complex, subunit Med8, fungi/metazoa / Mediator complex, subunit Med18 / RNA polymerase II transcription coactivator activity involved in preinitiation complex assembly / Subunit 17 of Mediator complex / TATA-binding related factor (TRF) of subunit 20 of Mediator complex / MED6 mediator sub complex component / chromosome, centromeric outer repeat region / Surfeit locus protein 5 subunit 22 of Mediator complex / Mediator of RNA polymerase II transcription complex subunit 8 / chromatin silencing at centromere outer repeat region / core mediator complex / Med18 protein / mediator complex / RNA polymerase II transcriptional preinitiation complex assembly / RNA polymerase II transcription cofactor activity / termination of RNA polymerase II transcription / cellular protein localization / activation of protein kinase activity / transcription by RNA polymerase II / transcription coactivator activity / regulation of transcription by RNA polymerase II / transcription, DNA-templated / nucleus / cytosol / Mediator of RNA polymerase II transcription subunit 20 / Mediator of RNA polymerase II transcription subunit 6 / Mediator of RNA polymerase II transcription subunit 14 / Mediator of RNA polymerase II transcription subunit 11 / Mediator of RNA polymerase II transcription subunit 27 / Mediator of RNA polymerase II transcription subunit 18 / Mediator of RNA polymerase II transcription subunit 17 / Mediator of RNA polymerase II transcription subunit 8 / Mediator of RNA polymerase II transcription subunit 21 / Mediator of RNA polymerase II transcription subunit 7 / Mediator of RNA polymerase II transcription subunit 22 / Uncharacterized protein C24C9.04 / Mediator of RNA polymerase II transcription subunit 31
Function and homology information
SourceSchizosaccharomyces pombe / / yeast /
MethodCryo EM / single particle reconstruction / 4.4 Å resolution
AuthorsTsai K-L / Yu X
CitationJournal: Nature / Year: 2017
Title: Mediator structure and rearrangements required for holoenzyme formation.
Authors: Kuang-Lei Tsai / Xiaodi Yu / Sneha Gopalan / Ti-Chun Chao / Ying Zhang / Laurence Florens / Michael P Washburn / Kenji Murakami / Ronald C Conaway / Joan W Conaway / Francisco J Asturias
Abstract: The conserved Mediator co-activator complex has an essential role in the regulation of RNA polymerase II transcription in all eukaryotes. Understanding the structure and interactions of Mediator is ...The conserved Mediator co-activator complex has an essential role in the regulation of RNA polymerase II transcription in all eukaryotes. Understanding the structure and interactions of Mediator is crucial for determining how the complex influences transcription initiation and conveys regulatory information to the basal transcription machinery. Here we present a 4.4 Å resolution cryo-electron microscopy map of Schizosaccharomyces pombe Mediator in which conserved Mediator subunits are individually resolved. The essential Med14 subunit works as a central backbone that connects the Mediator head, middle and tail modules. Comparison with a 7.8 Å resolution cryo-electron microscopy map of a Mediator-RNA polymerase II holoenzyme reveals that changes in the structure of Med14 facilitate a large-scale Mediator rearrangement that is essential for holoenzyme formation. Our study suggests that access to different conformations and crosstalk between structural elements are essential for the Mediator regulation mechanism, and could explain the capacity of the complex to integrate multiple regulatory signals.
Validation ReportPDB-ID: 5u0p

SummaryFull reportAbout validation report
DateDeposition: Nov 26, 2016 / Header (metadata) release: Mar 8, 2017 / Map release: Mar 8, 2017 / Last update: Apr 19, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.016
  • Imaged by UCSF CHIMERA
  • Download
  • Surface view colored by radius
  • Surface level: 0.016
  • Imaged by UCSF CHIMERA
  • Download
  • Surface view with fitted model
  • Atomic models: : PDB-5u0p
  • Surface level: 0.016
  • Imaged by UCSF CHIMERA
  • Download
3D viewer
Supplemental images

Downloads & links

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Map

Fileemd_8479.map.gz (map file in CCP4 format, 174457 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
352 pix
1.31 Å/pix.
= 461.12 Å
352 pix
1.31 Å/pix.
= 461.12 Å
352 pix
1.31 Å/pix.
= 461.12 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 1.31 Å
Density
Contour Level:0.016 (by author), 0.016 (movie #1):
Minimum - Maximum-0.017003836 - 0.07525921
Average (Standard dev.)-3.3825316E-5 (0.0022060836)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions352352352
Origin000
Limit351351351
Spacing352352352
CellA=B=C: 461.12 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.311.311.31
M x/y/z352352352
origin x/y/z0.0000.0000.000
length x/y/z461.120461.120461.120
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS352352352
D min/max/mean-0.0170.075-0.000

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Supplemental data

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Mask #1

Fileemd_8479_msk_1.map ( map file in CCP4 format, 174457 KB )
Projections & Slices
AxesZYX
Projections
Slices (1/2)
Density Histograms
Data typeImage stored as Reals
Annotation detailsSp Mediator, mask
Space group number1

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Mask #1~

Fileemd_8479_msk_1.map
Projections & Slices
AxesZYX
Projections
Slices (1/2)
Density Histograms

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Sample components

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Entire Mediator complex

EntireName: Mediator complex / Number of components: 17

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Component #1: protein, Mediator complex

ProteinName: Mediator complexMediator (coactivator) / Recombinant expression: No
SourceSpecies: Schizosaccharomyces pombe

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Component #2: protein, Mediator complex subunit 14

ProteinName: Mediator complex subunit 14Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 105.382461 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #3: protein, Mediator complex subunit 6

ProteinName: Mediator complex subunit 6Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 24.928947 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #4: protein, Mediator complex subunit 8

ProteinName: Mediator complex subunit 8Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 23.360068 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #5: protein, Mediator complex subunit 17

ProteinName: Mediator complex subunit 17Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 62.55141 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #6: protein, Mediator complex subunit 18

ProteinName: Mediator complex subunit 18Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 24.056377 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #7: protein, Mediator complex subunit 20

ProteinName: Mediator complex subunit 20Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 22.374775 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #8: protein, Mediator complex subunit 11

ProteinName: Mediator complex subunit 11Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 13.1188 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #9: protein, Mediator complex subunit 22

ProteinName: Mediator complex subunit 22Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 15.396165 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #10: protein, Mediator complex subunit 4

ProteinName: Mediator complex subunit 4Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 20.35801 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #11: protein, Mediator complex subunit 7

ProteinName: Mediator complex subunit 7Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 43.573094 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #12: protein, Mediator complex subunit 21

ProteinName: Mediator complex subunit 21Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 15.821031 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #13: protein, Mediator complex subunit 31

ProteinName: Mediator complex subunit 31Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 16.913975 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #14: protein, Mediator complex subunit 9

ProteinName: Mediator complex subunit 9Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 13.810964 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #15: protein, Mediator complex subunit 27

ProteinName: Mediator complex subunit 27Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 31.132693 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #16: protein, Mediator complex subunit 19

ProteinName: Mediator complex subunit 19Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 11.847596 kDa
SourceSpecies: Schizosaccharomyces pombe

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Component #17: protein, Mediator complex subunit 10

ProteinName: Mediator complex subunit 10Mediator (coactivator) / Recombinant expression: No
MassTheoretical: 11.251861 kDa
SourceSpecies: Schizosaccharomyces pombe

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: Cryo EM
Sample solutionSpecimen conc.: 1 mg/ml
Buffer solution: 20 mM HEPES, pH 7.4, 200 mM potassium acetate, 5 mM b-Mercaptoethanol, 0.01% NP-40
pH: 7.4
VitrificationCryogen name: ETHANE

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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: 9.6 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 22500 X (nominal) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1000 - 4000 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 11181

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 42484
3D reconstructionSoftware: RELION / Resolution: 4.4 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution assessment)

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Atomic model buiding

Output model

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