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- EMDB-40972: CryoEM map of TR-TRAP -

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Basic information

Entry
Database: EMDB / ID: EMD-40972
TitleCryoEM map of TR-TRAP
Map dataCryoEM map of TR-TRAP
Sample
  • Complex: TR-TRAP/Mediator coactivator complex
Keywordsmouse mediator / complex / GENE REGULATION / CKM module
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 7.01 Å
AuthorsZhao H / Asturias F
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R01-67167 United States
National Institutes of Health/National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH/NIAMS) United States
CitationJournal: Mol Cell / Year: 2024
Title: An IDR-dependent mechanism for nuclear receptor control of Mediator interaction with RNA polymerase II.
Authors: Haiyan Zhao / Jiaqin Li / Yufei Xiang / Sohail Malik / Supriya V Vartak / Giovana M B Veronezi / Natalie Young / McKayla Riney / Jens Kalchschmidt / Andrea Conte / Seol Kyoung Jung / ...Authors: Haiyan Zhao / Jiaqin Li / Yufei Xiang / Sohail Malik / Supriya V Vartak / Giovana M B Veronezi / Natalie Young / McKayla Riney / Jens Kalchschmidt / Andrea Conte / Seol Kyoung Jung / Srinivas Ramachandran / Robert G Roeder / Yi Shi / Rafael Casellas / Francisco J Asturias /
Abstract: The essential Mediator (MED) coactivator complex plays a well-understood role in regulation of basal transcription in all eukaryotes, but the mechanism underlying its role in activator-dependent ...The essential Mediator (MED) coactivator complex plays a well-understood role in regulation of basal transcription in all eukaryotes, but the mechanism underlying its role in activator-dependent transcription remains unknown. We investigated modulation of metazoan MED interaction with RNA polymerase II (RNA Pol II) by antagonistic effects of the MED26 subunit and the CDK8 kinase module (CKM). Biochemical analysis of CKM-MED showed that the CKM blocks binding of the RNA Pol II carboxy-terminal domain (CTD), preventing RNA Pol II interaction. This restriction is eliminated by nuclear receptor (NR) binding to CKM-MED, which enables CTD binding in a MED26-dependent manner. Cryoelectron microscopy (cryo-EM) and crosslinking-mass spectrometry (XL-MS) revealed that the structural basis for modulation of CTD interaction with MED relates to a large intrinsically disordered region (IDR) in CKM subunit MED13 that blocks MED26 and CTD interaction with MED but is repositioned upon NR binding. Hence, NRs can control transcription initiation by priming CKM-MED for MED26-dependent RNA Pol II interaction.
History
DepositionJun 2, 2023-
Header (metadata) releaseJun 12, 2024-
Map releaseJun 12, 2024-
UpdateJul 17, 2024-
Current statusJul 17, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_40972.png

Downloads & links

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Map

FileDownload / File: emd_40972.map.gz / Format: CCP4 / Size: 83.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM map of TR-TRAP
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.22 Å/pix.
x 280 pix.
= 621.6 Å		2.22 Å/pix.
x 280 pix.
= 621.6 Å		2.22 Å/pix.
x 280 pix.
= 621.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.22 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.286
Minimum - Maximum-1.3119551 - 2.1643
Average (Standard dev.)0.00016598255 (±0.0431204)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions280280280
Spacing280280280
CellA=B=C: 621.60004 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half Map 1

Fileemd_40972_half_map_1.map
AnnotationHalf Map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half Map 2

Fileemd_40972_half_map_2.map
AnnotationHalf Map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : TR-TRAP/Mediator coactivator complex

EntireName: TR-TRAP/Mediator coactivator complex
Components
  • Complex: TR-TRAP/Mediator coactivator complex

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Supramolecule #1: TR-TRAP/Mediator coactivator complex

SupramoleculeName: TR-TRAP/Mediator coactivator complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#26
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 1.0 MDa

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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

Concentration0.1 mg/mL
BufferpH: 7.9
Sugar embeddingMaterial: ice
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: HOMEMADE PLUNGER

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
TemperatureMin: 70.0 K / Max: 70.0 K
SoftwareName: Leginon (ver. 9.03)
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 100.0 e/Å2
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated defocus max: 3.5 µm / Calibrated defocus min: 0.7000000000000001 µm / Calibrated magnification: 36000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.5 µm / Nominal defocus min: 0.7000000000000001 µm / Nominal magnification: 36000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 7.01 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 72812
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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