+Open data
-Basic information
Entry | Database: PDB / ID: 5u0s | |||||||||
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Title | Cryo-EM structure of the Mediator-RNAPII complex | |||||||||
Components |
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Keywords | TRANSCRIPTION/TRANSFERASE / transcriptional / transcription / Mediator / Srb / RNA polymerase II / RNA / Pol2 / activation / complex / TRANSCRIPTION-TRANSFERASE complex | |||||||||
Function / homology | Function and homology information RNA Polymerase I Transcription Initiation / RNA polymerase II transcribes snRNA genes / Processing of Capped Intron-Containing Pre-mRNA / Formation of TC-NER Pre-Incision Complex / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / Estrogen-dependent gene expression / RNA Polymerase I Promoter Escape ...RNA Polymerase I Transcription Initiation / RNA polymerase II transcribes snRNA genes / Processing of Capped Intron-Containing Pre-mRNA / Formation of TC-NER Pre-Incision Complex / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / Estrogen-dependent gene expression / RNA Polymerase I Promoter Escape / RNA Polymerase III Transcription Initiation From Type 1 Promoter / RNA Polymerase III Transcription Initiation From Type 2 Promoter / mRNA Capping / RNA Pol II CTD phosphorylation and interaction with CE / Formation of the Early Elongation Complex / RNA Polymerase II Pre-transcription Events / TP53 Regulates Transcription of DNA Repair Genes / RNA Polymerase II Transcription Elongation / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / regulation of septum digestion after cytokinesis / regulatory ncRNA-mediated heterochromatin formation / DNA-templated transcription elongation / core mediator complex / mediator complex / : / : / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / termination of RNA polymerase II transcription / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / tRNA transcription by RNA polymerase III / transcription elongation by RNA polymerase I / positive regulation of translational initiation / RNA polymerase II activity / transcription-coupled nucleotide-excision repair / positive regulation of transcription initiation by RNA polymerase II / RNA polymerase I complex / transcription by RNA polymerase I / RNA polymerase III complex / transcription by RNA polymerase III / pericentric heterochromatin / RNA polymerase II, core complex / translation initiation factor binding / P-body / transcription initiation at RNA polymerase II promoter / transcription coregulator activity / euchromatin / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / single-stranded DNA binding / transcription by RNA polymerase II / nucleic acid binding / single-stranded RNA binding / transcription coactivator activity / protein dimerization activity / RNA polymerase II cis-regulatory region sequence-specific DNA binding / nucleotide binding / chromatin / nucleolus / regulation of transcription by RNA polymerase II / DNA binding / RNA binding / zinc ion binding / metal ion binding / nucleus / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Schizosaccharomyces pombe (fission yeast) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 7.8 Å | |||||||||
Authors | Tsai, K.-L. / Yu, X. / Gopalan, S. / Chao, T.-C. / Zhang, Y. / Florens, L. / Washburn, M.P. / Murakami, K. / Conaway, R.C. / Conaway, J.W. / Asturias, F. | |||||||||
Funding support | United States, 2items
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Citation | Journal: 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. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 5u0s.cif.gz | 1.3 MB | Display | PDBx/mmCIF format |
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PDB format | pdb5u0s.ent.gz | 1012.7 KB | Display | PDB format |
PDBx/mmJSON format | 5u0s.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/u0/5u0s ftp://data.pdbj.org/pub/pdb/validation_reports/u0/5u0s | HTTPS FTP |
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-Related structure data
Related structure data | 8480MC 8479C 5u0pC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Mediator complex subunit ... , 16 types, 16 molecules FHQRTKVNDGU32ISJ
#1: Protein | Mass: 24928.947 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q9US45 |
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#2: Protein | Mass: 23360.068 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O94646 |
#3: Protein | Mass: 62551.410 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: P87306 |
#4: Protein | Mass: 24056.377 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O14198 |
#5: Protein | Mass: 22374.775 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q10317 |
#6: Protein | Mass: 13118.800 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q9P6Q0 |
#7: Protein | Mass: 15396.165 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O14010 |
#8: Protein | Mass: 105382.461 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q9P7Y4 |
#9: Protein | Mass: 20358.010 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) |
#10: Protein | Mass: 43573.094 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O60104 |
#11: Protein | Mass: 15821.031 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O94376 |
#12: Protein | Mass: 16913.975 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q9USH1 |
#13: Protein | Mass: 31132.693 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q10477 |
#14: Protein | Mass: 13810.964 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O13964 |
#15: Protein | Mass: 11847.596 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) |
#16: Protein | Mass: 11251.861 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) |
-RNA polymerase II subunit ... , 12 types, 12 molecules abcdefghijkl
#17: Protein | Mass: 194372.625 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: P36594, DNA-directed RNA polymerase |
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#18: Protein | Mass: 138027.188 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q02061, DNA-directed RNA polymerase |
#19: Protein | Mass: 33748.816 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: P37382 |
#20: Protein | Mass: 15379.490 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O74825 |
#21: Protein | Mass: 23954.504 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q09191 |
#22: Protein | Mass: 15742.497 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: P36595 |
#23: Protein | Mass: 19121.982 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O14459 |
#24: Protein | Mass: 14317.318 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q92399 |
#25: Protein | Mass: 13192.771 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O74635 |
#26: Protein | Mass: 8286.801 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O13877 |
#27: Protein | Mass: 14143.276 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: P87123 |
#28: Protein | Mass: 7216.495 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: P48011 |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: complex of Mediator and RNAPII / Type: COMPLEX / Entity ID: all / Source: NATURAL |
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Molecular weight | Value: 1.5 MDa / Experimental value: NO |
Source (natural) | Organism: Schizosaccharomyces pombe (fission yeast) |
Buffer solution | pH: 7.4 Details: 20 mM HEPES, pH 7.4, 200 mM potassium acetate, 5 mM b-Mercaptoethanol, 0.01% NP-40 |
Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: C-flat-2/2 4C |
Vitrification | Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Humidity: 100 % |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 22500 X / Nominal defocus max: 4000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 7 sec. / Electron dose: 9.6 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||
3D reconstruction | Resolution: 7.8 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 3862 / Symmetry type: POINT |