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- EMDB-47263: Cryo-EM reconstruction of TC-NER transcription elongation complex... -

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Basic information

Entry
Database: EMDB / ID: EMD-47263
TitleCryo-EM reconstruction of TC-NER transcription elongation complex with STK19 (map II)
Map data
Sample
  • Complex: Transcription-coupled nucleotide excision repair complex
    • Complex: CSA, DDB1
    • Complex: CSB, UVSSA
    • Complex: DDA1, ELOF1, STK19
    • Complex: RNA Polymerase II proteins
    • Complex: Nucleic Acid
KeywordsDNA repair / RNA polymerase II / Complex / Co-transcriptional process / TRANSCRIPTION
Biological speciesHomo sapiens (human) / Sus scrofa (pig) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.1 Å
AuthorsMevissen TET / Kuemmecke M / Farnung L / Walter JC
Funding support United States, 3 items
OrganizationGrant numberCountry
Howard Hughes Medical Institute (HHMI) United States
National Institutes of Health/National Institute of Environmental Health Sciences (NIH/NIEHS)DP2-ES036404 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)HL098316 United States
CitationJournal: Cell / Year: 2024
Title: STK19 positions TFIIH for cell-free transcription-coupled DNA repair.
Authors: Tycho E T Mevissen / Maximilian Kümmecke / Ernst W Schmid / Lucas Farnung / Johannes C Walter /
Abstract: In transcription-coupled nucleotide excision repair (TC-NER), stalled RNA polymerase II (RNA Pol II) binds CSB and CRL4, which cooperate with UVSSA and ELOF1 to recruit TFIIH. To explore the ...In transcription-coupled nucleotide excision repair (TC-NER), stalled RNA polymerase II (RNA Pol II) binds CSB and CRL4, which cooperate with UVSSA and ELOF1 to recruit TFIIH. To explore the mechanism of TC-NER, we recapitulated this reaction in vitro. When a plasmid containing a site-specific lesion is transcribed in frog egg extract, error-free repair is observed that depends on CSB, CRL4, UVSSA, and ELOF1. Repair also requires STK19, a factor previously implicated in transcription recovery after UV exposure. A 1.9-Å cryo-electron microscopy structure shows that STK19 binds the TC-NER complex through CSA and the RPB1 subunit of RNA Pol II. Furthermore, AlphaFold predicts that STK19 interacts with the XPD subunit of TFIIH, and disrupting this interface impairs cell-free repair. Molecular modeling suggests that STK19 positions TFIIH ahead of RNA Pol II for lesion verification. Our analysis of cell-free TC-NER suggests that STK19 couples RNA Pol II stalling to downstream repair events.
History
DepositionOct 10, 2024-
Header (metadata) releaseDec 11, 2024-
Map releaseDec 11, 2024-
UpdateJan 8, 2025-
Current statusJan 8, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_47263.png

Downloads & links

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Map

FileDownload / File: emd_47263.map.gz / Format: CCP4 / Size: 391 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
0.94 Å/pix.
x 468 pix.
= 439.92 Å		0.94 Å/pix.
x 468 pix.
= 439.92 Å		0.94 Å/pix.
x 468 pix.
= 439.92 Å

Surface

Projections

Slices (1/3)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 0.94 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0495
Minimum - Maximum-0.1228975 - 0.30521968
Average (Standard dev.)-0.00024042414 (±0.0050328663)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions468468468
Spacing468468468
CellA=B=C: 439.92 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_47263_msk_1.map
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Mask #2

Fileemd_47263_msk_2.map
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Additional map: #1

Fileemd_47263_additional_1.map
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Half map: #1

Fileemd_47263_half_map_1.map
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Half map: #2

Fileemd_47263_half_map_2.map
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Sample components

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Entire : Transcription-coupled nucleotide excision repair complex

EntireName: Transcription-coupled nucleotide excision repair complex
Components
  • Complex: Transcription-coupled nucleotide excision repair complex
    • Complex: CSA, DDB1
    • Complex: CSB, UVSSA
    • Complex: DDA1, ELOF1, STK19
    • Complex: RNA Polymerase II proteins
    • Complex: Nucleic Acid

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Supramolecule #1: Transcription-coupled nucleotide excision repair complex

SupramoleculeName: Transcription-coupled nucleotide excision repair complex
type: complex / ID: 1 / Parent: 0

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Supramolecule #2: CSA, DDB1

SupramoleculeName: CSA, DDB1 / type: complex / ID: 2 / Parent: 1
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #3: CSB, UVSSA

SupramoleculeName: CSB, UVSSA / type: complex / ID: 3 / Parent: 1
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #4: DDA1, ELOF1, STK19

SupramoleculeName: DDA1, ELOF1, STK19 / type: complex / ID: 4 / Parent: 1
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #5: RNA Polymerase II proteins

SupramoleculeName: RNA Polymerase II proteins / type: complex / ID: 5 / Parent: 1
Source (natural)Organism: Sus scrofa (pig)

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Supramolecule #6: Nucleic Acid

SupramoleculeName: Nucleic Acid / type: complex / ID: 6 / Parent: 1
Source (natural)Organism: synthetic construct (others) / Synthetically produced: Yes

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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: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: TFS Selectris / Energy filter - Slit width: 10 eV
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Average electron dose: 52.4 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Nominal defocus max: 1.8 µm / Nominal defocus min: 0.6 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL
Final reconstructionResolution.type: BY AUTHOR / Resolution: 2.1 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 484012
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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