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- PDB-8b3g: C(N)RL4CSA-UVSSA-E2-ubiquitin complex. -

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Basic information

Entry
Database: PDB / ID: 8b3g
TitleC(N)RL4CSA-UVSSA-E2-ubiquitin complex.
Components
  • Cullin-4A
  • DNA damage-binding protein 1
  • DNA excision repair protein ERCC-8
  • E3 ubiquitin-protein ligase RBX1
  • NEDD8
  • UV-stimulated scaffold protein A
  • Ubiquitin
  • ubiquitin-conjugating enzyme E2 D2 isoform X3
KeywordsTRANSCRIPTION / DNA repair / ubiquitin / cryo-EM
Function / homology
Function and homology information


regulation of transcription-coupled nucleotide-excision repair / nucleotide-excision repair complex / negative regulation of granulocyte differentiation / single strand break repair / (E3-independent) E2 ubiquitin-conjugating enzyme / cullin-RING-type E3 NEDD8 transferase / NEDD8 transferase activity / cullin-RING ubiquitin ligase complex / positive regulation by virus of viral protein levels in host cell / regulation of DNA damage checkpoint ...regulation of transcription-coupled nucleotide-excision repair / nucleotide-excision repair complex / negative regulation of granulocyte differentiation / single strand break repair / (E3-independent) E2 ubiquitin-conjugating enzyme / cullin-RING-type E3 NEDD8 transferase / NEDD8 transferase activity / cullin-RING ubiquitin ligase complex / positive regulation by virus of viral protein levels in host cell / regulation of DNA damage checkpoint / cellular response to chemical stress / spindle assembly involved in female meiosis / Cul7-RING ubiquitin ligase complex / ubiquitin-dependent protein catabolic process via the C-end degron rule pathway / regulation of nucleotide-excision repair / epigenetic programming in the zygotic pronuclei / regulation of proteolysis / double-strand break repair via classical nonhomologous end joining / Loss of Function of FBXW7 in Cancer and NOTCH1 Signaling / Cul4-RING E3 ubiquitin ligase complex / UV-damage excision repair / positive regulation of protein autoubiquitination / protein neddylation / NEDD8 ligase activity / biological process involved in interaction with symbiont / Cul5-RING ubiquitin ligase complex / negative regulation of response to oxidative stress / ubiquitin-ubiquitin ligase activity / regulation of mitotic cell cycle phase transition / WD40-repeat domain binding / SCF ubiquitin ligase complex / Cul2-RING ubiquitin ligase complex / Cul4A-RING E3 ubiquitin ligase complex / Cul4B-RING E3 ubiquitin ligase complex / negative regulation of type I interferon production / SCF-dependent proteasomal ubiquitin-dependent protein catabolic process / E2 ubiquitin-conjugating enzyme / ubiquitin ligase complex scaffold activity / Cul3-RING ubiquitin ligase complex / Prolactin receptor signaling / negative regulation of reproductive process / negative regulation of developmental process / RNA polymerase II complex binding / TGF-beta receptor signaling activates SMADs / protein monoubiquitination / ubiquitin conjugating enzyme activity / viral release from host cell / cullin family protein binding / somatic stem cell population maintenance / hemopoiesis / response to X-ray / ectopic germ cell programmed cell death / anatomical structure morphogenesis / transcription-coupled nucleotide-excision repair / positive regulation of G1/S transition of mitotic cell cycle / protein autoubiquitination / proteasomal protein catabolic process / positive regulation of viral genome replication / protein K48-linked ubiquitination / Nuclear events stimulated by ALK signaling in cancer / response to UV / positive regulation of gluconeogenesis / Maturation of protein E / Maturation of protein E / ER Quality Control Compartment (ERQC) / Myoclonic epilepsy of Lafora / IRAK2 mediated activation of TAK1 complex / Alpha-protein kinase 1 signaling pathway / FLT3 signaling by CBL mutants / Prevention of phagosomal-lysosomal fusion / IRAK1 recruits IKK complex / IRAK1 recruits IKK complex upon TLR7/8 or 9 stimulation / Glycogen synthesis / IRAK2 mediated activation of TAK1 complex upon TLR7/8 or 9 stimulation / TICAM1,TRAF6-dependent induction of TAK1 complex / Regulation of TBK1, IKKε (IKBKE)-mediated activation of IRF3, IRF7 / Regulation of TBK1, IKKε-mediated activation of IRF3, IRF7 upon TLR3 ligation / Membrane binding and targetting of GAG proteins / Endosomal Sorting Complex Required For Transport (ESCRT) / Negative regulation of FLT3 / Constitutive Signaling by NOTCH1 HD Domain Mutants / PTK6 Regulates RTKs and Their Effectors AKT1 and DOK1 / TICAM1-dependent activation of IRF3/IRF7 / NOTCH2 Activation and Transmission of Signal to the Nucleus / Regulation of FZD by ubiquitination / APC/C:Cdc20 mediated degradation of Cyclin B / p75NTR recruits signalling complexes / VLDLR internalisation and degradation / positive regulation of TORC1 signaling / post-translational protein modification / Downregulation of ERBB4 signaling / intrinsic apoptotic signaling pathway / TRAF6-mediated induction of TAK1 complex within TLR4 complex / TRAF6 mediated IRF7 activation in TLR7/8 or 9 signaling / APC-Cdc20 mediated degradation of Nek2A / Regulation of innate immune responses to cytosolic DNA / regulation of cellular response to insulin stimulus / positive regulation of DNA repair / InlA-mediated entry of Listeria monocytogenes into host cells / NF-kB is activated and signals survival
Similarity search - Function
UV-stimulated scaffold protein A / : / : / Uncharacterized conserved protein (DUF2043) / UVSSA N-terminal domain / DNA excision repair protein Rad28/ERCC8/Ckn1/ATCSA-1 / Nedd8-like ubiquitin / ENTH/VHS / Zinc finger, RING-H2-type / RING-H2 zinc finger domain ...UV-stimulated scaffold protein A / : / : / Uncharacterized conserved protein (DUF2043) / UVSSA N-terminal domain / DNA excision repair protein Rad28/ERCC8/Ckn1/ATCSA-1 / Nedd8-like ubiquitin / ENTH/VHS / Zinc finger, RING-H2-type / RING-H2 zinc finger domain / Cullin protein neddylation domain / : / Cullin, conserved site / Cullin family signature. / Cullin repeat-like-containing domain superfamily / Cullin protein, neddylation domain / Cullin / Cullin protein neddylation domain / Cullin / Cullin, N-terminal / Cullin homology domain / Cullin homology domain superfamily / Cullin family / Cullin family profile. / Cleavage/polyadenylation specificity factor, A subunit, N-terminal / Mono-functional DNA-alkylating methyl methanesulfonate N-term / Cleavage/polyadenylation specificity factor, A subunit, C-terminal / : / CPSF A subunit region / Ubiquitin-conjugating enzyme, active site / Ubiquitin-conjugating (UBC) active site signature. / Ubiquitin-conjugating enzyme E2 / Ubiquitin-conjugating enzyme / Ubiquitin-conjugating (UBC) core domain profile. / Ubiquitin-conjugating enzyme E2, catalytic domain homologues / Ubiquitin-conjugating enzyme/RWD-like / Zinc finger RING-type profile. / Zinc finger, RING-type / : / Ubiquitin domain signature. / Ubiquitin conserved site / Ubiquitin domain / Ubiquitin family / Ubiquitin homologues / Ubiquitin domain profile. / Ubiquitin-like domain / Zinc finger, RING/FYVE/PHD-type / G-protein beta WD-40 repeat / Ubiquitin-like domain superfamily / Winged helix DNA-binding domain superfamily / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / Winged helix-like DNA-binding domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
Ubiquitin-conjugating enzyme E2 D2 / Polyubiquitin-C / E3 ubiquitin-protein ligase RBX1 / DNA excision repair protein ERCC-8 / Cullin-4A / NEDD8 / DNA damage-binding protein 1 / UV-stimulated scaffold protein A
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.4 Å
AuthorsKokic, G. / Cramer, P.
Funding support Germany, European Union, 2items
OrganizationGrant numberCountry
German Research Foundation (DFG)EXC 2067/1-390729940 Germany
European Research Council (ERC)Advanced Investigator Grant CHROMATRANS (grant agreement No. 882357)European Union
CitationJournal: Nat Struct Mol Biol / Year: 2024
Title: Structural basis for RNA polymerase II ubiquitylation and inactivation in transcription-coupled repair.
Authors: Goran Kokic / George Yakoub / Diana van den Heuvel / Annelotte P Wondergem / Paula J van der Meer / Yana van der Weegen / Aleksandar Chernev / Isaac Fianu / Thornton J Fokkens / Sonja Lorenz ...Authors: Goran Kokic / George Yakoub / Diana van den Heuvel / Annelotte P Wondergem / Paula J van der Meer / Yana van der Weegen / Aleksandar Chernev / Isaac Fianu / Thornton J Fokkens / Sonja Lorenz / Henning Urlaub / Patrick Cramer / Martijn S Luijsterburg /
Abstract: During transcription-coupled DNA repair (TCR), RNA polymerase II (Pol II) transitions from a transcriptionally active state to an arrested state that allows for removal of DNA lesions. This ...During transcription-coupled DNA repair (TCR), RNA polymerase II (Pol II) transitions from a transcriptionally active state to an arrested state that allows for removal of DNA lesions. This transition requires site-specific ubiquitylation of Pol II by the CRL4 ubiquitin ligase, a process that is facilitated by ELOF1 in an unknown way. Using cryogenic electron microscopy, biochemical assays and cell biology approaches, we found that ELOF1 serves as an adaptor to stably position UVSSA and CRL4 on arrested Pol II, leading to ligase neddylation and activation of Pol II ubiquitylation. In the presence of ELOF1, a transcription factor IIS (TFIIS)-like element in UVSSA gets ordered and extends through the Pol II pore, thus preventing reactivation of Pol II by TFIIS. Our results provide the structural basis for Pol II ubiquitylation and inactivation in TCR.
History
DepositionSep 16, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Sep 4, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
D: ubiquitin-conjugating enzyme E2 D2 isoform X3
N: NEDD8
R: E3 ubiquitin-protein ligase RBX1
U: Ubiquitin
a: DNA excision repair protein ERCC-8
c: UV-stimulated scaffold protein A
d: DNA damage-binding protein 1
e: Cullin-4A
hetero molecules


Theoretical massNumber of molelcules
Total (without water)386,13311
Polymers385,9378
Non-polymers1963
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 8 types, 8 molecules DNRUacde

#1: Protein ubiquitin-conjugating enzyme E2 D2 isoform X3


Mass: 16755.227 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: UBE2D2 / Production host: Escherichia coli (E. coli) / References: UniProt: A0A6P5C4T4
#2: Protein NEDD8 / Neddylin / Neural precursor cell expressed developmentally down-regulated protein 8 / NEDD-8 / ...Neddylin / Neural precursor cell expressed developmentally down-regulated protein 8 / NEDD-8 / Ubiquitin-like protein Nedd8


Mass: 8573.978 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: NEDD8 / Production host: Escherichia coli (E. coli) / References: UniProt: Q15843
#3: Protein E3 ubiquitin-protein ligase RBX1 / E3 ubiquitin-protein transferase RBX1 / Protein ZYP / RING finger protein 75 / RING-box protein 1 / ...E3 ubiquitin-protein transferase RBX1 / Protein ZYP / RING finger protein 75 / RING-box protein 1 / Rbx1 / Regulator of cullins 1 / ROC1


Mass: 12289.977 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: RBX1, RNF75, ROC1 / Production host: Trichoplusia ni (cabbage looper)
References: UniProt: P62877, RING-type E3 ubiquitin transferase, cullin-RING-type E3 NEDD8 transferase
#4: Protein Ubiquitin / Polyubiquitin-C


Mass: 8576.831 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: UBC / Production host: Escherichia coli (E. coli) / References: UniProt: P0CG48
#5: Protein DNA excision repair protein ERCC-8 / Cockayne syndrome WD repeat protein CSA


Mass: 44107.160 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: ERCC8, CKN1, CSA / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q13216
#6: Protein UV-stimulated scaffold protein A


Mass: 80721.680 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: UVSSA, KIAA1530 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q2YD98
#7: Protein DNA damage-binding protein 1 / DDB p127 subunit / DNA damage-binding protein a / DDBa / Damage-specific DNA-binding protein 1 / ...DDB p127 subunit / DNA damage-binding protein a / DDBa / Damage-specific DNA-binding protein 1 / HBV X-associated protein 1 / XAP-1 / UV-damaged DNA-binding factor / UV-damaged DNA-binding protein 1 / UV-DDB 1 / XPE-binding factor / XPE-BF / Xeroderma pigmentosum group E-complementing protein / XPCe


Mass: 127097.469 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DDB1, XAP1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q16531
#8: Protein Cullin-4A / CUL-4A


Mass: 87814.297 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: CUL4A / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q13619

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Non-polymers , 1 types, 3 molecules

#9: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Zn

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Details

Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: C(N)RL4CSA-UVSSA-E2-ubiquitin complex. / Type: COMPLEX / Entity ID: #1-#8 / Source: RECOMBINANT
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

MicroscopyModel: TFS GLACIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 3550 nm / Nominal defocus min: 400 nm
Image recordingElectron dose: 40.2 e/Å2 / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.18.2_3874: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 4.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 67365 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00521825
ELECTRON MICROSCOPYf_angle_d0.68129480
ELECTRON MICROSCOPYf_dihedral_angle_d6.4212894
ELECTRON MICROSCOPYf_chiral_restr0.0443316
ELECTRON MICROSCOPYf_plane_restr0.0053789

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