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- PDB-8tl6: Cryo-EM structure of DDB1deltaB-DDA1-DCAF5 -

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Basic information

Entry
Database: PDB / ID: 8tl6
TitleCryo-EM structure of DDB1deltaB-DDA1-DCAF5
Components
  • DDB1- and CUL4-associated factor 5
  • DET1- and DDB1-associated protein 1
  • DNA damage-binding protein 1
KeywordsLIGASE / E3 ligase / protein degradation / cryo-EM / WD40
Function / homology
Function and homology information


negative regulation of fatty acid biosynthetic process / positive regulation by virus of viral protein levels in host cell / epigenetic programming in the zygotic pronuclei / spindle assembly involved in female meiosis / Cul4-RING E3 ubiquitin ligase complex / UV-damage excision repair / biological process involved in interaction with symbiont / regulation of mitotic cell cycle phase transition / WD40-repeat domain binding / Cul4A-RING E3 ubiquitin ligase complex ...negative regulation of fatty acid biosynthetic process / positive regulation by virus of viral protein levels in host cell / epigenetic programming in the zygotic pronuclei / spindle assembly involved in female meiosis / Cul4-RING E3 ubiquitin ligase complex / UV-damage excision repair / biological process involved in interaction with symbiont / regulation of mitotic cell cycle phase transition / WD40-repeat domain binding / Cul4A-RING E3 ubiquitin ligase complex / Cul4B-RING E3 ubiquitin ligase complex / ubiquitin ligase complex scaffold activity / negative regulation of reproductive process / negative regulation of developmental process / cullin family protein binding / viral release from host cell / ectopic germ cell programmed cell death / positive regulation of viral genome replication / positive regulation of gluconeogenesis / proteasomal protein catabolic process / nucleotide-excision repair / Recognition of DNA damage by PCNA-containing replication complex / DNA Damage Recognition in GG-NER / regulation of circadian rhythm / Dual Incision in GG-NER / Transcription-Coupled Nucleotide Excision Repair (TC-NER) / Formation of TC-NER Pre-Incision Complex / Wnt signaling pathway / Formation of Incision Complex in GG-NER / protein polyubiquitination / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / positive regulation of protein catabolic process / cellular response to UV / rhythmic process / protein-macromolecule adaptor activity / positive regulation of proteasomal ubiquitin-dependent protein catabolic process / site of double-strand break / Neddylation / ubiquitin-dependent protein catabolic process / proteasome-mediated ubiquitin-dependent protein catabolic process / chromosome, telomeric region / damaged DNA binding / protein ubiquitination / DNA repair / apoptotic process / DNA damage response / protein-containing complex binding / nucleolus / negative regulation of apoptotic process / protein-containing complex / DNA binding / extracellular space / extracellular exosome / nucleoplasm / nucleus / cytoplasm
Similarity search - Function
DDB1- and CUL4-associated factor 8-like / DET1- and DDB1-associated protein 1, N-terminal / DET1- and DDB1-associated protein 1 / Det1 complexing ubiquitin ligase / 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 / WD domain, G-beta repeat / WD40 repeats ...DDB1- and CUL4-associated factor 8-like / DET1- and DDB1-associated protein 1, N-terminal / DET1- and DDB1-associated protein 1 / Det1 complexing ubiquitin ligase / 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 / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily
Similarity search - Domain/homology
DNA damage-binding protein 1 / DDB1- and CUL4-associated factor 5 / DET1- and DDB1-associated protein 1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.63 Å
AuthorsYue, H. / Hunkeler, M. / Roy Burman, S.S. / Fischer, E.S.
Funding support United States, 1items
OrganizationGrant numberCountry
The Mark Foundation6331505 United States
CitationJournal: Nature / Year: 2024
Title: Targeting DCAF5 suppresses SMARCB1-mutant cancer by stabilizing SWI/SNF.
Authors: Sandi Radko-Juettner / Hong Yue / Jacquelyn A Myers / Raymond D Carter / Alexis N Robertson / Priya Mittal / Zhexin Zhu / Baranda S Hansen / Katherine A Donovan / Moritz Hunkeler / Wojciech ...Authors: Sandi Radko-Juettner / Hong Yue / Jacquelyn A Myers / Raymond D Carter / Alexis N Robertson / Priya Mittal / Zhexin Zhu / Baranda S Hansen / Katherine A Donovan / Moritz Hunkeler / Wojciech Rosikiewicz / Zhiping Wu / Meghan G McReynolds / Shourya S Roy Burman / Anna M Schmoker / Nada Mageed / Scott A Brown / Robert J Mobley / Janet F Partridge / Elizabeth A Stewart / Shondra M Pruett-Miller / Behnam Nabet / Junmin Peng / Nathanael S Gray / Eric S Fischer / Charles W M Roberts /
Abstract: Whereas oncogenes can potentially be inhibited with small molecules, the loss of tumour suppressors is more common and is problematic because the tumour-suppressor proteins are no longer present to ...Whereas oncogenes can potentially be inhibited with small molecules, the loss of tumour suppressors is more common and is problematic because the tumour-suppressor proteins are no longer present to be targeted. Notable examples include SMARCB1-mutant cancers, which are highly lethal malignancies driven by the inactivation of a subunit of SWI/SNF (also known as BAF) chromatin-remodelling complexes. Here, to generate mechanistic insights into the consequences of SMARCB1 mutation and to identify vulnerabilities, we contributed 14 SMARCB1-mutant cell lines to a near genome-wide CRISPR screen as part of the Cancer Dependency Map Project. We report that the little-studied gene DDB1-CUL4-associated factor 5 (DCAF5) is required for the survival of SMARCB1-mutant cancers. We show that DCAF5 has a quality-control function for SWI/SNF complexes and promotes the degradation of incompletely assembled SWI/SNF complexes in the absence of SMARCB1. After depletion of DCAF5, SMARCB1-deficient SWI/SNF complexes reaccumulate, bind to target loci and restore SWI/SNF-mediated gene expression to levels that are sufficient to reverse the cancer state, including in vivo. Consequently, cancer results not from the loss of SMARCB1 function per se, but rather from DCAF5-mediated degradation of SWI/SNF complexes. These data indicate that therapeutic targeting of ubiquitin-mediated quality-control factors may effectively reverse the malignant state of some cancers driven by disruption of tumour suppressor complexes.
History
DepositionJul 26, 2023Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 3, 2024Provider: repository / Type: Initial release
Revision 1.1Apr 17, 2024Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: DNA damage-binding protein 1
B: DDB1- and CUL4-associated factor 5
E: DET1- and DDB1-associated protein 1


Theoretical massNumber of molelcules
Total (without water)219,9163
Polymers219,9163
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: 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: 96425.586 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
#2: Protein DDB1- and CUL4-associated factor 5 / Breakpoint cluster region protein 2 / BCRP2 / WD repeat-containing protein 22


Mass: 108948.688 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DCAF5, BCRG2, KIAA1824, WDR22 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q96JK2
#3: Protein DET1- and DDB1-associated protein 1 / Placenta cross-immune reaction antigen 1 / PCIA-1


Mass: 14542.154 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: DDA1, C19orf58, PCIA1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q9BW61

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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: Complex of DDB1deltaB-DDA1-DCAF5 / Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightValue: 0.22 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Trichoplusia ni (cabbage looper)
Buffer solutionpH: 7.5 / Details: 25mM HEPES, pH 7.4, 200mM NaCl, 4mM TCEP
Buffer component
IDConc.NameFormulaBuffer-ID
1200 mMsodium chlorideNaClSodium chloride1
225 mM2-[4-(2-Hydroxyethyl)piperazin-1-yl]ethane-1-sulfonic acidHEPES1
SpecimenConc.: 0.9 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: LEICA EM GP / Cryogen name: NITROGEN / Humidity: 90 % / Chamber temperature: 283 K

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 36000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 4.494 sec. / Electron dose: 53.349 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 1072 / Details: 50 frames per movie

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Processing

EM software
IDNameVersionCategory
1Topaz0.2.4particle selection
2SerialEMimage acquisition
4cryoSPARCv3.2CTF correction
7UCSF ChimeraX1.4model fitting
8Coot0.9model fitting
10cryoSPARCv3.2initial Euler assignment
11cryoSPARCv3.2final Euler assignment
12cryoSPARCv3.2classification
13cryoSPARCv4.1.23D reconstruction
20PHENIX1.19.2model refinement
CTF correctionDetails: implemented in cryoSPARCv3.2 / Type: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1404938
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 2.63 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 547805 / Algorithm: FOURIER SPACE / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingB value: 97 / Protocol: OTHER / Space: REAL
Atomic model building

3D fitting-ID: 1

IDPDB-IDPdb chain-IDAccession codeChain-IDInitial refinement model-IDSource nameType
16q0r

6q0r

A6q0rA1PDBexperimental model
26q0r

6q0r

E6q0rE1PDBexperimental model
3BRoseTTAFoldin silico model
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 41.71 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00239502
ELECTRON MICROSCOPYf_angle_d0.472312841
ELECTRON MICROSCOPYf_chiral_restr0.04551431
ELECTRON MICROSCOPYf_plane_restr0.00371664
ELECTRON MICROSCOPYf_dihedral_angle_d4.44921266

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