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- EMDB-44389: Cryo-EM structure of the ZBTB5 BTB domain filament -

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Basic information

Entry
Database: EMDB / ID: EMD-44389
TitleCryo-EM structure of the ZBTB5 BTB domain filament
Map dataprimary map
Sample
  • Complex: ZBTB5 BTB domain filament
    • Protein or peptide: Zinc finger and BTB domain-containing protein 5
KeywordsBTB domain / transcription factor / ZBTB protein / TRANSCRIPTION
Function / homology
Function and homology information


RNA polymerase II transcription regulatory region sequence-specific DNA binding / DNA-binding transcription repressor activity, RNA polymerase II-specific / DNA-binding transcription factor activity, RNA polymerase II-specific / chromatin / regulation of transcription by RNA polymerase II / negative regulation of transcription by RNA polymerase II / nucleus / metal ion binding
Similarity search - Function
BTB/POZ domain / BTB domain profile. / Broad-Complex, Tramtrack and Bric a brac / BTB/POZ domain / zinc finger / Zinc finger C2H2 type domain profile. / Zinc finger C2H2 superfamily / SKP1/BTB/POZ domain superfamily / Zinc finger C2H2 type domain signature. / Zinc finger C2H2-type
Similarity search - Domain/homology
Zinc finger and BTB domain-containing protein 5
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsPark J / Hunkeler M / Fischer ES
Funding support1 items
OrganizationGrant numberCountry
Not funded
CitationJournal: Mol Cell / Year: 2024
Title: Polymerization of ZBTB transcription factors regulates chromatin occupancy.
Authors: Paul M C Park / Jiho Park / Jared Brown / Moritz Hunkeler / Shourya S Roy Burman / Katherine A Donovan / Hojong Yoon / Radosław P Nowak / Mikołaj Słabicki / Benjamin L Ebert / Eric S Fischer /
Abstract: BCL6, an oncogenic transcription factor (TF), forms polymers in the presence of a small-molecule molecular glue that stabilizes a complementary interface between homodimers of BCL6's broad-complex, ...BCL6, an oncogenic transcription factor (TF), forms polymers in the presence of a small-molecule molecular glue that stabilizes a complementary interface between homodimers of BCL6's broad-complex, tramtrack, and bric-à-brac (BTB) domain. The BTB domains of other proteins, including a large class of TFs, have similar architectures and symmetries, raising the possibility that additional BTB proteins self-assemble into higher-order structures. Here, we surveyed 189 human BTB proteins with a cellular fluorescent reporter assay and identified 18 ZBTB TFs that show evidence of polymerization. Through biochemical and cryoelectron microscopy (cryo-EM) studies, we demonstrate that these ZBTB TFs polymerize into filaments. We found that BTB-domain-mediated polymerization of ZBTB TFs enhances chromatin occupancy within regions containing homotypic clusters of TF binding sites, leading to repression of target genes. Our results reveal a role of higher-order structures in regulating ZBTB TFs and suggest an underappreciated role for TF polymerization in modulating gene expression.
History
DepositionApr 3, 2024-
Header (metadata) releaseJul 24, 2024-
Map releaseJul 24, 2024-
UpdateJul 24, 2024-
Current statusJul 24, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_44389.png

Downloads & links

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Map

FileDownload / File: emd_44389.map.gz / Format: CCP4 / Size: 28.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationprimary map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.35 Å/pix.
x 196 pix.
= 264.6 Å		1.35 Å/pix.
x 196 pix.
= 264.6 Å		1.35 Å/pix.
x 196 pix.
= 264.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.35 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-0.21772067 - 0.54399884
Average (Standard dev.)0.0012841503 (±0.017068109)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions196196196
Spacing196196196
CellA=B=C: 264.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_44389_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_44389_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : ZBTB5 BTB domain filament

EntireName: ZBTB5 BTB domain filament
Components
  • Complex: ZBTB5 BTB domain filament
    • Protein or peptide: Zinc finger and BTB domain-containing protein 5

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Supramolecule #1: ZBTB5 BTB domain filament

SupramoleculeName: ZBTB5 BTB domain filament / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Homo sapiens (human)

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Macromolecule #1: Zinc finger and BTB domain-containing protein 5

MacromoleculeName: Zinc finger and BTB domain-containing protein 5 / type: protein_or_peptide / ID: 1 / Number of copies: 4 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 18.786326 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString:
MDWSHPQFEK SAVGLNDIFE AQKIEWHEGG GGSGENLYFQ GGGRMDFPGH FEQIFQQLNY QRLHGQLCDC VIVVGNRHFK AHRSVLAAC STHFRALFSV AEGDQTMNMI QLDSEVVTAE AFAALIDMMY TSTLMLGESN VMDVLLAASH LHLNSVVKAC K HYLTTRTL

UniProtKB: Zinc finger and BTB domain-containing protein 5

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statefilament

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Sample preparation

Concentration1.00 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
50.0 mMC8H18N2O4S/NaOHHEPES/NaOH pH 7.4
200.0 mMNaClNaCl
1.0 mMC32H58N2O8SCHAPSO
1.0 mMC9H15O6PTCEP

Details: 50 mM HEPES/NaOH pH 7.4, 200 mM NaCl, 1 mM CHAPSO, 1 mM TCEP
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 12 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 60 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.039 kPa
Details: Grids were glow-discharged for 60 s at 15-20 mA and 39 Pa.
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 283 K / Instrument: LEICA EM GP
Details: Grids were vitrified using a Leica EM GP plunge freezer operated at 90% humidity and 10 C with 10 s pre-blot, 3 s blot, 3 s post-blot..
DetailsElution fractions from Strep-tag affinity chromatography were dialyzed overnight against 50 mM HEPES/NaOH pH 7.4, 200 mM NaCl, 1 mM TCEP and concentrated by centrifugation.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Number real images: 10854 / Average exposure time: 2.1 sec. / Average electron dose: 50.7 e/Å2
Details: 2 movies (46 frames) were acquired per hole with 9 holes per stage position.
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 1.2 µm / Nominal magnification: 105000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 6344567
Startup modelType of model: OTHER / Details: Ab-initio reconstruction
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 4.4.1) / Number images used: 1352367
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.4.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 4.4.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelChain - Source name: AlphaFold / Chain - Initial model type: in silico model
DetailsReal-space refinement without local grid search
RefinementSpace: REAL / Protocol: OTHER / Overall B value: 194
Output model

PDB-9b9r:
Cryo-EM structure of the ZBTB5 BTB domain filament

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