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- EMDB-44391: Cryo-EM structure of the ZBTB9 BTB domain filament -

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Basic information

Entry
Database: EMDB / ID: EMD-44391
TitleCryo-EM structure of the ZBTB9 BTB domain filament
Map dataprimary map
Sample
  • Complex: ZBTB9 BTB domain filament
    • Protein or peptide: Zinc finger and BTB domain-containing protein 9
KeywordsBTB domain / transcription factor / ZBTB protein / transcription
Function / homology
Function and homology information


DNA-binding transcription factor activity, RNA polymerase II-specific / RNA polymerase II cis-regulatory region sequence-specific DNA binding / regulation of transcription by RNA polymerase II / identical protein binding / 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 9
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodhelical reconstruction / cryo EM / Resolution: 8.1 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_44391.png

Downloads & links

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Map

FileDownload / File: emd_44391.map.gz / Format: CCP4 / Size: 75.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationprimary map
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
2.35 Å/pix.
x 270 pix.
= 634.5 Å		2.35 Å/pix.
x 270 pix.
= 634.5 Å		2.35 Å/pix.
x 270 pix.
= 634.5 Å

Surface

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Slices (1/3)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 2.35 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.4
Minimum - Maximum-0.3420041 - 1.0533093
Average (Standard dev.)-0.00026321408 (±0.025548274)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions270270270
Spacing270270270
CellA=B=C: 634.5 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_44391_msk_1.map
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Additional map: sharpened

Fileemd_44391_additional_1.map
Annotationsharpened
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Half map: #2

Fileemd_44391_half_map_1.map
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Half map: #1

Fileemd_44391_half_map_2.map
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Sample components

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Entire : ZBTB9 BTB domain filament

EntireName: ZBTB9 BTB domain filament
Components
  • Complex: ZBTB9 BTB domain filament
    • Protein or peptide: Zinc finger and BTB domain-containing protein 9

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Supramolecule #1: ZBTB9 BTB domain filament

SupramoleculeName: ZBTB9 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 9

MacromoleculeName: Zinc finger and BTB domain-containing protein 9 / type: protein_or_peptide / ID: 1 / Number of copies: 14 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 19.387961 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
SequenceString:
MDWSHPQFEK SAVGLNDIFE AQKIEWHEGG GGSGENLYFQ GGGRNPAPRT IQIEFPQHSS SLLESLNRHR LEGKFCDVSL LVQGRELRA HKAVLAAASP YFHDKLLLGD APRLTLPSVI EADAFEGLLQ LIYSGRLRLP LDALPAHLLV ASGLQMWQVV D QCSEILRE LETSGGGI

UniProtKB: Zinc finger and BTB domain-containing protein 9

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Experimental details

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Structure determination

Methodcryo EM
Processinghelical reconstruction
Aggregation statefilament

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Sample preparation

Concentration0.95 mg/mL
BufferpH: 7.4
Component:
ConcentrationFormulaName
50.0 mMC8H18N2O4S/NaOHHEPES/NaOH pH 7.4
200.0 mMNaClNaCl
0.25 mMC32H58N2O8SCHAPSO
1.0 mMC9H15O6PTCEP

Details: 50 mM HEPES/NaOH pH 7.4, 200 mM NaCl, 0.25 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 TALOS ARCTICA
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Number grids imaged: 1 / Number real images: 1137 / Average exposure time: 4.5 sec. / Average electron dose: 50.1 e/Å2
Details: 1 movie (45 frames) was acquired per hole and stage position.
Electron beamAcceleration voltage: 200 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: 2.0 µm / Nominal defocus min: 0.8 µm / Nominal magnification: 36000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Helical parameters - Δz: 36.1 Å
Applied symmetry - Helical parameters - Δ&Phi: 101.9 °
Applied symmetry - Helical parameters - Axial symmetry: C1 (asymmetric)
Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 8.1 Å / Resolution method: FSC 0.143 CUT-OFF
Details: Asymmetric helical refinement followed by local refinement led to the final reconstruction. The helical symmetry values represent the angular rotation and axial rise for two dimers at the ...Details: Asymmetric helical refinement followed by local refinement led to the final reconstruction. The helical symmetry values represent the angular rotation and axial rise for two dimers at the core of the map, where the local resolution was the highest. These values, however, were not applied during helical refinement as they did not improve the quality of the final reconstruction.
Number images used: 147135
Segment selectionNumber selected: 2022676
Startup modelType of model: OTHER / Details: Ab-initio reconstruction
Final angle assignmentType: MAXIMUM LIKELIHOOD
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: 762
Output model

PDB-9b9v:
Cryo-EM structure of the ZBTB9 BTB domain filament

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