|Entry||Database: EMDB / ID: EMD-6838|
|Title||Nucleosome with ALB1 enhancer DNA sequence|
|Sample||Nucleosome with ALB1-DNA sequence|
|Function / homology|
Function and homology information
negative regulation of megakaryocyte differentiation / CENP-A containing nucleosome assembly / DNA replication-independent nucleosome assembly / telomere capping / telomere organization / DNA replication-dependent nucleosome assembly / nuclear nucleosome / rDNA heterochromatin assembly / negative regulation of gene expression, epigenetic / regulation of gene silencing by miRNA ...negative regulation of megakaryocyte differentiation / CENP-A containing nucleosome assembly / DNA replication-independent nucleosome assembly / telomere capping / telomere organization / DNA replication-dependent nucleosome assembly / nuclear nucleosome / rDNA heterochromatin assembly / negative regulation of gene expression, epigenetic / regulation of gene silencing by miRNA / nuclear chromosome / DNA-templated transcription, initiation / regulation of megakaryocyte differentiation / nucleosome assembly / nucleosome / double-strand break repair via nonhomologous end joining / nuclear chromosome, telomeric region / protein heterodimerization activity / protein domain specific binding / cellular protein metabolic process / host cell nucleus / protein-containing complex / RNA binding / DNA binding / extracellular exosome / membrane / extracellular region / nucleoplasm / nucleus
Histone H4, conserved site / CENP-T/Histone H4, histone fold / Histone-fold / TATA box binding protein associated factor (TAF) / Histone H4
|Biological species||Homo sapiens (human)|
|Method||single particle reconstruction / cryo EM / Resolution: 4 Å|
|Authors||Takizawa Y / Tanaka H / Machida S / Koyama M / Maehara K / Ohkawa Y / Wade PA / Wolf M / Kurumizaka H|
|Citation||Journal: Open Biol / Year: 2018|
Title: Cryo-EM structure of the nucleosome containing the enhancer DNA sequence.
Authors: Yoshimasa Takizawa / Hiroki Tanaka / Shinichi Machida / Masako Koyama / Kazumitsu Maehara / Yasuyuki Ohkawa / Paul A Wade / Matthias Wolf / Hitoshi Kurumizaka /
Abstract: Pioneer transcription factors specifically target their recognition DNA sequences within nucleosomes. FoxA is the pioneer transcription factor that binds to the gene enhancer in liver precursor ...Pioneer transcription factors specifically target their recognition DNA sequences within nucleosomes. FoxA is the pioneer transcription factor that binds to the gene enhancer in liver precursor cells, and is required for liver differentiation in embryos. The enhancer DNA sequence is reportedly incorporated into nucleosomes in cells, although the nucleosome structure containing the targeting sites for FoxA has not been clarified yet. In this study, we determined the nucleosome structure containing the enhancer (N1) sequence, by cryogenic electron microscopy at 4.0 Å resolution. The nucleosome structure with the enhancer DNA is not significantly different from the previously reported nucleosome structure with the Widom 601 DNA. Interestingly, in the nucleosomes, the enhancer DNA contains local flexible regions, as compared to the Widom 601 DNA. Consistently, DNaseI treatments revealed that, in the nucleosome, the enhancer (N1) DNA is more accessible than the Widom 601 sequence. The histones also associated less strongly with the enhancer (N1) DNA than the Widom 601 DNA in the nucleosome. Therefore, the local histone-DNA contacts may be responsible for the enhanced DNA accessibility in the nucleosome with the enhancer DNA.
|Structure viewer||EM map: |
Downloads & links
|File||Download / File: emd_6838.map.gz / Format: CCP4 / Size: 10.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)|
|Projections & slices|
Images are generated by Spider.
|Voxel size||X=Y=Z: 1.4 Å|
|Symmetry||Space group: 1|
CCP4 map header:
-Entire Nucleosome with ALB1-DNA sequence
|Entire||Name: Nucleosome with ALB1-DNA sequence / Number of components: 1|
-Component #1: protein, Nucleosome with ALB1-DNA sequence
|Protein||Name: Nucleosome with ALB1-DNA sequence / Recombinant expression: No|
|Mass||Theoretical: 200 kDa|
|Source||Species: Homo sapiens (human)|
|Source (engineered)||Expression System: Escherichia coli (E. coli)|
Vector: pET15b-H3.1, pET15b-H4, pET15b-H2A, pET15b-H2B, pGEM-T-Easy-(186 base-pair mouse ALB1 enhancer DNA)
|Specimen||Specimen state: Particle / Method: cryo EM|
|Sample solution||Specimen conc.: 0.8 mg/mL / pH: 7.5|
|Support film||Gatan Solarus|
|Vitrification||Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 289 K / Humidity: 100 % / Details: 3 second blot, 2.5uL.|
-Electron microscopy imaging
Model: Talos Arctica / Image courtesy: FEI Company
|Imaging||Microscope: FEI TALOS ARCTICA / Details: nanoprobe, parallel beam illumination|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 80 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Magnification: 73000.0 X (nominal), 100000.0 X (calibrated) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: -1500.0 - -3000.0 nm|
|Specimen Holder||Model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature: (77.0 - 100.0 K)|
|Camera||Detector: FEI FALCON III (4k x 4k)|
|Image acquisition||Number of digital images: 2312 / Sampling size: 14 µm|
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