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- EMDB-4280: CryoEM structure of INO80core Nucleosome complex in open grappler... -

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Basic information

Entry
Database: EMDB / ID: EMD-4280
TitleCryoEM structure of INO80core Nucleosome complex in open grappler conformation
Map data
Sample
  • Complex: INO80core Nucleosome complex
Function / homology
Function and homology information


RMTs methylate histone arginines / DASH complex / heterochromatin formation => GO:0031507 / protein transport along microtubule to mitotic spindle pole body / mitotic sister chromatid biorientation / Metalloprotease DUBs / UCH proteinases / Ub-specific processing proteases / attachment of spindle microtubules to kinetochore / chromatin => GO:0000785 ...RMTs methylate histone arginines / DASH complex / heterochromatin formation => GO:0031507 / protein transport along microtubule to mitotic spindle pole body / mitotic sister chromatid biorientation / Metalloprotease DUBs / UCH proteinases / Ub-specific processing proteases / attachment of spindle microtubules to kinetochore / chromatin => GO:0000785 / Ino80 complex / attachment of mitotic spindle microtubules to kinetochore / negative regulation of megakaryocyte differentiation / protein localization to CENP-A containing chromatin / ATP-dependent activity, acting on DNA / Chromatin modifying enzymes / Replacement of protamines by nucleosomes in the male pronucleus / CENP-A containing nucleosome / Packaging Of Telomere Ends / Recognition and association of DNA glycosylase with site containing an affected purine / Cleavage of the damaged purine / Deposition of new CENPA-containing nucleosomes at the centromere / Recognition and association of DNA glycosylase with site containing an affected pyrimidine / Cleavage of the damaged pyrimidine / Inhibition of DNA recombination at telomere / Meiotic synapsis / telomere organization / RNA Polymerase I Promoter Opening / Interleukin-7 signaling / SUMOylation of chromatin organization proteins / Assembly of the ORC complex at the origin of replication / DNA methylation / Condensation of Prophase Chromosomes / HCMV Late Events / Chromatin modifications during the maternal to zygotic transition (MZT) / ERCC6 (CSB) and EHMT2 (G9a) positively regulate rRNA expression / SIRT1 negatively regulates rRNA expression / innate immune response in mucosa / PRC2 methylates histones and DNA / helicase activity / Defective pyroptosis / HDACs deacetylate histones / RNA Polymerase I Promoter Escape / Nonhomologous End-Joining (NHEJ) / Transcriptional regulation by small RNAs / Formation of the beta-catenin:TCF transactivating complex / RUNX1 regulates genes involved in megakaryocyte differentiation and platelet function / Activated PKN1 stimulates transcription of AR (androgen receptor) regulated genes KLK2 and KLK3 / G2/M DNA damage checkpoint / NoRC negatively regulates rRNA expression / B-WICH complex positively regulates rRNA expression / HDMs demethylate histones / DNA Damage/Telomere Stress Induced Senescence / Metalloprotease DUBs / mitotic spindle / kinetochore / PKMTs methylate histone lysines / RMTs methylate histone arginines / Meiotic recombination / Pre-NOTCH Transcription and Translation / nucleosome assembly / Activation of anterior HOX genes in hindbrain development during early embryogenesis / HCMV Early Events / Transcriptional regulation of granulopoiesis / structural constituent of chromatin / UCH proteinases / nucleosome / antimicrobial humoral immune response mediated by antimicrobial peptide / E3 ubiquitin ligases ubiquitinate target proteins / Recruitment and ATM-mediated phosphorylation of repair and signaling proteins at DNA double strand breaks / RUNX1 regulates transcription of genes involved in differentiation of HSCs / chromatin organization / Factors involved in megakaryocyte development and platelet production / Processing of DNA double-strand break ends / HATs acetylate histones / antibacterial humoral response / Senescence-Associated Secretory Phenotype (SASP) / Oxidative Stress Induced Senescence / DNA helicase / Estrogen-dependent gene expression / chromosome, telomeric region / Ub-specific processing proteases / defense response to Gram-positive bacterium / chromatin remodeling / Amyloid fiber formation / protein heterodimerization activity / DNA repair / enzyme binding / ATP hydrolysis activity / protein-containing complex / DNA binding / extracellular space / RNA binding / extracellular exosome / extracellular region / nucleoplasm / ATP binding / membrane / identical protein binding / nucleus
Similarity search - Function
DASH complex subunit Dad4 / DASH complex subunit Dad4 / INO80 complex subunit B-like conserved region / INO80 complex, subunit Ies2 / INO80 complex, subunit Ies6 / PAPA-1-like conserved region / PAPA-1 / Vps72/YL1, C-terminal / YL1 nuclear protein C-terminal domain / YL1 nuclear protein C-terminal domain ...DASH complex subunit Dad4 / DASH complex subunit Dad4 / INO80 complex subunit B-like conserved region / INO80 complex, subunit Ies2 / INO80 complex, subunit Ies6 / PAPA-1-like conserved region / PAPA-1 / Vps72/YL1, C-terminal / YL1 nuclear protein C-terminal domain / YL1 nuclear protein C-terminal domain / RuvB-like / RuvB-like, AAA-lid domain / RuvBL1/2, DNA/RNA binding domain / TIP49 P-loop domain / TIP49 AAA-lid domain / TIP49, P-loop domain / Actin / Actin family / Actin / Histone H2B signature. / Histone H2B / Histone H2B / Histone H2A conserved site / Histone H2A signature. / Histone H2A, C-terminal domain / C-terminus of histone H2A / Histone H2A / Histone 2A / Histone H4, conserved site / Histone H4 signature. / Histone H4 / Histone H4 / CENP-T/Histone H4, histone fold / Centromere kinetochore component CENP-T histone fold / TATA box binding protein associated factor / TATA box binding protein associated factor (TAF), histone-like fold domain / Histone H3 signature 1. / Histone H3 signature 2. / Histone H3 / Histone H3/CENP-A / Histone H2A/H2B/H3 / Core histone H2A/H2B/H3/H4 / ATPase, nucleotide binding domain / Histone-fold / ATPases associated with a variety of cellular activities / AAA+ ATPase domain / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
INO80 complex subunit B-like conserved region domain-containing protein / RuvB-like helicase / RuvB-like helicase / Uncharacterized protein / Vps72/YL1 C-terminal domain-containing protein / Histone H2A type 1 / Histone H2A type 1 / Histone H4 / Histone H2B type 1-C/E/F/G/I / Histone H3.2
Similarity search - Component
Biological speciesChaetomium thermophilum var. thermophilum DSM 1495 (fungus)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.62 Å
AuthorsEustermann S / Schall K / Strauss M / Hopfner KP
CitationJournal: Nature / Year: 2018
Title: Structural basis for ATP-dependent chromatin remodelling by the INO80 complex.
Authors: Sebastian Eustermann / Kevin Schall / Dirk Kostrewa / Kristina Lakomek / Mike Strauss / Manuela Moldt / Karl-Peter Hopfner /
Abstract: In the eukaryotic nucleus, DNA is packaged in the form of nucleosomes, each of which comprises about 147 base pairs of DNA wrapped around a histone protein octamer. The position and histone ...In the eukaryotic nucleus, DNA is packaged in the form of nucleosomes, each of which comprises about 147 base pairs of DNA wrapped around a histone protein octamer. The position and histone composition of nucleosomes is governed by ATP-dependent chromatin remodellers such as the 15-subunit INO80 complex . INO80 regulates gene expression, DNA repair and replication by sliding nucleosomes, the exchange of histone H2A.Z with H2A, and the positioning of + 1 and -1 nucleosomes at promoter DNA. The structures and mechanisms of these remodelling reactions are currently unknown. Here we report the cryo-electron microscopy structure of the evolutionarily conserved core of the INO80 complex from the fungus Chaetomium thermophilum bound to a nucleosome, at a global resolution of 4.3 Å and with major parts at 3.7 Å. The INO80 core cradles one entire gyre of the nucleosome through multivalent DNA and histone contacts. An Rvb1/Rvb2 AAA ATPase heterohexamer is an assembly scaffold for the complex and acts as a 'stator' for the motor and nucleosome-gripping subunits. The Swi2/Snf2 ATPase motor binds to nucleosomal DNA at superhelical location -6, unwraps approximately 15 base pairs, disrupts the H2A-DNA contacts and is poised to pump entry DNA into the nucleosome. Arp5 and Ies6 bind superhelical locations -2 and -3 to act as a counter grip for the motor, on the other side of the H2A-H2B dimer. The Arp5 insertion domain forms a grappler element that binds the nucleosome dyad, connects the Arp5 actin-fold and entry DNA over a distance of about 90 Å and packs against histone H2A-H2B near the 'acidic patch'. Our structure together with biochemical data suggests a unified mechanism for nucleosome sliding and histone editing by INO80. The motor is part of a macromolecular ratchet, persistently pumping entry DNA across the H2A-H2B dimer against the Arp5 grip until a large nucleosome translocation step occurs. The transient exposure of H2A-H2B by motor activity as well as differential recognition of H2A.Z and H2A may regulate histone exchange.
History
DepositionJan 31, 2018-
Header (metadata) releaseMar 14, 2018-
Map releaseApr 25, 2018-
UpdateAug 29, 2018-
Current statusAug 29, 2018Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.024
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.024
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4280.png

Downloads & links

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Map

FileDownload / File: emd_4280.map.gz / Format: CCP4 / Size: 149.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.06 Å
Density
Contour LevelBy AUTHOR: 0.024 / Movie #1: 0.024
Minimum - Maximum-0.077622674 - 0.13234276
Average (Standard dev.)-0.000032947533 (±0.004677772)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions340340340
Spacing340340340
CellA=B=C: 360.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.061.061.06
M x/y/z340340340
origin x/y/z0.0000.0000.000
length x/y/z360.400360.400360.400
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS340340340
D min/max/mean-0.0780.132-0.000

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Supplemental data

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Sample components

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Entire : INO80core Nucleosome complex

EntireName: INO80core Nucleosome complex
Components
  • Complex: INO80core Nucleosome complex

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Supramolecule #1: INO80core Nucleosome complex

SupramoleculeName: INO80core Nucleosome complex / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#15
Source (natural)Organism: Chaetomium thermophilum var. thermophilum DSM 1495 (fungus)
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper)
Molecular weightTheoretical: 1 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration1 mg/mL
BufferpH: 8
Details: 20 mM HEPES pH 8, 60 mM KCl, 0.5% glycerol, 0.25 mM CaCl2, 20 uM ZnCl2, 0.25 mM DTT, 0.05% Octyl-beta-glucoside
GridModel: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 281 K / Instrument: LEICA EM GP
DetailsMonodisperse sample: INO80core complex reconstituted with nucleosomal substrate was purified by gelfiltration. Addition of nucleotides or crosslinking was not required

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated defocus min: 1.3 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.5 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-40 / Number grids imaged: 1 / Number real images: 3992 / Average electron dose: 59.6 e/Å2
Details: Images were collected in movie mode with 4 frames per second and 10s total aquisition
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 251692
CTF correctionSoftware - Name: CTFFIND (ver. 4)
Startup modelType of model: OTHER
Details: An initial ab initio model was generated using CryoSPARC
Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: cryoSPARC
Details: An initial ab initio model was generated using CryoSPARC
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2.1.1b)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.62 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1.1b) / Number images used: 17263
FSC plot (resolution estimation)

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