National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)
5R01GM092895-08
United States
Citation
Journal: Nat Struct Mol Biol / Year: 2021 Title: Structural insights into assembly and function of the RSC chromatin remodeling complex. Authors: Richard W Baker / Janice M Reimer / Peter J Carman / Bengi Turegun / Tsutomu Arakawa / Roberto Dominguez / Andres E Leschziner / Abstract: SWI/SNF chromatin remodelers modify the position and spacing of nucleosomes and, in humans, are linked to cancer. To provide insights into the assembly and regulation of this protein family, we ...SWI/SNF chromatin remodelers modify the position and spacing of nucleosomes and, in humans, are linked to cancer. To provide insights into the assembly and regulation of this protein family, we focused on a subcomplex of the Saccharomyces cerevisiae RSC comprising its ATPase (Sth1), the essential actin-related proteins (ARPs) Arp7 and Arp9 and the ARP-binding protein Rtt102. Cryo-EM and biochemical analyses of this subcomplex shows that ARP binding induces a helical conformation in the helicase-SANT-associated (HSA) domain of Sth1. Surprisingly, the ARP module is rotated 120° relative to the full RSC about a pivot point previously identified as a regulatory hub in Sth1, suggesting that large conformational changes are part of Sth1 regulation and RSC assembly. We also show that a conserved interaction between Sth1 and the nucleosome acidic patch enhances remodeling. As some cancer-associated mutations dysregulate rather than inactivate SWI/SNF remodelers, our insights into RSC complex regulation advance a mechanistic understanding of chromatin remodeling in disease states.
History
Deposition
Feb 28, 2020
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Header (metadata) release
Dec 2, 2020
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Map release
Dec 2, 2020
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Update
Jan 27, 2021
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Current status
Jan 27, 2021
Processing site: RCSB / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
Entire : cryo-EM map of Sth1-Arp7-Arp9-Rtt102 in the ADP BeF3 state bound ...
Entire
Name: cryo-EM map of Sth1-Arp7-Arp9-Rtt102 in the ADP BeF3 state bound to the nucleosome with a peeled DNA conformation
Components
Complex: cryo-EM map of Sth1-Arp7-Arp9-Rtt102 in the ADP BeF3 state bound to the nucleosome with a peeled DNA conformation
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Supramolecule #1: cryo-EM map of Sth1-Arp7-Arp9-Rtt102 in the ADP BeF3 state bound ...
Supramolecule
Name: cryo-EM map of Sth1-Arp7-Arp9-Rtt102 in the ADP BeF3 state bound to the nucleosome with a peeled DNA conformation type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#12
Organism: Escherichia coli (E. coli) / Recombinant strain: BL21 Rosetta
Molecular weight
Theoretical: 440 KDa
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Experimental details
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Structure determination
Method
cryo EM
Processing
single particle reconstruction
Aggregation state
particle
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Sample preparation
Concentration
1 mg/mL
Buffer
pH: 7.5
Grid
Model: UltrAuFoil / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Details: Glow discharge for 30 seconds at 25 mAmp
Vitrification
Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: 4 second blot time, blot force 20.
Details
Sample was crosslinked using the GRAFIX protocol
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Electron microscopy
Microscope
FEI TALOS ARCTICA
Image recording
Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average exposure time: 7.0 sec. / Average electron dose: 53.0 e/Å2
Electron beam
Acceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
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