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Yorodumi- EMDB-10409: In situ cryo-electron tomogram from Chlamydomonas reinhardtii of ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-10409 | |||||||||
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Title | In situ cryo-electron tomogram from Chlamydomonas reinhardtii of a proteasome cluster at the endoplasmic reticulum | |||||||||
Map data | In situ cryo-electron tomogram from Chlamydomonas reinhardtii, showing a cluster of proteasomes at the ER membrane. | |||||||||
Sample |
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Biological species | Chlamydomonas reinhardtii (plant) | |||||||||
Method | electron tomography / cryo EM | |||||||||
Authors | Albert S / Schaffer M / Baumeister W / Engel BD | |||||||||
Citation | Journal: Proc Natl Acad Sci U S A / Year: 2020 Title: Direct visualization of degradation microcompartments at the ER membrane. Authors: Sahradha Albert / Wojciech Wietrzynski / Chia-Wei Lee / Miroslava Schaffer / Florian Beck / Jan M Schuller / Patrice A Salomé / Jürgen M Plitzko / Wolfgang Baumeister / Benjamin D Engel / Abstract: To promote the biochemical reactions of life, cells can compartmentalize molecular interaction partners together within separated non-membrane-bound regions. It is unknown whether this strategy is ...To promote the biochemical reactions of life, cells can compartmentalize molecular interaction partners together within separated non-membrane-bound regions. It is unknown whether this strategy is used to facilitate protein degradation at specific locations within the cell. Leveraging in situ cryo-electron tomography to image the native molecular landscape of the unicellular alga , we discovered that the cytosolic protein degradation machinery is concentrated within ∼200-nm foci that contact specialized patches of endoplasmic reticulum (ER) membrane away from the ER-Golgi interface. These non-membrane-bound microcompartments exclude ribosomes and consist of a core of densely clustered 26S proteasomes surrounded by a loose cloud of Cdc48. Active proteasomes in the microcompartments directly engage with putative substrate at the ER membrane, a function canonically assigned to Cdc48. Live-cell fluorescence microscopy revealed that the proteasome clusters are dynamic, with frequent assembly and fusion events. We propose that the microcompartments perform ER-associated degradation, colocalizing the degradation machinery at specific ER hot spots to enable efficient protein quality control. #1: Journal: To Be Published Title: Direct visualization of degradation microcompartments at the ER membrane Authors: Albert S / Wietrzynski W / Lee CW / Schaffer M / Beck F / Schuller JM / Salome PA / Plitzko JM / Baumeister W / Engel BD | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_10409.map.gz | 337 MB | EMDB map data format | |
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Header (meta data) | emd-10409-v30.xml emd-10409.xml | 12.8 KB 12.8 KB | Display Display | EMDB header |
Images | emd_10409.png | 185.5 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-10409 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-10409 | HTTPS FTP |
-Validation report
Summary document | emd_10409_validation.pdf.gz | 217 KB | Display | EMDB validaton report |
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Full document | emd_10409_full_validation.pdf.gz | 216.1 KB | Display | |
Data in XML | emd_10409_validation.xml.gz | 3.9 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10409 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-10409 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_10409.map.gz / Format: CCP4 / Size: 762.2 MB / Type: IMAGE STORED AS SIGNED INTEGER (2 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | In situ cryo-electron tomogram from Chlamydomonas reinhardtii, showing a cluster of proteasomes at the ER membrane. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 13.68 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : Whole Chlamydomonas cells
Entire | Name: Whole Chlamydomonas cells |
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Components |
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-Supramolecule #1: Whole Chlamydomonas cells
Supramolecule | Name: Whole Chlamydomonas cells / type: cell / ID: 1 / Parent: 0 Details: Grown suspended in TAP media, with normal atmosphere aeration and constant light |
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Source (natural) | Organism: Chlamydomonas reinhardtii (plant) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | electron tomography |
Aggregation state | cell |
-Sample preparation
Buffer | pH: 7 / Details: TAP media |
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Grid | Model: Quantifoil R2/1 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE-PROPANE / Instrument: FEI VITROBOT MARK IV / Details: blotted from back for 10 seconds before plunging. |
Sectioning | Focused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 kV / Focused ion beam - Current: 0.03 nA / Focused ion beam - Duration: 2000 sec. / Focused ion beam - Temperature: 95 K / Focused ion beam - Initial thickness: 1000 nm / Focused ion beam - Final thickness: 150 nm Focused ion beam - Details: See https://bio-protocol.org/e1575 for detailed procedure.. The value given for _emd_sectioning_focused_ion_beam.instrument is FEI Quanta FIB. This is not in a list of ...Focused ion beam - Details: See https://bio-protocol.org/e1575 for detailed procedure.. The value given for _emd_sectioning_focused_ion_beam.instrument is FEI Quanta FIB. This is not in a list of allowed values set(['DB235', 'OTHER']) so OTHER is written into the XML file. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Specialist optics | Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV |
Details | Bidirectional tilt scheme, separated at 0 degrees |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average exposure time: 1.5 sec. / Average electron dose: 100.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Calibrated defocus max: 5.6 µm / Calibrated defocus min: 4.6 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 5.0 µm / Nominal magnification: 42000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Algorithm: BACK PROJECTION / Software - Name: IMOD / Number images used: 60 |
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CTF correction | Software - Name: IMOD |