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Yorodumi- EMDB-3967: In situ cryo-electron tomogram from Chlamydomonas reinhardtii of ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-3967 | |||||||||
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Title | In situ cryo-electron tomogram from Chlamydomonas reinhardtii of the cellular environment around the nuclear envelope | |||||||||
Map data | Cryo-electron tomogram of Chlamydomonas reinhardtii nuclear membrane | |||||||||
Sample |
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Biological species | Chlamydomonas reinhardtii (plant) | |||||||||
Method | electron tomography / cryo EM | |||||||||
Authors | Albert S / Schaffer M / Beck F / Mosalaganti S / Asano S / Thomas HF / Plitzko J / Beck M / Baumeister W / Engel BD | |||||||||
Citation | Journal: Proc Natl Acad Sci U S A / Year: 2017 Title: Proteasomes tether to two distinct sites at the nuclear pore complex. Authors: Sahradha Albert / Miroslava Schaffer / Florian Beck / Shyamal Mosalaganti / Shoh Asano / Henry F Thomas / Jürgen M Plitzko / Martin Beck / Wolfgang Baumeister / Benjamin D Engel / Abstract: The partitioning of cellular components between the nucleus and cytoplasm is the defining feature of eukaryotic life. The nuclear pore complex (NPC) selectively gates the transport of macromolecules ...The partitioning of cellular components between the nucleus and cytoplasm is the defining feature of eukaryotic life. The nuclear pore complex (NPC) selectively gates the transport of macromolecules between these compartments, but it is unknown whether surveillance mechanisms exist to reinforce this function. By leveraging in situ cryo-electron tomography to image the native cellular environment of , we observed that nuclear 26S proteasomes crowd around NPCs. Through a combination of subtomogram averaging and nanometer-precision localization, we identified two classes of proteasomes tethered via their Rpn9 subunits to two specific NPC locations: binding sites on the NPC basket that reflect its eightfold symmetry and more abundant binding sites at the inner nuclear membrane that encircle the NPC. These basket-tethered and membrane-tethered proteasomes, which have similar substrate-processing state frequencies as proteasomes elsewhere in the cell, are ideally positioned to regulate transcription and perform quality control of both soluble and membrane proteins transiting the NPC. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_3967.map.gz | 340 MB | EMDB map data format | |
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Header (meta data) | emd-3967-v30.xml emd-3967.xml | 14.3 KB 14.3 KB | Display Display | EMDB header |
Images | emd_3967.png | 164.6 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-3967 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-3967 | HTTPS FTP |
-Validation report
Summary document | emd_3967_validation.pdf.gz | 221.1 KB | Display | EMDB validaton report |
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Full document | emd_3967_full_validation.pdf.gz | 220.2 KB | Display | |
Data in XML | emd_3967_validation.xml.gz | 4.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-3967 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-3967 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_3967.map.gz / Format: CCP4 / Size: 762.2 MB / Type: IMAGE STORED AS SIGNED INTEGER (2 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Cryo-electron tomogram of Chlamydomonas reinhardtii nuclear membrane | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. generated in cubic-lattice coordinate | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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) / Strain: mat3-4 |
-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 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR |
Vitrification | Cryogen name: ETHANE-PROPANE / Chamber humidity: 90 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK IV Details: Blotted for 10 seconds with 10 blot force before plunging.. |
Details | The cells were frozen onto grids, then thinned using cryo-focused ion beam milling. |
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 - Dose rate: 2 / Focused ion beam - Duration: 2400 sec. / Focused ion beam - Temperature: 80 K / Focused ion beam - Initial thickness: 5000 / Focused ion beam - Final thickness: 100 Focused ion beam - Details: Starting curent: 0.5 nA, Final current: 0.03 nA. The value given for _emd_sectioning_focused_ion_beam.instrument is FEI Scios DB-FIB. This is not in a list of allowed ...Focused ion beam - Details: Starting curent: 0.5 nA, Final current: 0.03 nA. The value given for _emd_sectioning_focused_ion_beam.instrument is FEI Scios DB-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 - Lower energy threshold: 0 eV / Energy filter - Upper energy threshold: 20 eV |
Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 3838 pixel / Digitization - Dimensions - Height: 3710 pixel / Average exposure time: 1.5 sec. / Average electron dose: 1.5 e/Å2 Details: Images were collected in movie mode at 12 frames per second |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 6.0 µm / Nominal defocus min: 4.0 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 / Details: This is a bin4 (twice binned) tomogram. / Number images used: 61 |
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CTF correction | Software - Name: IMOD |