+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-3104 | |||||||||
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Title | The human Nuclear Pore Complex with nucleoporin 358 knockdown | |||||||||
Map data | Reconstruction of human nuclear pore complex after Nup358 knockdown | |||||||||
Sample |
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Keywords | nuclear pore complex / Nup358 knockdown | |||||||||
Function / homology | Function and homology information GATOR2 complex / nephron development / nuclear pore inner ring / Seh1-associated complex / protein localization to nuclear inner membrane / protein exit from endoplasmic reticulum / COPII-coated vesicle budding / nuclear envelope organization / transcription-dependent tethering of RNA polymerase II gene DNA at nuclear periphery / telomere tethering at nuclear periphery ...GATOR2 complex / nephron development / nuclear pore inner ring / Seh1-associated complex / protein localization to nuclear inner membrane / protein exit from endoplasmic reticulum / COPII-coated vesicle budding / nuclear envelope organization / transcription-dependent tethering of RNA polymerase II gene DNA at nuclear periphery / telomere tethering at nuclear periphery / COPII-coated vesicle cargo loading / nuclear pore organization / nuclear pore complex assembly / nuclear pore outer ring / atrial cardiac muscle cell action potential / post-transcriptional tethering of RNA polymerase II gene DNA at nuclear periphery / nuclear pore cytoplasmic filaments / COPII vesicle coat / somite development / Nuclear Pore Complex (NPC) Disassembly / nuclear inclusion body / nuclear pore nuclear basket / paraxial mesoderm development / Transport of Ribonucleoproteins into the Host Nucleus / Regulation of Glucokinase by Glucokinase Regulatory Protein / Defective TPR may confer susceptibility towards thyroid papillary carcinoma (TPC) / Amino acids regulate mTORC1 / miRNA processing / attachment of mitotic spindle microtubules to kinetochore / Transport of the SLBP independent Mature mRNA / Transport of the SLBP Dependant Mature mRNA / NS1 Mediated Effects on Host Pathways / SUMOylation of SUMOylation proteins / Transport of Mature mRNA Derived from an Intronless Transcript / protein-containing complex localization / RNA export from nucleus / positive regulation of mRNA splicing, via spliceosome / structural constituent of nuclear pore / Rev-mediated nuclear export of HIV RNA / Nuclear import of Rev protein / SUMOylation of RNA binding proteins / Transport of Mature mRNA derived from an Intron-Containing Transcript / tRNA processing in the nucleus / NEP/NS2 Interacts with the Cellular Export Machinery / COPII-mediated vesicle transport / Postmitotic nuclear pore complex (NPC) reformation / neural tube development / nucleocytoplasmic transport / poly(A)+ mRNA export from nucleus / lamellipodium assembly / Viral Messenger RNA Synthesis / nuclear localization sequence binding / mitotic metaphase chromosome alignment / female gonad development / macrophage chemotaxis / SUMOylation of ubiquitinylation proteins / Vpr-mediated nuclear import of PICs / cellular response to nutrient levels / SUMOylation of DNA replication proteins / Hydrolases; Acting on peptide bonds (peptidases); Serine endopeptidases / positive regulation of TOR signaling / Regulation of HSF1-mediated heat shock response / mRNA transport / Amplification of signal from unattached kinetochores via a MAD2 inhibitory signal / SUMOylation of DNA damage response and repair proteins / mRNA export from nucleus / nuclear pore / Mitotic Prometaphase / EML4 and NUDC in mitotic spindle formation / negative regulation of TORC1 signaling / positive regulation of TORC1 signaling / Resolution of Sister Chromatid Cohesion / cellular response to amino acid starvation / MHC class II antigen presentation / neurogenesis / nuclear periphery / serine-type peptidase activity / SUMOylation of chromatin organization proteins / HCMV Late Events / chromosome segregation / RHO GTPases Activate Formins / promoter-specific chromatin binding / Antigen Presentation: Folding, assembly and peptide loading of class I MHC / Transcriptional regulation by small RNAs / intracellular protein transport / ER to Golgi transport vesicle membrane / spindle / kinetochore / ISG15 antiviral mechanism / HCMV Early Events / protein import into nucleus / Separation of Sister Chromatids / protein transport / nuclear envelope / snRNP Assembly / nuclear membrane / transcription coactivator activity / nuclear body / nuclear speck / defense response to Gram-positive bacterium Similarity search - Function | |||||||||
Biological species | Homo sapiens (human) | |||||||||
Method | subtomogram averaging / cryo EM / Resolution: 37.5 Å | |||||||||
Authors | von Appen A / Kosinski J / Sparks L / Ori A / DiGuilio A / Vollmer B / Mackmull M / Banterle N / Parca L / Kastritis P ...von Appen A / Kosinski J / Sparks L / Ori A / DiGuilio A / Vollmer B / Mackmull M / Banterle N / Parca L / Kastritis P / Buczak K / Mosalaganti S / Hagen W / Andres-Pons A / Lemke EA / Bork P / Antonin W / Glavy JS / Bui KH / Beck M | |||||||||
Citation | Journal: Nature / Year: 2015 Title: In situ structural analysis of the human nuclear pore complex. Authors: Alexander von Appen / Jan Kosinski / Lenore Sparks / Alessandro Ori / Amanda L DiGuilio / Benjamin Vollmer / Marie-Therese Mackmull / Niccolo Banterle / Luca Parca / Panagiotis Kastritis / ...Authors: Alexander von Appen / Jan Kosinski / Lenore Sparks / Alessandro Ori / Amanda L DiGuilio / Benjamin Vollmer / Marie-Therese Mackmull / Niccolo Banterle / Luca Parca / Panagiotis Kastritis / Katarzyna Buczak / Shyamal Mosalaganti / Wim Hagen / Amparo Andres-Pons / Edward A Lemke / Peer Bork / Wolfram Antonin / Joseph S Glavy / Khanh Huy Bui / Martin Beck / Abstract: Nuclear pore complexes are fundamental components of all eukaryotic cells that mediate nucleocytoplasmic exchange. Determining their 110-megadalton structure imposes a formidable challenge and ...Nuclear pore complexes are fundamental components of all eukaryotic cells that mediate nucleocytoplasmic exchange. Determining their 110-megadalton structure imposes a formidable challenge and requires in situ structural biology approaches. Of approximately 30 nucleoporins (Nups), 15 are structured and form the Y and inner-ring complexes. These two major scaffolding modules assemble in multiple copies into an eight-fold rotationally symmetric structure that fuses the inner and outer nuclear membranes to form a central channel of ~60 nm in diameter. The scaffold is decorated with transport-channel Nups that often contain phenylalanine-repeat sequences and mediate the interaction with cargo complexes. Although the architectural arrangement of parts of the Y complex has been elucidated, it is unclear how exactly it oligomerizes in situ. Here we combine cryo-electron tomography with mass spectrometry, biochemical analysis, perturbation experiments and structural modelling to generate, to our knowledge, the most comprehensive architectural model of the human nuclear pore complex to date. Our data suggest previously unknown protein interfaces across Y complexes and to inner-ring complex members. We show that the transport-channel Nup358 (also known as Ranbp2) has a previously unanticipated role in Y-complex oligomerization. Our findings blur the established boundaries between scaffold and transport-channel Nups. We conclude that, similar to coated vesicles, several copies of the same structural building block--although compositionally identical--engage in different local sets of interactions and conformations. | |||||||||
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_3104.map.gz | 16.5 MB | EMDB map data format | |
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Header (meta data) | emd-3104-v30.xml emd-3104.xml | 9.8 KB 9.8 KB | Display Display | EMDB header |
Images | EMD-3104.png | 141.6 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-3104 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-3104 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_3104.map.gz / Format: CCP4 / Size: 17.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Reconstruction of human nuclear pore complex after Nup358 knockdown | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 8.6 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 : Human nuclear pore complex with nucleoporin 358 knockdown
Entire | Name: Human nuclear pore complex with nucleoporin 358 knockdown |
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Components |
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-Supramolecule #1000: Human nuclear pore complex with nucleoporin 358 knockdown
Supramolecule | Name: Human nuclear pore complex with nucleoporin 358 knockdown type: sample / ID: 1000 / Number unique components: 1 |
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Molecular weight | Experimental: 120 MDa / Theoretical: 120 MDa |
-Macromolecule #1: nuclear pore complex
Macromolecule | Name: nuclear pore complex / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Recombinant expression: No |
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Source (natural) | Organism: Homo sapiens (human) / synonym: Human / Organelle: Nucleus / Location in cell: Nuclear envelope |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | subtomogram averaging |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 / Details: 20mM Tris, 0.2-0.4% Trehalose |
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Grid | Details: 200 mesh Quantifoil Copper Holey Carbon Grid R2/1 |
Vitrification | Cryogen name: ETHANE-PROPANE MIXTURE / Instrument: HOMEMADE PLUNGER |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Calibrated magnification: 34883 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 3.0 µm / Nominal magnification: 19500 |
Specialist optics | Energy filter - Name: GIF2002 / Energy filter - Lower energy threshold: 0.0 eV / Energy filter - Upper energy threshold: 20.0 eV |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Tilt series - Axis1 - Min angle: -44 ° / Tilt series - Axis1 - Max angle: 60 ° |
Date | Sep 20, 2014 |
Image recording | Category: CCD / Film or detector model: GATAN MULTISCAN / Average electron dose: 100 e/Å2 / Bits/pixel: 16 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
Final reconstruction | Applied symmetry - Point group: C8 (8 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 37.5 Å / Resolution method: OTHER / Software - Name: IMOD, TOM, AV3 / Number subtomograms used: 7360 |
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Details | The subtomograms were selected manually and further processed by iterative, symmetry-independent averaging. |