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- EMDB-1919: Visualizing branched actin filaments in lamellipodia by electron ... -

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Basic information

Entry
Database: EMDB / ID: EMD-1919
TitleVisualizing branched actin filaments in lamellipodia by electron tomography
Map dataTomogram of a lamellipodium of a 3T3 cell spread on polylysine
Sample
  • Sample: Tomogram of a lamellipodium of a 3T3 cell
  • Organelle or cellular component: Actin
Keywordsactin / branching / Arp2/3 / lamellipodium
Biological speciesMus musculus (house mouse)
Methodelectron tomography / negative staining
AuthorsSmall JV / Winkler C / Vinzenz M / Schmeiser C
CitationJournal: Nat Cell Biol / Year: 2010
Title: Electron tomography reveals unbranched networks of actin filaments in lamellipodia.
Authors: Edit Urban / Sonja Jacob / Maria Nemethova / Guenter P Resch / J Victor Small /
Abstract: Eukaryotic cells can initiate movement using the forces exerted by polymerizing actin filaments to extend lamellipodial and filopodial protrusions. In the current model, actin filaments in ...Eukaryotic cells can initiate movement using the forces exerted by polymerizing actin filaments to extend lamellipodial and filopodial protrusions. In the current model, actin filaments in lamellipodia are organized in a branched, dendritic network. We applied electron tomography to vitreously frozen 'live' cells, fixed cells and cytoskeletons, embedded in vitreous ice or in deep-negative stain. In lamellipodia from four cell types, including rapidly migrating fish keratocytes, we found that actin filaments are almost exclusively unbranched. The vast majority of apparent filament junctions proved to be overlapping filaments, rather than branched end-to-side junctions. Analysis of the tomograms revealed that actin filaments terminate at the membrane interface within a zone several hundred nanometres wide at the lamellipodium front, and yielded the first direct measurements of filament densities. Actin filament pairs were also identified as lamellipodium components and bundle precursors. These data provide a new structural basis for understanding actin-driven protrusion during cell migration.
History
DepositionJul 1, 2011-
Header (metadata) releaseJul 8, 2011-
Map releaseJul 8, 2011-
UpdateJul 20, 2016-
Current statusJul 20, 2016Processing site: PDBe / Status: Released

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Structure visualization

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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

EMD-1919.tif

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Map

FileDownload / File: emd_1919.map.gz / Format: CCP4 / Size: 252.3 MB / Type: IMAGE STORED AS SIGNED BYTE
AnnotationTomogram of a lamellipodium of a 3T3 cell spread on polylysine
Voxel sizeX=Y=Z: 7.46 Å
Density
Contour LevelBy AUTHOR: 27.800000000000001
Minimum - Maximum-128.0 - 127.0
Average (Standard dev.)57.786975859999998 (±8.30877113)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-112-114136
Dimensions1792186681
Spacing1792186681
CellA: 13920.36 Å / B: 13368.32 Å / C: 604.26 Å
α=β=γ: 90.0 °

CCP4 map header:

modeenvelope stored as signed bytes (from -128 lowest to 127 highest)
Å/pix. X/Y/Z7.467.467.46
M x/y/z1866179281
origin x/y/z0.0000.0000.000
length x/y/z13920.36013368.320604.260
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-114-112136
NC/NR/NS1866179281
D min/max/mean-128.000127.00057.787

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Supplemental data

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Sample components

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Entire : Tomogram of a lamellipodium of a 3T3 cell

EntireName: Tomogram of a lamellipodium of a 3T3 cell
Components
  • Sample: Tomogram of a lamellipodium of a 3T3 cell
  • Organelle or cellular component: Actin

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Supramolecule #1000: Tomogram of a lamellipodium of a 3T3 cell

SupramoleculeName: Tomogram of a lamellipodium of a 3T3 cell / type: sample / ID: 1000 / Number unique components: 1

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Supramolecule #1: Actin

SupramoleculeName: Actin / type: organelle_or_cellular_component / ID: 1 / Name.synonym: Actin / Oligomeric state: Multimer / Recombinant expression: No / Database: NCBI
Source (natural)Organism: Mus musculus (house mouse) / synonym: House mouse / Cell: NIH 3t3 / Location in cell: Lamellipodium

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Experimental details

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Structure determination

Methodnegative staining
Processingelectron tomography

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Sample preparation

StainingType: NEGATIVE / Details: 4% SST
VitrificationCryogen name: NONE / Instrument: OTHER

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Electron microscopy

MicroscopeFEI POLARA 300
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 31000
Sample stageSpecimen holder: Eucentric / Specimen holder model: PHILIPS ROTATION HOLDER / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 °
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Final reconstructionSoftware - Name: IMOD / Details: double tilt

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