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- EMDB-15638: Tomogram of lamellipodia -

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Basic information

Entry
Database: EMDB / ID: EMD-15638
TitleTomogram of lamellipodia
Map data
Sample
  • Cell: MEF cell
Keywordsactin network / CELL ADHESION
Biological speciesMus musculus (house mouse)
Methodelectron tomography / cryo EM
AuthorsWen-Lu C / Ohad M
Funding support Switzerland, European Union, 2 items
OrganizationGrant numberCountry
Swiss National Science Foundation Switzerland
European Research Council (ERC)European Union
CitationJournal: Commun Biol / Year: 2022
Title: A network of mixed actin polarity in the leading edge of spreading cells.
Authors: Wen-Lu Chung / Matthias Eibauer / Wenhong Li / Rajaa Boujemaa-Paterski / Benjamin Geiger / Ohad Medalia /
Abstract: Physical interactions of cells with the underlying extracellular matrix (ECM) play key roles in multiple cellular processes. The actin cytoskeleton is a central driver and regulator of cellular ...Physical interactions of cells with the underlying extracellular matrix (ECM) play key roles in multiple cellular processes. The actin cytoskeleton is a central driver and regulator of cellular dynamics, that produces membrane-protrusions such as lamellipodia and filopodia. Here, we examined actin organization in expanding lamellipodia during early stages of cell spreading. To gain insight into the 3D actin organization, we plated fibroblasts on galectin-8 coated EM grids, an ECM protein presents in disease states. We then combined cryo-electron tomography with advanced image processing tools for reconstructing the structure of F-actin in the lamellipodia. This approach enabled us to resolve the polarity and orientation of filaments, and the structure of the Arp2/3 complexes associated with F-actin branches. We show that F-actin in lamellipodial protrusions forms a dense network with three distinct sub-domains. One consists primarily of radial filaments, with their barbed ends pointing towards the membrane, the other is enriched with parallel filaments that run between the radial fibers, in addition to an intermediate sub-domain. Surprisingly, a minor, yet significant (~10%) population of actin filaments, are oriented with their barbed-ends towards the cell center. Our results provide structural insights into F-actin assembly and dynamic reorganization in the leading edge of spreading cells.
History
DepositionAug 22, 2022-
Header (metadata) releaseDec 14, 2022-
Map releaseDec 14, 2022-
UpdateMar 27, 2024-
Current statusMar 27, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_15638.map.gz / Format: CCP4 / Size: 285.4 MB / Type: IMAGE STORED AS SIGNED BYTE
Voxel sizeX=Y=Z: 8.82603 Å
Density
Minimum - Maximum-128.0 - 127.0
Average (Standard dev.)62.916804999999997 (±6.4880075)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin16-16-173
Dimensions901960346
Spacing960901346
CellA: 8472.988 Å / B: 7952.253 Å / C: 3053.8064 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : MEF cell

EntireName: MEF cell
Components
  • Cell: MEF cell

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Supramolecule #1: MEF cell

SupramoleculeName: MEF cell / type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Mus musculus (house mouse)

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statecell

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Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE
SectioningOther: NO SECTIONING
Fiducial markerManufacturer: Aurion / Diameter: 10 nm

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 4.0 µm / Nominal defocus min: 4.0 µm
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Average electron dose: 2.46 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionNumber images used: 41

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