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- EMDB-13195: Cryo-ET of axons from human cerebral organoids, expressing membra... -

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Basic information

Entry
Database: EMDB / ID: EMD-13195
TitleCryo-ET of axons from human cerebral organoids, expressing membrane-targeted GFP
Map datacryo-ET of axons from human cerebral organoids, expressing fGFP.
Sample
  • Tissue: Axon bundles grown on EM grids from human air-liquid interface cerebral organoid cultures.
Biological speciesHomo sapiens (human)
Methodelectron tomography / cryo EM
AuthorsHoffmann PC / Giandomenico SL / Ganeva I / Wozny MR / Sutcliffe M / Lancaster MA / Kukulski W
Funding support United Kingdom, European Union, 3 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MC_UP_1201/8 United Kingdom
Medical Research Council (MRC, United Kingdom)MC_UP_1201/9 United Kingdom
European Research Council (ERC)ERC STG 757710European Union
CitationJournal: Elife / Year: 2021
Title: Electron cryo-tomography reveals the subcellular architecture of growing axons in human brain organoids.
Authors: Patrick C Hoffmann / Stefano L Giandomenico / Iva Ganeva / Michael R Wozny / Magdalena Sutcliffe / Madeline A Lancaster / Wanda Kukulski /
Abstract: During brain development, axons must extend over great distances in a relatively short amount of time. How the subcellular architecture of the growing axon sustains the requirements for such rapid ...During brain development, axons must extend over great distances in a relatively short amount of time. How the subcellular architecture of the growing axon sustains the requirements for such rapid build-up of cellular constituents has remained elusive. Human axons have been particularly poorly accessible to imaging at high resolution in a near-native context. Here, we present a method that combines cryo-correlative light microscopy and electron tomography with human cerebral organoid technology to visualize growing axon tracts. Our data reveal a wealth of structural details on the arrangement of macromolecules, cytoskeletal components, and organelles in elongating axon shafts. In particular, the intricate shape of the endoplasmic reticulum is consistent with its role in fulfilling the high demand for lipid biosynthesis to support growth. Furthermore, the scarcity of ribosomes within the growing shaft suggests limited translational competence during expansion of this compartment. These findings establish our approach as a powerful resource for investigating the ultrastructure of defined neuronal compartments.
History
DepositionJul 14, 2021-
Header (metadata) releaseNov 3, 2021-
Map releaseNov 3, 2021-
UpdateNov 3, 2021-
Current statusNov 3, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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Supplemental images

emd_13195.png

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Map

FileDownload / File: emd_13195.map.gz / Format: CCP4 / Size: 199.4 MB / Type: IMAGE STORED AS SIGNED BYTE
Annotationcryo-ET of axons from human cerebral organoids, expressing fGFP.
Voxel sizeX=Y=Z: 14.71 Å
Density
Minimum - Maximum-128.0 - 127.0
Average (Standard dev.)9.502569 (±13.699769)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin16-16-134
Dimensions927960235
Spacing960927235
CellA: 14121.6 Å / B: 13636.17 Å / C: 3456.85 Å
α=β=γ: 90.0 °

CCP4 map header:

modeenvelope stored as signed bytes (from -128 lowest to 127 highest)
Å/pix. X/Y/Z14.7114.7114.71
M x/y/z960927235
origin x/y/z0.0000.0000.000
length x/y/z14121.60013636.1703456.850
α/β/γ90.00090.00090.000
start NX/NY/NZ799196
NX/NY/NZ149138117
MAP C/R/S123
start NC/NR/NS-1616-134
NC/NR/NS960927235
D min/max/mean-128.000127.0009.503

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Supplemental data

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Sample components

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Entire : Axon bundles grown on EM grids from human air-liquid interface ce...

EntireName: Axon bundles grown on EM grids from human air-liquid interface cerebral organoid cultures.
Components
  • Tissue: Axon bundles grown on EM grids from human air-liquid interface cerebral organoid cultures.

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Supramolecule #1: Axon bundles grown on EM grids from human air-liquid interface ce...

SupramoleculeName: Axon bundles grown on EM grids from human air-liquid interface cerebral organoid cultures.
type: tissue / ID: 1 / Parent: 0
Source (natural)Organism: Homo sapiens (human)

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statetissue

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Sample preparation

BufferpH: 7
VitrificationCryogen name: ETHANE
SectioningOther: NO SECTIONING

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 1.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionNumber images used: 113

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