[English] 日本語
Yorodumi
- EMDB-18490: Cryo-electron tomogram of lift-out lamella from cell-derived extr... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-18490
TitleCryo-electron tomogram of lift-out lamella from cell-derived extracellular matrix (example 1)
Map dataTomogram acquired on cryo-lift-out lamella of cell-derived matrix showing intra- and extracellular structures.
Sample
  • Cell: Cryo-lamella prepared by lift-out cryo-FIB milling of cell-derived matrix from human telomerase-immortalized foreskin fibroblasts (TIFFs)
KeywordsExtracellular matrix / cell-derived matrix / collagen / PROTEIN FIBRIL
Biological speciesHomo sapiens (human)
Methodelectron tomography / cryo EM
AuthorsZens B / Faessler F / Hansen J / Hauschild R / Datler J / Hodirnau V-V / Zheden V / Alanko J / Sixt MK / Schur FKM
Funding support Austria, United States, European Union, 4 items
OrganizationGrant numberCountry
Austrian Science FundP33367 Austria
Austrian Science FundE435 Austria
Chan Zuckerberg InitiativeDAF2021-234754 United States
European Research Council (ERC)724373European Union
CitationJournal: J Cell Biol / Year: 2024
Title: Lift-out cryo-FIBSEM and cryo-ET reveal the ultrastructural landscape of extracellular matrix.
Authors: Bettina Zens / Florian Fäßler / Jesse M Hansen / Robert Hauschild / Julia Datler / Victor-Valentin Hodirnau / Vanessa Zheden / Jonna Alanko / Michael Sixt / Florian K M Schur /
Abstract: The extracellular matrix (ECM) serves as a scaffold for cells and plays an essential role in regulating numerous cellular processes, including cell migration and proliferation. Due to limitations in ...The extracellular matrix (ECM) serves as a scaffold for cells and plays an essential role in regulating numerous cellular processes, including cell migration and proliferation. Due to limitations in specimen preparation for conventional room-temperature electron microscopy, we lack structural knowledge on how ECM components are secreted, remodeled, and interact with surrounding cells. We have developed a 3D-ECM platform compatible with sample thinning by cryo-focused ion beam milling, the lift-out extraction procedure, and cryo-electron tomography. Our workflow implements cell-derived matrices (CDMs) grown on EM grids, resulting in a versatile tool closely mimicking ECM environments. This allows us to visualize ECM for the first time in its hydrated, native context. Our data reveal an intricate network of extracellular fibers, their positioning relative to matrix-secreting cells, and previously unresolved structural entities. Our workflow and results add to the structural atlas of the ECM, providing novel insights into its secretion and assembly.
History
DepositionSep 20, 2023-
Header (metadata) releaseMar 13, 2024-
Map releaseMar 13, 2024-
UpdateApr 3, 2024-
Current statusApr 3, 2024Processing site: PDBe / Status: Released

-
Structure visualization

Supplemental images

emd_18490.png

Downloads & links

-
Map

FileDownload / File: emd_18490.map.gz / Format: CCP4 / Size: 350.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTomogram acquired on cryo-lift-out lamella of cell-derived matrix showing intra- and extracellular structures.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
17.1 Å/pix.
x 250 pix.
= 4274. Å		17.1 Å/pix.
x 720 pix.
= 12309.12 Å		17.1 Å/pix.
x 510 pix.
= 8718.96 Å

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

generated in cubic-lattice coordinate

Voxel sizeX=Y=Z: 17.096 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-6.8400183 - 4.1225986
Average (Standard dev.)-0.43350792 (±0.28823164)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions720510250
Spacing510720250
CellA: 8718.96 Å / B: 12309.12 Å / C: 4274.0 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Sample components

-
Entire : Cryo-lamella prepared by lift-out cryo-FIB milling of cell-derive...

EntireName: Cryo-lamella prepared by lift-out cryo-FIB milling of cell-derived matrix from human telomerase-immortalized foreskin fibroblasts (TIFFs)
Components
  • Cell: Cryo-lamella prepared by lift-out cryo-FIB milling of cell-derived matrix from human telomerase-immortalized foreskin fibroblasts (TIFFs)

-
Supramolecule #1: Cryo-lamella prepared by lift-out cryo-FIB milling of cell-derive...

SupramoleculeName: Cryo-lamella prepared by lift-out cryo-FIB milling of cell-derived matrix from human telomerase-immortalized foreskin fibroblasts (TIFFs)
type: cell / ID: 1 / Parent: 0
Details: Cell-derived matrix was generated by growing TIFF cells on EM grids over 14 days of growth time.
Source (natural)Organism: Homo sapiens (human) / Organ: Skin / Tissue: Foreskin

-
Experimental details

-
Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statecell

-
Sample preparation

BufferpH: 7.4 / Component - Concentration: 10.0 Percent / Component - Name: Dextran / Details: 10% Dextran in 0.1M Phosphate Buffer, pH 7.4
GridModel: Quantifoil R2/2 / Material: GOLD / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 120 sec. / Pretreatment - Atmosphere: AIR / Details: ELMO Glow Discharge Unit
VitrificationCryogen name: NITROGEN
DetailsCell-derived matrix obtained from TIFF-cells
High pressure freezingInstrument: OTHER
Details: High pressure freezing carriers were 3.0mm in diameter and 0.5mm thick. One carrier was flat, the second carrier had a central cavity of 2.0mm diameter and 0.02mm depth. Carriers were coated ...Details: High pressure freezing carriers were 3.0mm in diameter and 0.5mm thick. One carrier was flat, the second carrier had a central cavity of 2.0mm diameter and 0.02mm depth. Carriers were coated in 1-hexadecene prior to high pressure freezing.. The value given for _em_high_pressure_freezing.instrument is Bal-tec HPM010. This is not in a list of allowed values {'BAL-TEC HPM 010', 'EMS-002 RAPID IMMERSION FREEZER', 'LEICA EM PACT', 'LEICA EM PACT2', 'LEICA EM HPM100', 'OTHER'} so OTHER is written into the XML file.
Cryo protectant10% Dextran (high MW)
SectioningFocused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 / Focused ion beam - Current: 0.03 / Focused ion beam - Duration: 3600 / Focused ion beam - Temperature: 80 K / Focused ion beam - Initial thickness: 1000 / Focused ion beam - Final thickness: 250
Focused ion beam - Details: A block of sample was extracted from the sample bulk by cryo-lift out FIB milling, with a thickness of 5000-8000nm. It was attached to a second grid, where it was then ...Focused ion beam - Details: A block of sample was extracted from the sample bulk by cryo-lift out FIB milling, with a thickness of 5000-8000nm. It was attached to a second grid, where it was then milled down to 200-250nm thickness step by step. Lamellae were milled in consecutive steps with 1nA (down to 3000nm thickness), 0.5nA (2000nm thickness), 0.1nA (1000nm), 50pA (500nm) and 30pA (200nm). See associated publication for details.. The value given for _em_focused_ion_beam.instrument is TFS Aquilos II. This is not in a list of allowed values {'OTHER', 'DB235'} so OTHER is written into the XML file.

-
Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Average exposure time: 0.5 sec. / Average electron dose: 2.137 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated magnification: 42000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 8.0 µm / Nominal defocus min: 8.0 µm / Nominal magnification: 42000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

-
Image processing

Final reconstructionAlgorithm: BACK PROJECTION
Details: Reconstructed with AreTomo (version 1.3.4., Feb22) and processed with IsoNet (version 0.2).
Number images used: 61

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more