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- EMDB-16455: Electron cryo-tomography of HeLa cells expressing Cidec-EGFP -

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Basic information

Entry
Database: EMDB / ID: EMD-16455
TitleElectron cryo-tomography of HeLa cells expressing Cidec-EGFP
Map dataRepresentative tomogram of HeLa cells expressing Cidec-EGFP. Binned by factor 4.
Sample
  • Cell: HeLa cells expressing Cidec-EGFP
Biological speciesHomo sapiens (human)
Methodelectron tomography / cryo EM
AuthorsGaneva I / Lim K / Boulanger J / Hoffmann PC / Muriel O / Borgeaud AC / Hagen WJH / Savage DB / Kukulski W
Funding support United Kingdom, Switzerland, 5 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MC_UP_1201/8 United Kingdom
Swiss National Science Foundation201158 Switzerland
Swiss National Science Foundation185544 Switzerland
Wellcome TrustWT 219417 United Kingdom
Medical Research Council (MRC, United Kingdom)MRC_MC_UU_12012.1 United Kingdom
CitationJournal: Cell Rep / Year: 2023
Title: The architecture of Cidec-mediated interfaces between lipid droplets.
Authors: Iva Ganeva / Koini Lim / Jerome Boulanger / Patrick C Hoffmann / Olivia Muriel / Alicia C Borgeaud / Wim J H Hagen / David B Savage / Wanda Kukulski /
Abstract: Lipid droplets (LDs) are intracellular organelles responsible for storing surplus energy as neutral lipids. Their size and number vary enormously. In white adipocytes, LDs can reach 100 μm in ...Lipid droplets (LDs) are intracellular organelles responsible for storing surplus energy as neutral lipids. Their size and number vary enormously. In white adipocytes, LDs can reach 100 μm in diameter, occupying >90% of the cell. Cidec, which is strictly required for the formation of large LDs, is concentrated at interfaces between adjacent LDs and facilitates directional flux of neutral lipids from the smaller to the larger LD. The mechanism of lipid transfer is unclear, in part because the architecture of interfaces between LDs remains elusive. Here we visualize interfaces between LDs by electron cryo-tomography and analyze the kinetics of lipid transfer by quantitative live fluorescence microscopy. We show that transfer occurs through closely apposed monolayers, is slowed down by increasing the distance between the monolayers, and follows exponential kinetics. Our data corroborate the notion that Cidec facilitates pressure-driven transfer of neutral lipids through two "leaky" monolayers between LDs.
History
DepositionJan 13, 2023-
Header (metadata) releaseFeb 22, 2023-
Map releaseFeb 22, 2023-
UpdateMar 1, 2023-
Current statusMar 1, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16455.png

Downloads & links

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Map

FileDownload / File: emd_16455.map.gz / Format: CCP4 / Size: 42.8 MB / Type: IMAGE STORED AS SIGNED INTEGER (2 BYTES)
AnnotationRepresentative tomogram of HeLa cells expressing Cidec-EGFP. Binned by factor 4.
Voxel sizeX=Y=Z: 29.8 Å
Density
Minimum - Maximum-791.0 - 591.0
Average (Standard dev.)7.090758 (±35.638996)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin8-8-38
Dimensions463480101
Spacing480463101
CellA: 14304.0 Å / B: 13797.399 Å / C: 3009.7998 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : HeLa cells expressing Cidec-EGFP

EntireName: HeLa cells expressing Cidec-EGFP
Components
  • Cell: HeLa cells expressing Cidec-EGFP

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Supramolecule #1: HeLa cells expressing Cidec-EGFP

SupramoleculeName: HeLa cells expressing Cidec-EGFP / type: cell / 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 statecell

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Sample preparation

BufferpH: 7.2
GridModel: Quantifoil R2/2 / Material: GOLD / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER
SectioningFocused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 kV / Focused ion beam - Current: 1 nA / Focused ion beam - Duration: 2000 sec. / Focused ion beam - Temperature: 100 K / Focused ion beam - Initial thickness: 1000 nm / Focused ion beam - Final thickness: 100 nm
Focused ion beam - Details: Rough milling: 30 kV, 1-0.1 nA. Fine milling: 16kV, 23 pA.. The value given for _em_focused_ion_beam.instrument is FEI Scios. This is not in a list of allowed values ...Focused ion beam - Details: Rough milling: 30 kV, 1-0.1 nA. Fine milling: 16kV, 23 pA.. The value given for _em_focused_ion_beam.instrument is FEI Scios. This is not in a list of allowed values {'OTHER', 'DB235'} so OTHER is written into the XML file.

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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: 5.0 µm / Nominal defocus min: 5.0 µm
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 1.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionSoftware - Name: IMOD / Number images used: 105

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