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- EMDB-16871: Subtomogram average of short bridges of the yeast ER-mitochondria... -

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Entry
Database: EMDB / ID: EMD-16871
TitleSubtomogram average of short bridges of the yeast ER-mitochondria encounter structure (ERMES). The population half containing shorter bridge structures was averaged.
Map dataSubtomogram average (STA) map of short ERMES bridge structures. The data set of all bridges was split into halves according to bridge length. The map has been aligned to the STA map of all bridges.
Sample
  • Cell: Budding yeast cells expressing Mdm34-mNeonGreen, a component of the ER-mitochondria encounter structure (ERMES).
KeywordsERMES / lipid transfer protein / membrane contact sites / SMP domains / LIPID TRANSPORT
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsubtomogram averaging / cryo EM / Resolution: 29.0 Å
AuthorsWozny MR / Di Luca A / Morado DR / Picco A / Khaddaj R / Campomanes P / Ivanovic L / Hoffmann PC / Miller EA / Vanni S / Kukulski W
Funding support United Kingdom, Canada, Switzerland, European Union, 7 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom)MC_UP_1201/8 United Kingdom
Medical Research Council (MRC, United Kingdom)MC_UP_1201/10 United Kingdom
Natural Sciences and Engineering Research Council (NSERC, Canada) Canada
Swiss National Science Foundation185544 Switzerland
Swiss National Science Foundation201158 Switzerland
Swiss National Science Foundation194807 Switzerland
European Research Council (ERC)803952European Union
CitationJournal: Nature / Year: 2023
Title: In situ architecture of the ER-mitochondria encounter structure.
Authors: Michael R Wozny / Andrea Di Luca / Dustin R Morado / Andrea Picco / Rasha Khaddaj / Pablo Campomanes / Lazar Ivanović / Patrick C Hoffmann / Elizabeth A Miller / Stefano Vanni / Wanda Kukulski /
Abstract: The endoplasmic reticulum and mitochondria are main hubs of eukaryotic membrane biogenesis that rely on lipid exchange via membrane contact sites, but the underpinning mechanisms remain poorly ...The endoplasmic reticulum and mitochondria are main hubs of eukaryotic membrane biogenesis that rely on lipid exchange via membrane contact sites, but the underpinning mechanisms remain poorly understood. In yeast, tethering and lipid transfer between the two organelles is mediated by the endoplasmic reticulum-mitochondria encounter structure (ERMES), a four-subunit complex of unresolved stoichiometry and architecture. Here we determined the molecular organization of ERMES within Saccharomyces cerevisiae cells using integrative structural biology by combining quantitative live imaging, cryo-correlative microscopy, subtomogram averaging and molecular modelling. We found that ERMES assembles into approximately 25 discrete bridge-like complexes distributed irregularly across a contact site. Each bridge consists of three synaptotagmin-like mitochondrial lipid binding protein domains oriented in a zig-zag arrangement. Our molecular model of ERMES reveals a pathway for lipids. These findings resolve the in situ supramolecular architecture of a major inter-organelle lipid transfer machinery and provide a basis for the mechanistic understanding of lipid fluxes in eukaryotic cells.
History
DepositionMar 19, 2023-
Header (metadata) releaseMar 29, 2023-
Map releaseMar 29, 2023-
UpdateJun 7, 2023-
Current statusJun 7, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_16871.png

Downloads & links

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Map

FileDownload / File: emd_16871.map.gz / Format: CCP4 / Size: 6.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram average (STA) map of short ERMES bridge structures. The data set of all bridges was split into halves according to bridge length. The map has been aligned to the STA map of all bridges.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
5.37 Å/pix.
x 120 pix.
= 644.16 Å		5.37 Å/pix.
x 120 pix.
= 644.16 Å		5.37 Å/pix.
x 120 pix.
= 644.16 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 5.368 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.2
Minimum - Maximum-0.60045236 - 1.0418863
Average (Standard dev.)0.0002155369 (±0.08242928)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions120120120
Spacing120120120
CellA=B=C: 644.16003 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Half map b, masked. Together with masked half...

Fileemd_16871_additional_1.map
AnnotationHalf map b, masked. Together with masked half map a, this was used for resolution estimate.
Projections & Slices
AxesZYX

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Histogram

Histogram (log scale)

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Additional map: Half map a, masked. Together with masked half...

Fileemd_16871_additional_2.map
AnnotationHalf map a, masked. Together with masked half map b, this was used for resolution estimate.
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Additional map: This is the main STA map of short...

Fileemd_16871_additional_3.map
AnnotationThis is the main STA map of short bridges, not aligned to the STA map of all bridges.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map a, unmasked

Fileemd_16871_half_map_1.map
AnnotationHalf map a, unmasked
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map b, unmasked

Fileemd_16871_half_map_2.map
AnnotationHalf map b, unmasked
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Budding yeast cells expressing Mdm34-mNeonGreen, a component of t...

EntireName: Budding yeast cells expressing Mdm34-mNeonGreen, a component of the ER-mitochondria encounter structure (ERMES).
Components
  • Cell: Budding yeast cells expressing Mdm34-mNeonGreen, a component of the ER-mitochondria encounter structure (ERMES).

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Supramolecule #1: Budding yeast cells expressing Mdm34-mNeonGreen, a component of t...

SupramoleculeName: Budding yeast cells expressing Mdm34-mNeonGreen, a component of the ER-mitochondria encounter structure (ERMES).
type: cell / ID: 1 / Parent: 0
Details: Yeast cells were cryo-FIB-milled, imaged by cryo-fluorescence microscopy and electron cryo-tomograms were acquired at localisations of Mdm34-mNeonGreen fluorescent signals.
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: S288C

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 6
Details: Synthetic complete medium without tryptophan, with 2% glucose and 15% dextran
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER
DetailsPlunge-frozen grids with yeast cells were subjected to cryo-FIB milling followed by cryo-fluorescence microscopy prior to cryo-ET.

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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: 6.0 µm / Nominal defocus min: 3.5 µm
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 1.3 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

ExtractionNumber tomograms: 51 / Number images used: 1133
Final angle assignmentType: OTHER
Final reconstructionResolution.type: BY AUTHOR / Resolution: 29.0 Å / Resolution method: FSC 0.143 CUT-OFF
Details: This STA map was obtained using half of the subtomograms that were used to obtain the map of all ERMES bridges (1098 subtomograms). The data set was split according to bridge length; here ...Details: This STA map was obtained using half of the subtomograms that were used to obtain the map of all ERMES bridges (1098 subtomograms). The data set was split according to bridge length; here the half containing shorter bridges was used.
Number subtomograms used: 549
FSC plot (resolution estimation)

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