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- EMDB-19779: in-situ subtomogram average of C. elegans centrioles in centrosomes -

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Basic information

Entry
Database: EMDB / ID: EMD-19779
Titlein-situ subtomogram average of C. elegans centrioles in centrosomes
Map dataFull EM map
Sample
  • Cell: Dissociated C. elegans embryonic cells
Keywordscentriole / microtubule / 13-protofilaments / cryo-ET / cryo-FIB / C. elegans / interphase / mitosis / centrosome / CELL CYCLE
Biological speciesCaenorhabditis elegans (invertebrata)
Methodsubtomogram averaging / cryo EM / Resolution: 44.0 Å
AuthorsTollervey F / Rios MU / Zagoriy I / Woodruff JB / Mahamid J
Funding supportEuropean Union, 1 items
OrganizationGrant numberCountry
European Research Council (ERC)760067European Union
CitationJournal: bioRxiv / Year: 2024
Title: Native molecular architectures of centrosomes in embryos.
Authors: Fergus Tollervey / Manolo U Rios / Evgenia Zagoriy / Jeffrey B Woodruff / Julia Mahamid /
Abstract: Centrosomes organize microtubules that are essential for mitotic divisions in animal cells. They consist of centrioles surrounded by Pericentriolar Material (PCM). Questions related to mechanisms of ...Centrosomes organize microtubules that are essential for mitotic divisions in animal cells. They consist of centrioles surrounded by Pericentriolar Material (PCM). Questions related to mechanisms of centriole assembly, PCM organization, and microtubule formation remain unanswered, in part due to limited availability of molecular-resolution structural analyses . Here, we use cryo-electron tomography to visualize centrosomes across the cell cycle in cells isolated from embryos. We describe a pseudo-timeline of centriole assembly and identify distinct structural features including a cartwheel in daughter centrioles, and incomplete microtubule doublets surrounded by a star-shaped density in mother centrioles. We find that centriole and PCM microtubules differ in protofilament number (13 versus 11) indicating distinct nucleation mechanisms. This difference could be explained by atypical γ-tubulin ring complexes with 11-fold symmetry identified at the minus ends of short PCM microtubules. We further characterize a porous and disordered network that forms the interconnected PCM. Thus, our work builds a three-dimensional structural atlas that helps explain how centrosomes assemble, grow, and achieve function.
History
DepositionMar 3, 2024-
Header (metadata) releaseApr 24, 2024-
Map releaseApr 24, 2024-
UpdateApr 24, 2024-
Current statusApr 24, 2024Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_19779.png

Downloads & links

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Map

FileDownload / File: emd_19779.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFull EM map
Projections & slices

Image control

Size
Brightness
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Others
AxesZ (Sec.)Y (Row.)X (Col.)
6.74 Å/pix.
x 180 pix.
= 1213.272 Å		6.74 Å/pix.
x 180 pix.
= 1213.272 Å		6.74 Å/pix.
x 180 pix.
= 1213.272 Å

Surface

Projections

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Slices (1/2)

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Images are generated by Spider.

Voxel sizeX=Y=Z: 6.7404 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 2.21
Minimum - Maximum-6.608727 - 7.6268005
Average (Standard dev.)0.01017788 (±0.9010375)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 1213.272 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_19779_msk_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map even

Fileemd_19779_half_map_1.map
AnnotationHalf map even
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map odd

Fileemd_19779_half_map_2.map
AnnotationHalf map odd
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Dissociated C. elegans embryonic cells

EntireName: Dissociated C. elegans embryonic cells
Components
  • Cell: Dissociated C. elegans embryonic cells

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Supramolecule #1: Dissociated C. elegans embryonic cells

SupramoleculeName: Dissociated C. elegans embryonic cells / type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Caenorhabditis elegans (invertebrata) / Tissue: Embryo

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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: 7.4
GridModel: Quantifoil / Material: GOLD / Mesh: 200 / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 45 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.037 kPa
VitrificationCryogen name: ETHANE

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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 / Cs: 2.7 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 2.5 µm
Specialist opticsPhase plate: VOLTA PHASE PLATE / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Detector mode: COUNTING / Average electron dose: 120.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

ExtractionNumber tomograms: 10 / Number images used: 800 / Reference model: featureless tube
Final angle assignmentType: NOT APPLICABLE / Software - Name: Dynamo (ver. 1.1.520)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 44.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: Dynamo (ver. 1.1.520) / Number subtomograms used: 800
FSC plot (resolution estimation)

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