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- EMDB-79047: Microtubule bundles in metaphase RPE1 spindles -

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Basic information

Entry
Database: EMDB / ID: EMD-79047
TitleMicrotubule bundles in metaphase RPE1 spindles
Map dataTomogram of microtubules in a human metaphase mitotic spindle
Sample
  • Cell: Microtubule bundles in metaphase RPE1 cells
Keywordsspindle / microtubule / mitosis / cell division / CELL CYCLE
Biological speciesHomo sapiens (human)
Methodelectron tomography / cryo EM
AuthorsConway W / Bobe D / Fabig G / Zimyanin V / de Marco A / Redemann S
Funding support United States, 1 items
OrganizationGrant numberCountry
Simons Foundation00012463 United States
CitationJournal: bioRxiv / Year: 2026
Title: Eg5 activity and density-driven bundling organize the human metaphase mitotic spindle independently of spindle bipolarity.
Authors: William Conway / Daija Bobe / Vitaly Zimyanin / Gunar Fabig / Alex de Marco / Stefanie Redemann /
Abstract: The mitotic spindle segregates chromosomes through the coordinated actions of microtubules and molecular motors. Classic models propose that microtubules nucleate at spindle poles and grow inward to ...The mitotic spindle segregates chromosomes through the coordinated actions of microtubules and molecular motors. Classic models propose that microtubules nucleate at spindle poles and grow inward to capture chromosomes; however, recent structural studies show that spindles contain short microtubules that do not span the distance between poles and chromosomes. It is unclear how these short microtubules assemble a bipolar spindle. Using cryo-electron tomography to map microtubule polarity in human metaphase spindles, we find that microtubules form locally antiparallel bundles with consistent 8 nm wall-to-wall spacing. We utilized motor perturbations and centriole depletion, which generated motor-active monopolar spindles, to reveal that the kinesin-5 motor Eg5 organizes local antiparallel overlap independently of spindle bipolarity. We found that bundles are organized by density-driven steric interactions rather than motor-mediated crosslinking. These findings support a self-organized, bottom-up model in which local microtubule-motor interactions within dense bundles generate forces that build the bipolar spindle, challenging pole-centric models.
History
DepositionSep 12, 2026-
Header (metadata) releaseSep 30, 2026-
Map releaseSep 30, 2026-
UpdateSep 30, 2026-
Current statusSep 30, 2026Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_79047.png

Downloads & links

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Map

FileDownload / File: emd_79047.map.gz / Format: CCP4 / Size: 528.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationTomogram of microtubules in a human metaphase mitotic spindle
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
26.8 Å/pix.
x 376 pix.
= 10076.8 Å		26.8 Å/pix.
x 720 pix.
= 19296. Å		26.8 Å/pix.
x 512 pix.
= 13721.6 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: 26.8 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-0.15673694 - 0.18557456
Average (Standard dev.)0.0000035650926 (±0.014841738)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions720512376
Spacing512720376
CellA: 13721.6 Å / B: 19296.0 Å / C: 10076.8 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : Microtubule bundles in metaphase RPE1 cells

EntireName: Microtubule bundles in metaphase RPE1 cells
Components
  • Cell: Microtubule bundles in metaphase RPE1 cells

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Supramolecule #1: Microtubule bundles in metaphase RPE1 cells

SupramoleculeName: Microtubule bundles in metaphase RPE1 cells / 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: 7.4 / Details: DMEM
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 310 K / Instrument: OTHER
Details: The value given for _em_vitrification.instrument is LEICA EM GP2. This is not in a list of allowed values {'REICHERT-JUNG PLUNGER', 'OTHER', 'SPT LABTECH CHAMELEON', 'FEI VITROBOT MARK IV', ...Details: The value given for _em_vitrification.instrument is LEICA EM GP2. This is not in a list of allowed values {'REICHERT-JUNG PLUNGER', 'OTHER', 'SPT LABTECH CHAMELEON', 'FEI VITROBOT MARK IV', 'LEICA EM GP', 'FEI VITROBOT MARK III', 'LEICA PLUNGER', 'GATAN CRYOPLUNGE 3', 'FEI VITROBOT MARK I', 'CRYOSOL VITROJET', 'SPOTITON', 'ZEISS PLUNGE FREEZER CRYOBOX', 'LEICA EM CPC', 'HOMEMADE PLUNGER', 'FEI VITROBOT MARK II', 'LEICA KF80', 'EMS-002 RAPID IMMERSION FREEZER'} so OTHER is written into the XML file.
High pressure freezingInstrument: OTHER
Details: High pressure frozen via the Waffle method on 200 mesh carbon grids.. The value given for _em_high_pressure_freezing.instrument is Wohlwend. This is not in a list of allowed values {'LEICA ...Details: High pressure frozen via the Waffle method on 200 mesh carbon grids.. The value given for _em_high_pressure_freezing.instrument is Wohlwend. This is not in a list of allowed values {'LEICA EM PACT', 'LEICA EM PACT2', 'BAL-TEC HPM 010', 'OTHER', 'EMS-002 RAPID IMMERSION FREEZER', 'LEICA EM HPM100'} so OTHER is written into the XML file.
Cryo protectant5% glycerol
SectioningFocused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 / Focused ion beam - Current: 0.05 / Focused ion beam - Duration: 60 / Focused ion beam - Temperature: 79 K / Focused ion beam - Initial thickness: 600 / Focused ion beam - Final thickness: 150
Focused ion beam - Details: Polishing item varied per sample. Lamella were hand polished after overnight automated milling. The value given for _em_focused_ion_beam.instrument is Aquilos 2. This is ...Focused ion beam - Details: Polishing item varied per sample. Lamella were hand polished after overnight automated milling. The value given for _em_focused_ion_beam.instrument is Aquilos 2. This is not in a list of allowed values {'DB235', 'OTHER'} so OTHER is written into the XML file.

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Electron microscopy

MicroscopeTFS KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum
Image recordingFilm or detector model: GATAN K3 BIOCONTINUUM (6k x 4k) / Average exposure time: 1.6 sec. / Average electron dose: 3.1 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 5.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 26000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionSoftware - Name: RELION (ver. 4.0.1) / Number images used: 90832
CTF correctionSoftware:
Namedetails
Warp (ver. 1.10b)Fit/determine CTF
RELION (ver. 4.0.1)Apply CTF correction

Type: PHASE FLIPPING AND AMPLITUDE CORRECTION

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