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- EMDB-29693: 96-nm doublet microtubule from combined Tetrahymena thermophila s... -

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Basic information

Entry
Database: EMDB / ID: EMD-29693
Title96-nm doublet microtubule from combined Tetrahymena thermophila strains CU428 and K40R
Map datamerged map of 96-nm doublet from Tetrahymena CU428 and K40R combined
Sample
  • Organelle or cellular component: 96-nm repeat unit of microtubule doublet from Tetrahymena thermophila
KeywordsCilia / Axoneme / Doublet Microtubule / Microtubule Inner Protein / STRUCTURAL PROTEIN
Biological speciesTetrahymena thermophila (eukaryote)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.75 Å
AuthorsBlack CS / Kubo S / Yang SK / Bui KH
Funding support Canada, 1 items
OrganizationGrant numberCountry
Canadian Institutes of Health Research (CIHR) Canada
CitationJournal: Nat Commun / Year: 2023
Title: Native doublet microtubules from Tetrahymena thermophila reveal the importance of outer junction proteins.
Authors: Shintaroh Kubo / Corbin S Black / Ewa Joachimiak / Shun Kai Yang / Thibault Legal / Katya Peri / Ahmad Abdelzaher Zaki Khalifa / Avrin Ghanaeian / Caitlyn L McCafferty / Melissa Valente- ...Authors: Shintaroh Kubo / Corbin S Black / Ewa Joachimiak / Shun Kai Yang / Thibault Legal / Katya Peri / Ahmad Abdelzaher Zaki Khalifa / Avrin Ghanaeian / Caitlyn L McCafferty / Melissa Valente-Paterno / Chelsea De Bellis / Phuong M Huynh / Zhe Fan / Edward M Marcotte / Dorota Wloga / Khanh Huy Bui /
Abstract: Cilia are ubiquitous eukaryotic organelles responsible for cellular motility and sensory functions. The ciliary axoneme is a microtubule-based cytoskeleton consisting of two central singlets and nine ...Cilia are ubiquitous eukaryotic organelles responsible for cellular motility and sensory functions. The ciliary axoneme is a microtubule-based cytoskeleton consisting of two central singlets and nine outer doublet microtubules. Cryo-electron microscopy-based studies have revealed a complex network inside the lumen of both tubules composed of microtubule-inner proteins (MIPs). However, the functions of most MIPs remain unknown. Here, we present single-particle cryo-EM-based analyses of the Tetrahymena thermophila native doublet microtubule and identify 42 MIPs. These data shed light on the evolutionarily conserved and diversified roles of MIPs. In addition, we identified MIPs potentially responsible for the assembly and stability of the doublet outer junction. Knockout of the evolutionarily conserved outer junction component CFAP77 moderately diminishes Tetrahymena swimming speed and beat frequency, indicating the important role of CFAP77 and outer junction stability in cilia beating generation and/or regulation.
History
DepositionFeb 7, 2023-
Header (metadata) releaseJun 14, 2023-
Map releaseJun 14, 2023-
UpdateJun 14, 2023-
Current statusJun 14, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

Downloads & links

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Map

FileDownload / File: emd_29693.map.gz / Format: CCP4 / Size: 247.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationmerged map of 96-nm doublet from Tetrahymena CU428 and K40R combined
Voxel sizeX=Y=Z: 1.745 Å
Density
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-0.004379463 - 2.2860177
Average (Standard dev.)0.012709907 (±0.08375859)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions402402402
Spacing402402402
CellA=B=C: 701.49 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_29693_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: half map 1 of 96-nm doublet from Tetrahymena CU428 and K40R combined

Fileemd_29693_half_map_1.map
Annotationhalf map 1 of 96-nm doublet from Tetrahymena CU428 and K40R combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Half map: half map 2 of 96-nm doublet from Tetrahymena CU428 and K40R combined

Fileemd_29693_half_map_2.map
Annotationhalf map 2 of 96-nm doublet from Tetrahymena CU428 and K40R combined
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire : 96-nm repeat unit of microtubule doublet from Tetrahymena thermophila

EntireName: 96-nm repeat unit of microtubule doublet from Tetrahymena thermophila
Components
  • Organelle or cellular component: 96-nm repeat unit of microtubule doublet from Tetrahymena thermophila

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Supramolecule #1: 96-nm repeat unit of microtubule doublet from Tetrahymena thermophila

SupramoleculeName: 96-nm repeat unit of microtubule doublet from Tetrahymena thermophila
type: organelle_or_cellular_component / ID: 1 / Parent: 0
Details: Single Particle Analysis of the 96-nm repeating unit of the axoneme from intact cilia purified from Tetrahymena strains CU428 and K40R
Source (natural)Organism: Tetrahymena thermophila (eukaryote) / Strain: CU428 / Organelle: Cilia
Molecular weightTheoretical: 7.6 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation statecell

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Sample preparation

Concentration2.2 mg/mL
BufferpH: 7.4
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeTFS 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: 3.0 µm / Nominal defocus min: 1.0 µm
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 45.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: EMDB MAP
EMDB ID:
Initial angle assignmentType: NOT APPLICABLE
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.75 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 172223
FSC plot (resolution estimation)

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