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- EMDB-45648: Representative cryo electron tomography reconstructions showing c... -

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Basic information

Entry
Database: EMDB / ID: EMD-45648
TitleRepresentative cryo electron tomography reconstructions showing central pair microtubule alignment in a wild type Trypanosoma brucei cell
Map dataa tomogram of the extracted flagellum in a wild type cell
Sample
  • Organelle or cellular component: central pair microtubule organization in wild type cells
KeywordsIntraflagellar transport (IFT) / Arl13b / Arl3 / ODA16 / cilia / flagella / small GTPase / Trypanosoma brucei / ciliopathy / PROTEIN TRANSPORT
Biological speciesTrypanosoma brucei (eukaryote)
Methodelectron tomography / cryo EM / Resolution: 11.02 Å
AuthorsSun SY / Huang Y
Funding support Singapore, 2 items
OrganizationGrant numberCountry
Ministry of Education (MoE, Singapore)MOE2017-T2-2-109 Singapore
Ministry of Education (MoE, Singapore)MOE-T2EP30121-0003 Singapore
CitationJournal: Sci Adv / Year: 2024
Title: ARL3 GTPases facilitate ODA16 unloading from IFT in motile cilia.
Authors: Yameng Huang / Xiaoduo Dong / Stella Y Sun / Teck-Kwang Lim / Qingsong Lin / Cynthia Y He /
Abstract: Eukaryotic cilia and flagella are essential for cell motility and sensory functions. Their biogenesis and maintenance rely on the intraflagellar transport (IFT). Several cargo adapters have been ...Eukaryotic cilia and flagella are essential for cell motility and sensory functions. Their biogenesis and maintenance rely on the intraflagellar transport (IFT). Several cargo adapters have been identified to aid IFT cargo transport, but how ciliary cargos are discharged from the IFT remains largely unknown. During our explorations of small GTPases ARL13 and ARL3 in , we found that ODA16, a known IFT cargo adapter present exclusively in motile cilia, is a specific effector of ARL3. In the cilia, active ARL3 GTPases bind to ODA16 and dissociate ODA16 from the IFT complex. Depletion of ARL3 GTPases stabilizes ODA16 interaction with the IFT, leading to ODA16 accumulation in cilia and defects in axonemal assembly. The interactions between human ODA16 homolog HsDAW1 and ARL GTPases are conserved, and these interactions are altered in HsDAW1 disease variants. These findings revealed a conserved function of ARL GTPases in IFT transport of motile ciliary components, and a mechanism of cargo unloading from the IFT.
History
DepositionJul 9, 2024-
Header (metadata) releaseSep 11, 2024-
Map releaseSep 11, 2024-
UpdateSep 11, 2024-
Current statusSep 11, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45648.png

Downloads & links

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Map

FileDownload / File: emd_45648.map.gz / Format: CCP4 / Size: 1.4 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationa tomogram of the extracted flagellum in a wild type cell
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
11.02 Å/pix.
x 350 pix.
= 3858.4 Å		11.02 Å/pix.
x 1024 pix.
= 11288.576 Å		11.02 Å/pix.
x 1024 pix.
= 11288.576 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: 11.024 Å
Density

Histogram

Histogram (log scale)
Minimum - Maximum-35.294246999999999 - 20.53406
Average (Standard dev.)0.000000003997413 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-512-512-175
Dimensions10241024350
Spacing10241024350
CellA: 11288.576 Å / B: 11288.576 Å / C: 3858.4001 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: Feature annotation of the paraflagellar rod

Fileemd_45648_additional_1.map
AnnotationFeature annotation of the paraflagellar rod
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Feature annotation of the central pair microtubules

Fileemd_45648_additional_2.map
AnnotationFeature annotation of the central pair microtubules
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Feature annotation of the axoneme microtubules

Fileemd_45648_additional_3.map
AnnotationFeature annotation of the axoneme microtubules
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : central pair microtubule organization in wild type cells

EntireName: central pair microtubule organization in wild type cells
Components
  • Organelle or cellular component: central pair microtubule organization in wild type cells

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Supramolecule #1: central pair microtubule organization in wild type cells

SupramoleculeName: central pair microtubule organization in wild type cells
type: organelle_or_cellular_component / ID: 1 / Parent: 0
Details: detergent extracted flagellum from Trypanosoma brucei cells
Source (natural)Organism: Trypanosoma brucei (eukaryote)

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statefilament

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Sample preparation

BufferpH: 7.2
GridModel: Quantifoil / Material: COPPER / Mesh: 200 / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Instrument: LEICA EM GP
SectioningOther: NO SECTIONING
Fiducial marker
ManufacturerDiameter
EMS10 nm
EMS15 nm

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 2.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: OTHER / Imaging mode: DARK FIELD / Cs: 2.7 mm / Nominal defocus max: 1.0 µm / Nominal defocus min: 1.0 µm
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 reconstructionResolution.type: BY AUTHOR / Resolution: 11.02 Å / Resolution method: OTHER / Software - Name: EMAN2 / Number images used: 51

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