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- EMDB-20685: Average of the TZ linker from bovine respiratory cilia -

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Basic information

Entry
Database: EMDB / ID: EMD-20685
TitleAverage of the TZ linker from bovine respiratory cilia
Map dataAverage of the TZ linker from bovine respiratory cilia
Sample
  • Organelle or cellular component: Nascent axoneme from bovine respiratory epithelial cells
Biological speciesBos taurus (domestic cattle)
Methodsubtomogram averaging / cryo EM / Resolution: 33.2 Å
AuthorsGreenan GA / Vale RD / Agard DA
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical SciencesGM118099 United States
National Institutes of Health/National Institute of General Medical SciencesGM118106 United States
CitationJournal: J Cell Biol / Year: 2020
Title: Electron cryotomography of intact motile cilia defines the basal body to axoneme transition.
Authors: Garrett A Greenan / Ronald D Vale / David A Agard /
Abstract: Cells use motile cilia to generate force in the extracellular space. The structure of a cilium can be classified into three subdomains: the intracellular basal body (BB) that templates cilium ...Cells use motile cilia to generate force in the extracellular space. The structure of a cilium can be classified into three subdomains: the intracellular basal body (BB) that templates cilium formation, the extracellular axoneme that generates force, and the transition zone (TZ) that bridges them. While the BB is composed of triplet microtubules (TMTs), the axoneme is composed of doublet microtubules (DMTs), meaning the cilium must convert between different microtubule geometries. Here, we performed electron cryotomography to define this conversion, and our reconstructions reveal identifying structural features of the BB, TZ, and axoneme. Each region is distinct in terms of microtubule number and geometry, microtubule inner proteins, and microtubule linkers. TMT to DMT conversion occurs within the BB, and microtubule geometry changes to axonemal by the end of the TZ, followed by the addition of axoneme-specific components essential for cilium motility. Our results provide the highest-resolution images of the motile cilium to date and reveal how BBs template axonemes.
History
DepositionSep 4, 2019-
Header (metadata) releaseDec 4, 2019-
Map releaseDec 4, 2019-
UpdateDec 4, 2019-
Current statusDec 4, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1.6
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 1.6
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20685.png

Downloads & links

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Map

FileDownload / File: emd_20685.map.gz / Format: CCP4 / Size: 12.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationAverage of the TZ linker from bovine respiratory cilia
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
8.18 Å/pix.
x 150 pix.
= 1227. Å		8.18 Å/pix.
x 150 pix.
= 1227. Å		8.18 Å/pix.
x 150 pix.
= 1227. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 8.18 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 1.6 / Movie #1: 1.6
Minimum - Maximum-19.624691 - 18.979658
Average (Standard dev.)0.013934474 (±0.98903865)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions150150150
Spacing150150150
CellA=B=C: 1227.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z8.188.188.18
M x/y/z150150150
origin x/y/z0.0000.0000.000
length x/y/z1227.0001227.0001227.000
α/β/γ90.00090.00090.000
start NX/NY/NZ111-94150
NX/NY/NZ111123111
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS150150150
D min/max/mean-19.62518.9800.014

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Supplemental data

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Sample components

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Entire : Nascent axoneme from bovine respiratory epithelial cells

EntireName: Nascent axoneme from bovine respiratory epithelial cells
Components
  • Organelle or cellular component: Nascent axoneme from bovine respiratory epithelial cells

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Supramolecule #1: Nascent axoneme from bovine respiratory epithelial cells

SupramoleculeName: Nascent axoneme from bovine respiratory epithelial cells
type: organelle_or_cellular_component / ID: 1 / Parent: 0
Source (natural)Organism: Bos taurus (domestic cattle) / Organelle: cilium

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statefilament

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Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV / Details: 15 second blot.

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Electron microscopy

MicroscopeFEI POLARA 300
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 0.8 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company

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Image processing

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 33.2 Å / Resolution method: FSC 0.143 CUT-OFF / Number subtomograms used: 824
ExtractionNumber tomograms: 21 / Number images used: 9450
Final angle assignmentType: MAXIMUM LIKELIHOOD

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