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- EMDB-20064: Three-dimensional architecture of epithelial primary cilia -

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Basic information

Entry
Database: EMDB / ID: EMD-20064
TitleThree-dimensional architecture of epithelial primary cilia
Map dataUCSF Chimera: contour level 89 with "hide dust" (9e 03)
Sample
  • Organelle or cellular component: primary cilium #B
Biological speciesMus musculus (house mouse)
Methodelectron tomography / negative staining
AuthorsSun S / Fisher RL / Bowser SS / Pentecost BT / Sui H
CitationJournal: Proc Natl Acad Sci U S A / Year: 2019
Title: Three-dimensional architecture of epithelial primary cilia.
Authors: Shufeng Sun / Rebecca L Fisher / Samuel S Bowser / Brian T Pentecost / Haixin Sui /
Abstract: We report a complete 3D structural model of typical epithelial primary cilia based on structural maps of full-length primary cilia obtained by serial section electron tomography. Our data demonstrate ...We report a complete 3D structural model of typical epithelial primary cilia based on structural maps of full-length primary cilia obtained by serial section electron tomography. Our data demonstrate the architecture of primary cilia differs extensively from the commonly acknowledged 9+0 paradigm. The axoneme structure is relatively stable but gradually evolves from base to tip with a decreasing number of microtubule complexes (MtCs) and a reducing diameter. The axonemal MtCs are cross-linked by previously unrecognized fibrous protein networks. Such an architecture explains why primary cilia can elastically withstand liquid flow for mechanosensing. The nine axonemal MtCs in a cilium are found to differ significantly in length indicating intraflagellar transport processes in primary cilia may be more complicated than that reported for motile cilia. The 3D maps of microtubule doublet-singlet transitions generally display longitudinal gaps at the inner junction between the A- and B-tubules, which indicates the inner junction protein is a major player in doublet-singlet transitions. In addition, vesicles releasing from kidney primary cilia were observed in the structural maps, supporting that ciliary vesicles budding may serve as ectosomes for cell-cell communication.
History
DepositionApr 5, 2019-
Header (metadata) releaseMay 1, 2019-
Map releaseMay 8, 2019-
UpdateMay 22, 2019-
Current statusMay 22, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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  • Solid view (volume rendering)
  • Imaged by UCSF Chimera
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Movie viewer
Supplemental images

emd_20064.png

Downloads & links

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Map

FileDownload / File: emd_20064.map.gz / Format: CCP4 / Size: 707.6 MB / Type: IMAGE STORED AS SIGNED BYTE
AnnotationUCSF Chimera: contour level 89 with "hide dust" (9e 03)
Voxel sizeX=Y=Z: 19.56 Å
Density
Minimum - Maximum0.0 - 127.0
Average (Standard dev.)80.493610000000004 (±4.0639696)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin484277-61
Dimensions7687681258
Spacing7687681258
CellA: 15022.08 Å / B: 15022.08 Å / C: 24606.479 Å
α=β=γ: 90.0 °

CCP4 map header:

modeenvelope stored as signed bytes (from -128 lowest to 127 highest)
Å/pix. X/Y/Z19.5619.5619.559999205087
M x/y/z7687681258
origin x/y/z0.0000.0000.000
length x/y/z15022.08015022.08024606.479
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ513513513
MAP C/R/S123
start NC/NR/NS277484-61
NC/NR/NS7687681258
D min/max/mean0.000127.00080.494

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Supplemental data

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Sample components

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Entire : primary cilium #B

EntireName: primary cilium #B
Components
  • Organelle or cellular component: primary cilium #B

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Supramolecule #1: primary cilium #B

SupramoleculeName: primary cilium #B / type: organelle_or_cellular_component / ID: 1 / Parent: 0
Details: Primary cilium #B is related to Fig.S3b, S4b, and movie 7.
Source (natural)Organism: Mus musculus (house mouse) / Organ: kidney / Cell: IMCD3

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Experimental details

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Structure determination

Methodnegative staining
Processingelectron tomography
Aggregation statecell

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Sample preparation

BufferpH: 7.4 / Details: PBS
StainingType: POSITIVE / Material: uranyl acetate, lead citrate
Details: 2% uranyl acetate and Reynolds lead citrate solution
Sugar embeddingMaterial: epoxy resin
Details: The samples were infiltrated with a graded series of resins (SPI-PON 812, SPI-CHEM, USA) in acetone over 24 hours. After infiltration with pure resin (3 x within 24 hours), the specimens in ...Details: The samples were infiltrated with a graded series of resins (SPI-PON 812, SPI-CHEM, USA) in acetone over 24 hours. After infiltration with pure resin (3 x within 24 hours), the specimens in resin were polymerized at 60 degrees C for 2 days.
GridDetails: unspecified
SectioningUltramicrotomy - Instrument: UC6 microtome / Ultramicrotomy - Temperature: 298 K / Ultramicrotomy - Final thickness: 120 nm
Fiducial markerManufacturer: Sigma-Aldrich / Diameter: 10 nm

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Electron microscopy

MicroscopeFEI TECNAI 20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: TVIPS TEMCAM-F415 (4k x 4k) / Average electron dose: 6.0 e/Å2

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Image processing

Final reconstructionSoftware - Name: TOMO3D
Details: This SSET work used a series of dual-axis tilt data sets.
Number images used: 6552

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