|Entry||Database: EMDB / ID: EMD-20161|
|Title||The central pair apparatus focusing on the C1a-e-c supercomplex extracted from the cryo-electron tomography and subtomographic average of isolated Chlamydomonas fap92 mutant axoneme|
|Sample||The C1a-e-c supercomplex of central pair apparatus averaged from Chlamydomonas fap92 mutant cilia|
|Biological species||Chlamydomonas reinhardtii (plant)|
|Method||subtomogram averaging / cryo EM / Resolution: 22 Å|
|Authors||Fu G / Nicastro D|
|Funding support|| United States, 4 items |
|Citation||Journal: J. Cell Biol. / Year: 2019|
Title: Structural organization of the C1a-e-c supercomplex within the ciliary central apparatus.
Authors: Gang Fu / Lei Zhao / Erin Dymek / Yuqing Hou / Kangkang Song / Nhan Phan / Zhiguo Shang / Elizabeth F Smith / George B Witman / Daniela Nicastro /
Abstract: Nearly all motile cilia contain a central apparatus (CA) composed of two connected singlet microtubules with attached projections that play crucial roles in regulating ciliary motility. Defects in CA ...Nearly all motile cilia contain a central apparatus (CA) composed of two connected singlet microtubules with attached projections that play crucial roles in regulating ciliary motility. Defects in CA assembly usually result in motility-impaired or paralyzed cilia, which in humans causes disease. Despite their importance, the protein composition and functions of the CA projections are largely unknown. Here, we integrated biochemical and genetic approaches with cryo-electron tomography to compare the CA of wild-type with CA mutants. We identified a large (>2 MD) complex, the C1a-e-c supercomplex, that requires the PF16 protein for assembly and contains the CA components FAP76, FAP81, FAP92, and FAP216. We localized these subunits within the supercomplex using nanogold labeling and show that loss of any one of them results in impaired ciliary motility. These data provide insight into the subunit organization and 3D structure of the CA, which is a prerequisite for understanding the molecular mechanisms by which the CA regulates ciliary beating.
|Structure viewer||EM map: |
Downloads & links
|File||Download / File: emd_20161.map.gz / Format: CCP4 / Size: 3.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)|
|Projections & slices|
Images are generated by Spider.
generated in cubic-lattice coordinate
|Voxel size||X=Y=Z: 5.523 Å|
|Symmetry||Space group: 1|
CCP4 map header:
-Entire The C1a-e-c supercomplex of central pair apparatus averaged from ...
|Entire||Name: The C1a-e-c supercomplex of central pair apparatus averaged from Chlamydomonas fap92 mutant cilia|
Number of components: 1
-Component #1: protein, The C1a-e-c supercomplex of central pair apparatus avera...
|Protein||Name: The C1a-e-c supercomplex of central pair apparatus averaged from Chlamydomonas fap92 mutant cilia|
Recombinant expression: No
|Source||Species: Chlamydomonas reinhardtii (plant)|
|Source (natural)||Organelle: cilia|
|Specimen||Specimen state: Cell / Method: cryo EM|
|Sample solution||pH: 7.4|
|Vitrification||Instrument: HOMEMADE PLUNGER / Cryogen name: ETHANE / Temperature: 298 K|
Details: back-side blotting with No.1 Whatman filter for 1.5-2.5 seconds before plunging.
-Electron microscopy imaging
Model: Titan Krios / Image courtesy: FEI Company
|Imaging||Microscope: FEI TITAN KRIOS|
|Electron gun||Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 1.57 e/Å2 / Illumination mode: FLOOD BEAM|
|Lens||Magnification: 26000.0 X (calibrated) / Imaging mode: BRIGHT FIELD / Energy filter: GIF Bioquantum|
|Specimen Holder||Model: FEI TITAN KRIOS AUTOGRID HOLDER|
|Camera||Detector: GATAN K2 SUMMIT (4k x 4k)|
|Processing||Method: subtomogram averaging / Number of subtomograms: 882|
|3D reconstruction||Software: IMOD / Resolution: 22 Å / Resolution method: FSC 0.5 CUT-OFF|
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