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- EMDB-22446: Chlamydomonas reinhardtii radial spoke head and neck (recombinant) -

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Open data


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Basic information

Entry
Database: EMDB / ID: EMD-22446
TitleChlamydomonas reinhardtii radial spoke head and neck (recombinant)
Map data
SampleChlamydomonas reinhardtii radial spoke head and neck
  • (Radial spoke protein ...) x 2
  • (Flagellar radial spoke protein ...) x 10
  • Nucleoside diphosphate kinase 6Nucleoside-diphosphate kinase
  • unknown protein
Function / homology
Function and homology information


radial spoke / positive regulation of cilium-dependent cell motility / cilium movement involved in cell motility / axoneme assembly / motile cilium assembly / motile cilium / Set1C/COMPASS complex / nucleoside-diphosphate kinase / MLL3/4 complex / CTP biosynthetic process ...radial spoke / positive regulation of cilium-dependent cell motility / cilium movement involved in cell motility / axoneme assembly / motile cilium assembly / motile cilium / Set1C/COMPASS complex / nucleoside-diphosphate kinase / MLL3/4 complex / CTP biosynthetic process / UTP biosynthetic process / GTP biosynthetic process / Oxidoreductases / nucleoside diphosphate kinase activity / cilium assembly / axoneme / cilium / calmodulin binding / cytoskeleton / oxidoreductase activity / cytoplasm
IQ motif, EF-hand binding site / NADP-dependent oxidoreductase domain / MORN motif / Nucleoside diphosphate kinase / Radial spoke 3 / Dpy-30 motif / Nucleoside diphosphate kinase, active site / Nucleoside diphosphate kinase-like domain / NADP-dependent oxidoreductase domain superfamily / Nucleoside diphosphate kinase 6 ...IQ motif, EF-hand binding site / NADP-dependent oxidoreductase domain / MORN motif / Nucleoside diphosphate kinase / Radial spoke 3 / Dpy-30 motif / Nucleoside diphosphate kinase, active site / Nucleoside diphosphate kinase-like domain / NADP-dependent oxidoreductase domain superfamily / Nucleoside diphosphate kinase 6 / Sdc1/DPY30 / Radial spokehead-like protein / Nucleoside diphosphate kinase-like domain superfamily
Radial spoke protein 9 / Nucleoside diphosphate kinase 6 / Flagellar radial spoke protein 5 / Flagellar radial spoke protein 3 / Radial spoke protein 10 / Flagellar radial spoke protein 1 / Flagellar radial spoke protein 6 / Flagellar radial spoke protein 4 / Flagellar radial spoke protein 2
Biological speciesChlamydomonas reinhardtii (plant)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.03 Å
AuthorsGrossman-Haham I / Coudray N / Yu Z / Wang F / Zhang N / Bhabha G / Vale RD
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R00GM112982 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM103310 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM118106 United States
CitationJournal: Nat Struct Mol Biol / Year: 2021
Title: Structure of the radial spoke head and insights into its role in mechanoregulation of ciliary beating.
Authors: Iris Grossman-Haham / Nicolas Coudray / Zanlin Yu / Feng Wang / Nan Zhang / Gira Bhabha / Ronald D Vale /
Abstract: Motile cilia power cell locomotion and drive extracellular fluid flow by propagating bending waves from their base to tip. The coordinated bending of cilia requires mechanoregulation by the radial ...Motile cilia power cell locomotion and drive extracellular fluid flow by propagating bending waves from their base to tip. The coordinated bending of cilia requires mechanoregulation by the radial spoke (RS) protein complexes and the microtubule central pair (CP). Despite their importance for ciliary motility across eukaryotes, the molecular function of the RSs is unknown. Here, we reconstituted the Chlamydomonas reinhardtii RS head that abuts the CP and determined its structure using single-particle cryo-EM to 3.1-Å resolution, revealing a flat, negatively charged surface supported by a rigid core of tightly intertwined proteins. Mutations in this core, corresponding to those involved in human ciliopathies, compromised the stability of the recombinant complex, providing a molecular basis for disease. Partially reversing the negative charge on the RS surface impaired motility in C. reinhardtii. We propose that the RS-head architecture is well-suited for mechanoregulation of ciliary beating through physical collisions with the CP.
Validation ReportSummary, Full report, XML, About validation report
History
DepositionAug 12, 2020-
Header (metadata) releaseDec 16, 2020-
Map releaseDec 16, 2020-
UpdateJan 27, 2021-
Current statusJan 27, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.015
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.015
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-7jrj
  • Surface level: 0.015
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_22446.map.gz / Format: CCP4 / Size: 600.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.84 Å/pix.
x 540 pix.
= 450.9 Å
0.84 Å/pix.
x 540 pix.
= 450.9 Å
0.84 Å/pix.
x 540 pix.
= 450.9 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.835 Å
Density
Contour LevelBy AUTHOR: 0.015 / Movie #1: 0.015
Minimum - Maximum-0.035352327 - 0.0930655
Average (Standard dev.)0.0001230415 (±0.0018453321)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions540540540
Spacing540540540
CellA=B=C: 450.9 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.8350.8350.835
M x/y/z540540540
origin x/y/z0.0000.0000.000
length x/y/z450.900450.900450.900
α/β/γ90.00090.00090.000
start NX/NY/NZ1081340
NX/NY/NZ13584349
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS540540540
D min/max/mean-0.0350.0930.000

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Supplemental data

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Additional map: Focused map for the RSP1 protrusion

Fileemd_22446_additional_1.map
AnnotationFocused map for the RSP1 protrusion
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Focused map for the RSP3 protrusion

Fileemd_22446_additional_2.map
AnnotationFocused map for the RSP3 protrusion
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Focused map for the RSP2 protrusion

Fileemd_22446_additional_3.map
AnnotationFocused map for the RSP2 protrusion
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Focused map for the RSP10 protrusion

Fileemd_22446_additional_4.map
AnnotationFocused map for the RSP10 protrusion
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: Focused map for the radial spoke arm region

Fileemd_22446_additional_5.map
AnnotationFocused map for the radial spoke arm region
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Additional map: composite map

Fileemd_22446_additional_6.map
Annotationcomposite map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

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Sample components

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Entire Chlamydomonas reinhardtii radial spoke head and neck

EntireName: Chlamydomonas reinhardtii radial spoke head and neck / Number of components: 15

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Component #1: protein, Chlamydomonas reinhardtii radial spoke head and neck

ProteinName: Chlamydomonas reinhardtii radial spoke head and neck / Recombinant expression: No
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #2: protein, Radial spoke protein 10

ProteinName: Radial spoke protein 10 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 23.553943 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #3: protein, Flagellar radial spoke protein 1

ProteinName: Flagellar radial spoke protein 1 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 87.873445 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #4: protein, Nucleoside diphosphate kinase 6

ProteinName: Nucleoside diphosphate kinase 6Nucleoside-diphosphate kinase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 61.422211 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #5: protein, Flagellar radial spoke protein 2

ProteinName: Flagellar radial spoke protein 2 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 77.445297 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #6: protein, unknown protein

ProteinName: unknown protein / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.358434 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #7: protein, Flagellar radial spoke protein 3

ProteinName: Flagellar radial spoke protein 3 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 39.361293 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #8: protein, Flagellar radial spoke protein 5

ProteinName: Flagellar radial spoke protein 5 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 55.92818 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #9: protein, Radial spoke protein 9

ProteinName: Radial spoke protein 9 / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 29.572348 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #10: protein, Flagellar radial spoke protein 4

ProteinName: Flagellar radial spoke protein 4 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 49.90082 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #11: protein, Flagellar radial spoke protein 6

ProteinName: Flagellar radial spoke protein 6 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 48.884551 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #12: protein, Flagellar radial spoke protein 6

ProteinName: Flagellar radial spoke protein 6 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.528644 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #13: protein, Flagellar radial spoke protein 10

ProteinName: Flagellar radial spoke protein 10 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 0.443539 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #14: protein, Flagellar radial spoke protein 1

ProteinName: Flagellar radial spoke protein 1 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 7.59235 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Component #15: protein, Flagellar radial spoke protein 2

ProteinName: Flagellar radial spoke protein 2 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 5.890252 kDa
SourceSpecies: Chlamydomonas reinhardtii (plant)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm)

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.1 mg/mL / Buffer solution: 30 mM HEPES, 300 mM NaCl, 1 mM TCEP / pH: 7.4
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 295 K / Humidity: 95 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 70 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 105000 X (nominal) / Imaging mode: BRIGHT FIELD / Defocus: 800.0 - 1500.0 nm
Specimen HolderModel: OTHER
CameraDetector: OTHER

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Image acquisition

Image acquisitionNumber of digital images: 7572

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 136659
3D reconstructionSoftware: RELION / Resolution: 3.03 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: rigid body / Refinement space: REAL
Input PDB model: 7JR9
Output model

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