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- EMDB-12162: Microtubule doublet structure from WT Chlamydomonas as a control ... -

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Basic information

Entry
Database: EMDB / ID: EMD-12162
TitleMicrotubule doublet structure from WT Chlamydomonas as a control for MOT7 mutants
Map dataCilia from WT Chlamydomonas as a control data for mutants, EMD-12160 and EMD-12161
Sample
  • Organelle or cellular component: doublet microtubules from cilia
Biological speciesChlamydomonas reinhardtii (plant)
Methodsubtomogram averaging / cryo EM / Resolution: 25.0 Å
AuthorsObbineni JM / Kutomi O / Yamamoto R / Nakagiri Y / Imai H / Noga A / Zimmermann N / Ishikawa T / Wakabayashi K / Kon T / Inaba K
Funding support Switzerland, Japan, 2 items
OrganizationGrant numberCountry
Swiss National Science FoundationNF310030_192644 Switzerland
Japan Society for the Promotion of Science (JSPS)17H01440 Japan
CitationJournal: Sci Adv / Year: 2021
Title: A dynein-associated photoreceptor protein prevents ciliary acclimation to blue light.
Authors: Osamu Kutomi / Ryosuke Yamamoto / Keiko Hirose / Katsutoshi Mizuno / Yuuhei Nakagiri / Hiroshi Imai / Akira Noga / Jagan Mohan Obbineni / Noemi Zimmermann / Masako Nakajima / Daisuke Shibata ...Authors: Osamu Kutomi / Ryosuke Yamamoto / Keiko Hirose / Katsutoshi Mizuno / Yuuhei Nakagiri / Hiroshi Imai / Akira Noga / Jagan Mohan Obbineni / Noemi Zimmermann / Masako Nakajima / Daisuke Shibata / Misa Shibata / Kogiku Shiba / Masaki Kita / Hideo Kigoshi / Yui Tanaka / Yuya Yamasaki / Yuma Asahina / Chihong Song / Mami Nomura / Mamoru Nomura / Ayako Nakajima / Mia Nakachi / Lixy Yamada / Shiori Nakazawa / Hitoshi Sawada / Kazuyoshi Murata / Kaoru Mitsuoka / Takashi Ishikawa / Ken-Ichi Wakabayashi / Takahide Kon / Kazuo Inaba /
Abstract: Light-responsive regulation of ciliary motility is known to be conducted through modulation of dyneins, but the mechanism is not fully understood. Here, we report a novel subunit of the two-headed ...Light-responsive regulation of ciliary motility is known to be conducted through modulation of dyneins, but the mechanism is not fully understood. Here, we report a novel subunit of the two-headed f/I1 inner arm dynein, named DYBLUP, in animal spermatozoa and a unicellular green alga. This subunit contains a BLUF (sensors of blue light using FAD) domain that appears to directly modulate dynein activity in response to light. DYBLUP (dynein-associated BLUF protein) mediates the connection between the f/I1 motor domain and the tether complex that links the motor to the doublet microtubule. lacking the DYBLUP ortholog shows both positive and negative phototaxis but becomes acclimated and attracted to high-intensity blue light. These results suggest a mechanism to avoid toxic strong light via direct photoregulation of dyneins.
History
DepositionDec 26, 2020-
Header (metadata) releaseMar 17, 2021-
Map releaseMar 17, 2021-
UpdateMar 17, 2021-
Current statusMar 17, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3900
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 3900
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_12162.png

Downloads & links

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Map

FileDownload / File: emd_12162.map.gz / Format: CCP4 / Size: 16.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCilia from WT Chlamydomonas as a control data for mutants, EMD-12160 and EMD-12161
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
8.4 Å/pix.
x 164 pix.
= 1377.6 Å		8.4 Å/pix.
x 164 pix.
= 1377.6 Å		8.4 Å/pix.
x 164 pix.
= 1377.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 8.4 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 3900.0 / Movie #1: 3900
Minimum - Maximum0.0 - 10000.0
Average (Standard dev.)3457.861 (±890.5959)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions164164164
Spacing164164164
CellA=B=C: 1377.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z8.48.48.4
M x/y/z164164164
origin x/y/z0.0000.0000.000
length x/y/z1377.6001377.6001377.600
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS164164164
D min/max/mean0.00010000.0003457.861

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Supplemental data

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Sample components

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Entire : doublet microtubules from cilia

EntireName: doublet microtubules from cilia
Components
  • Organelle or cellular component: doublet microtubules from cilia

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Supramolecule #1: doublet microtubules from cilia

SupramoleculeName: doublet microtubules from cilia / type: organelle_or_cellular_component / ID: 1 / Parent: 0
Details: Average of subtomograms extracted from microtubule doublet parts of cilia
Source (natural)Organism: Chlamydomonas reinhardtii (plant) / Strain: MOT7-BCCP / Organelle: flagella/cilia / Location in cell: flagella/cilia

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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 / Details: TAP medium
GridModel: Quantifoil R3.5/1 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Pretreatment - Type: PLASMA CLEANING
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Instrument: GATAN CRYOPLUNGE 3
Detailscilia isolated from the cell and demembraned

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 25 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 3 / Average exposure time: 1.0 sec. / Average electron dose: 60.0 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 / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 4.0 µm / Nominal magnification: 30000
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 reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 25.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMOD / Number subtomograms used: 3000
ExtractionNumber tomograms: 20 / Number images used: 4000
CTF correctionSoftware - Name: IMOD
Final angle assignmentType: PROJECTION MATCHING

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