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- EMDB-29013: 96nm doublet microtubule repeat from wild type mouse sperm -

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Basic information

Entry
Database: EMDB / ID: EMD-29013
Title96nm doublet microtubule repeat from wild type mouse sperm
Map data96nm doublet microtubule repeat from WT mouse sperm
Sample
  • Cell: Mouse sperm
Keywords96 nm doublet microtubule from mouse sperm / STRUCTURAL PROTEIN
Biological speciesMus (mice)
Methodsubtomogram averaging / cryo EM / Resolution: 34.0 Å
AuthorsHwang JY / Chai P / Nawaz S
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/Eunice Kennedy Shriver National Institute of Child Health & Human Development (NIH/NICHD)R01HD096745 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM142959 United States
CitationJournal: Elife / Year: 2023
Title: LRRC23 truncation impairs radial spoke 3 head assembly and sperm motility underlying male infertility.
Authors: Jae Yeon Hwang / Pengxin Chai / Shoaib Nawaz / Jungmin Choi / Francesc Lopez-Giraldez / Shabir Hussain / Kaya Bilguvar / Shrikant Mane / Richard P Lifton / Wasim Ahmad / Kai Zhang / Jean-Ju Chung /
Abstract: Radial spokes (RS) are T-shaped multiprotein complexes on the axonemal microtubules. Repeated RS1, RS2, and RS3 couple the central pair to modulate ciliary and flagellar motility. Despite the cell ...Radial spokes (RS) are T-shaped multiprotein complexes on the axonemal microtubules. Repeated RS1, RS2, and RS3 couple the central pair to modulate ciliary and flagellar motility. Despite the cell type specificity of RS3 substructures, their molecular components remain largely unknown. Here, we report that a leucine-rich repeat-containing protein, LRRC23, is an RS3 head component essential for its head assembly and flagellar motility in mammalian spermatozoa. From infertile male patients with defective sperm motility, we identified a splice site variant of . A mutant mouse model mimicking this variant produces a truncated LRRC23 at the C-terminus that fails to localize to the sperm tail, causing male infertility due to defective sperm motility. LRRC23 was previously proposed to be an ortholog of the RS stalk protein RSP15. However, we found that purified recombinant LRRC23 interacts with an RS head protein RSPH9, which is abolished by the C-terminal truncation. Evolutionary and structural comparison also shows that LRRC34, not LRRC23, is the RSP15 ortholog. Cryo-electron tomography clearly revealed that the absence of the RS3 head and the sperm-specific RS2-RS3 bridge structure in LRRC23 mutant spermatozoa. Our study provides new insights into the structure and function of RS3 in mammalian spermatozoa and the molecular pathogenicity of LRRC23 underlying reduced sperm motility in infertile human males.
History
DepositionDec 4, 2022-
Header (metadata) releaseNov 1, 2023-
Map releaseNov 1, 2023-
UpdateDec 27, 2023-
Current statusDec 27, 2023Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_29013.png

Downloads & links

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Map

FileDownload / File: emd_29013.map.gz / Format: CCP4 / Size: 15.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotation96nm doublet microtubule repeat from WT mouse sperm
Voxel sizeX=Y=Z: 10.2 Å
Density
Contour LevelBy AUTHOR: 0.15
Minimum - Maximum-0.97795314 - 1.4890715
Average (Standard dev.)0.0029622489 (±0.121874586)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions160160160
Spacing160160160
CellA=B=C: 1632.0 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: unmasked 96nm doublet microtubule repeat from WT mouse sperm

Fileemd_29013_additional_1.map
Annotationunmasked 96nm doublet microtubule repeat from WT mouse sperm
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 2 of 96nm doublet microtubule repeat from WT mouse sperm

Fileemd_29013_half_map_1.map
Annotationhalf map 2 of 96nm doublet microtubule repeat from WT mouse sperm
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 1 of 96nm doublet microtubule repeat from WT mouse sperm

Fileemd_29013_half_map_2.map
Annotationhalf map 1 of 96nm doublet microtubule repeat from WT mouse sperm
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Mouse sperm

EntireName: Mouse sperm
Components
  • Cell: Mouse sperm

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Supramolecule #1: Mouse sperm

SupramoleculeName: Mouse sperm / type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Mus (mice)

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

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Sample preparation

BufferpH: 5.5
GridModel: Quantifoil R2/2 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: AIR
VitrificationCryogen name: ETHANE / Chamber humidity: 30 % / Chamber temperature: 298 K / Instrument: HOMEMADE PLUNGER

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 0.5 µm / Nominal defocus min: 0.2 µm
Specialist opticsPhase plate: VOLTA PHASE PLATE
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 3.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

ExtractionNumber tomograms: 8 / Number images used: 1212 / Software - Name: IMOD (ver. 4.9)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 4)
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 34.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 4) / Number subtomograms used: 1212
FSC plot (resolution estimation)

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