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- EMDB-45671: Subtomogram average of the Polar Tube Outer Filament layer from E... -

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Entry
Database: EMDB / ID: EMD-45671
TitleSubtomogram average of the Polar Tube Outer Filament layer from Encephalitozoon intestinalis microsporidian spores
Map dataFiltered map
Sample
  • Cell: Encephalitozoon intestinalis microsporidian spores
KeywordsMicrosporidia / polar tube / subtomogram averaging / cryo-ET / CELL INVASION
Biological speciesEncephalitozoon intestinalis (fungus)
Methodsubtomogram averaging / cryo EM / Resolution: 36.0 Å
AuthorsUsmani M / Coudray N / Bobe D / Kopylov M / Ekiert DC / Bhabha G
Funding support United States, 8 items
OrganizationGrant numberCountry
American Heart Association915749 United States
Other privateSearle Scholars Program, SSP-2018-2737
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)R01AI147131 United States
Other privateHirschl Career Scientist Award
The Pew Charitable TrustsPEW-00033055 United States
Other governmentNIH Common Fund Transformative High Resolution Cryo-Electron Microscopy, U24 GM129539
Simons FoundationSF349247 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)9 P41 GM103310 United States
CitationJournal: bioRxiv / Year: 2024
Title: Cryo-ET reveals the architecture of the polar tube invasion apparatus from microsporidian parasites.
Authors: Mahrukh Usmani / Nicolas Coudray / Margot Riggi / Rishwanth Raghu / Harshita Ramchandani / Daija Bobe / Mykhailo Kopylov / Ellen D Zhong / Janet H Iwasa / Damian C Ekiert / Gira Bhabha
Abstract: Microsporidia are divergent fungal pathogens that employ a harpoon-like apparatus called the polar tube (PT) to invade host cells. The PT architecture and its association with neighboring organelles ...Microsporidia are divergent fungal pathogens that employ a harpoon-like apparatus called the polar tube (PT) to invade host cells. The PT architecture and its association with neighboring organelles remain poorly understood. Here, we use cryo-electron tomography to investigate the structural cell biology of the PT in dormant spores from the human-infecting microsporidian species, . Segmentation and subtomogram averaging of the PT reveal at least four layers: two protein-based layers surrounded by a membrane, and filled with a dense core. Regularly spaced protein filaments form the structural skeleton of the PT. Combining cryo-electron tomography with cellular modeling, we propose a model for the 3-dimensional organization of the polaroplast, an organelle that is continuous with the membrane layer that envelops the PT. Our results reveal the ultrastructure of the microsporidian invasion apparatus , laying the foundation for understanding infection mechanisms.
History
DepositionJul 10, 2024-
Header (metadata) releaseFeb 26, 2025-
Map releaseFeb 26, 2025-
UpdateMar 5, 2025-
Current statusMar 5, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_45671.png

Downloads & links

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Map

FileDownload / File: emd_45671.map.gz / Format: CCP4 / Size: 1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationFiltered map
Projections & slices

Image control

Size
Brightness
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AxesZ (Sec.)Y (Row.)X (Col.)
6.86 Å/pix.
x 64 pix.
= 439.168 Å		6.86 Å/pix.
x 64 pix.
= 439.168 Å		6.86 Å/pix.
x 64 pix.
= 439.168 Å

Surface

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Images are generated by Spider.

Voxel sizeX=Y=Z: 6.862 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.6
Minimum - Maximum-1.0041461 - 1.148324
Average (Standard dev.)0.022266291 (±0.16539502)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions646464
Spacing646464
CellA=B=C: 439.168 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_45671_msk_1.map
Projections & Slices
AxesZYX

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Additional map: Raw unfiltered map

Fileemd_45671_additional_1.map
AnnotationRaw unfiltered map
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Half map: #2

Fileemd_45671_half_map_1.map
Projections & Slices
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Half map: #1

Fileemd_45671_half_map_2.map
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Sample components

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Entire : Encephalitozoon intestinalis microsporidian spores

EntireName: Encephalitozoon intestinalis microsporidian spores
Components
  • Cell: Encephalitozoon intestinalis microsporidian spores

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Supramolecule #1: Encephalitozoon intestinalis microsporidian spores

SupramoleculeName: Encephalitozoon intestinalis microsporidian spores / type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Encephalitozoon intestinalis (fungus)

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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: 7
Component:
ConcentrationFormulaName
200.0 mg/LKClPotassium Chloride
200.0 mg/LKH2PO4Potassium Phosphate monobasic
8000.0 mg/LNaClSodium Chloride
2160.0 mg/LNa2HPO4-7H2)Sodium Phosphate dibasic

Details: Buffer details: DPBS (Gibco 14190144)
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 20 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec.
VitrificationCryogen name: NITROGEN / Details: Wohlwend Compact 01 HPF.
Details1.3e8 spores per uL

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number real images: 1 / Average exposure time: 1.0 sec. / Average electron dose: 2.87 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 5.5 µm / Nominal defocus min: 4.0 µm / Nominal magnification: 26000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 36.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION / Number subtomograms used: 1657
ExtractionNumber tomograms: 1 / Number images used: 7353 / Software - Name: Dynamo
Final 3D classificationSoftware - Name: RELION
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION

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