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- EMDB-14686: Subtomogram average of the chikungunya virus neck complex, unsymm... -

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Basic information

Entry
Database: EMDB / ID: EMD-14686
TitleSubtomogram average of the chikungunya virus neck complex, unsymmetrized
Map dataSubtomogram average of chikungunya virus neck complex, without symmetrization.
Sample
  • Organelle or cellular component: Chikungunya virus neck complex
Biological speciesChikungunya virus
Methodsubtomogram averaging / cryo EM / Resolution: 34.0 Å
AuthorsLaurent T / Carlson LA
Funding support Sweden, 1 items
OrganizationGrant numberCountry
Knut and Alice Wallenberg Foundation Sweden
CitationJournal: Elife / Year: 2022
Title: Architecture of the chikungunya virus replication organelle.
Authors: Timothée Laurent / Pravin Kumar / Susanne Liese / Farnaz Zare / Mattias Jonasson / Andreas Carlson / Lars-Anders Carlson /
Abstract: are mosquito-borne viruses that cause serious disease in humans and other mammals. Along with its mosquito vector, the s chikungunya virus (CHIKV) has spread explosively in the last 20 years, and ... are mosquito-borne viruses that cause serious disease in humans and other mammals. Along with its mosquito vector, the s chikungunya virus (CHIKV) has spread explosively in the last 20 years, and there is no approved treatment for chikungunya fever. On the plasma membrane of the infected cell, CHIKV generates dedicated organelles for viral RNA replication, so-called spherules. Whereas structures exist for several viral proteins that make up the spherule, the architecture of the full organelle is unknown. Here, we use cryo-electron tomography to image CHIKV spherules in their cellular context. This reveals that the viral protein nsP1 serves as a base for the assembly of a larger protein complex at the neck of the membrane bud. Biochemical assays show that the viral helicase-protease nsP2, while having no membrane affinity on its own, is recruited to membranes by nsP1. The tomograms further reveal that full-sized spherules contain a single copy of the viral genome in double-stranded form. Finally, we present a mathematical model that explains the membrane remodeling of the spherule in terms of the pressure exerted on the membrane by the polymerizing RNA, which provides a good agreement with the experimental data. The energy released by RNA polymerization is found to be sufficient to remodel the membrane to the characteristic spherule shape.
History
DepositionMar 30, 2022-
Header (metadata) releaseNov 2, 2022-
Map releaseNov 2, 2022-
UpdateNov 2, 2022-
Current statusNov 2, 2022Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_14686.png

Downloads & links

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Map

FileDownload / File: emd_14686.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSubtomogram average of chikungunya virus neck complex, without symmetrization.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
4.29 Å/pix.
x 240 pix.
= 1029.6 Å		4.29 Å/pix.
x 240 pix.
= 1029.6 Å		4.29 Å/pix.
x 240 pix.
= 1029.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 4.29 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.68
Minimum - Maximum-1.961254 - 1.9264829
Average (Standard dev.)0.00023738852 (±0.053279918)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 1029.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_14686_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1

Fileemd_14686_half_map_1.map
AnnotationHalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: half map 2

Fileemd_14686_half_map_2.map
Annotationhalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Chikungunya virus neck complex

EntireName: Chikungunya virus neck complex
Components
  • Organelle or cellular component: Chikungunya virus neck complex

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Supramolecule #1: Chikungunya virus neck complex

SupramoleculeName: Chikungunya virus neck complex / type: organelle_or_cellular_component / ID: 1 / Parent: 0
Details: Protein complex at the membrane neck of chikungunya virus spherules.
Source (natural)Organism: Chikungunya virus

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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: 7.4
GridModel: Quantifoil / Material: GOLD / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY ARRAY / Support film - Film thickness: 10.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.0037 kPa
VitrificationCryogen name: ETHANE-PROPANE / Chamber humidity: 70 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV / Details: 5 seconds blot.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 3.0 µm / Nominal magnification: 33000
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Average electron dose: 1.5 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

ExtractionNumber tomograms: 9 / Number images used: 76 / Method: manual
CTF correctionSoftware: (Name: CTFFIND, CTFPHASEFLIP)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: Dynamo
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 34.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: Dynamo
Details: 34A resolution was calculated using the uploaded mask
Number subtomograms used: 64

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