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- EMDB-54375: Tomogram showing an NA membrane in an A549wt cell infected with W... -

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Basic information

Entry
Database: EMDB / ID: EMD-54375
TitleTomogram showing an NA membrane in an A549wt cell infected with WSNdeltaHA at 16 hpi.
Map dataTomogram showing a NA membrane in an A549wt cell infected with WSN-delHA at 16 hpi.
Sample
  • Cell: A549wt cell infected with WSNdeltaHA virus at 16 hpi.
KeywordsInfluenza A virus / VIRUS
Biological speciesHomo sapiens (human)
Methodelectron tomography / cryo EM
AuthorsWachsmuth-Melm M / Chlanda P
Funding support Germany, 3 items
OrganizationGrant numberCountry
Other privateChica and Heinz Schaller Foundation Germany
German Research Foundation (DFG)437060729 Germany
German Research Foundation (DFG)240245660 Germany
CitationJournal: Nat Commun / Year: 2025
Title: Visualizing influenza A virus assembly by in situ cryo-electron tomography.
Authors: Moritz Wachsmuth-Melm / Sarah Peterl / Aidan O'Riain / Jana Makroczyová / Konstantin Fischer / Tim Krischuns / Sílvia Vale-Costa / Maria João Amorim / Petr Chlanda /
Abstract: Influenza A virus (IAV) forms pleomorphic particles that package eight ribonucleoprotein complexes (vRNPs), each carrying a distinct RNA genome segment. vRNPs assemble in the nucleus and undergo ...Influenza A virus (IAV) forms pleomorphic particles that package eight ribonucleoprotein complexes (vRNPs), each carrying a distinct RNA genome segment. vRNPs assemble in the nucleus and undergo selective sorting during Rab11a-mediated trafficking to the plasma membrane. Virion assembly is orchestrated by matrix protein 1 (M1), which forms a layer beneath the viral envelope containing hemagglutinin (HA) and neuraminidase (NA). However, molecular details of vRNP distribution, cytosolic trafficking, and coordination of IAV assembly remains unclear. Using in situ cryo-ET, we reveal that HA-containing membranes provide Rab11a-dependent platforms for membrane-assisted vRNP clustering, reducing inter-vRNP distances. In the absence of HA, vRNPs cluster on NA-containing membranes and virus assembly remains intact, indicating that vRNP clustering and trafficking is membrane-assisted but HA independent. The characteristic 7 + 1 vRNP bundle forms concomitantly with budding and is orchestrated by M1 layer assembly that precedes plasma membrane attachment. We further reveal that intracellular M1 forms multilayered helical assemblies of antiparallel dimers, structurally distinct from the M1 layer in virions. These assemblies are compact in the nucleus but partially dissociate in the cytoplasm, likely serving as a reservoir for budding. Together, our findings uncover membrane-assisted vRNP clustering and molecular details of M1 coordinated influenza virus assembly.
History
DepositionJul 15, 2025-
Header (metadata) releaseNov 5, 2025-
Map releaseNov 5, 2025-
UpdateNov 5, 2025-
Current statusNov 5, 2025Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_54375.png

Downloads & links

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Map

FileDownload / File: emd_54375.map.gz / Format: CCP4 / Size: 203.9 MB / Type: IMAGE STORED AS SIGNED BYTE
AnnotationTomogram showing a NA membrane in an A549wt cell infected with WSN-delHA at 16 hpi.
Voxel sizeX=Y=Z: 10.36 Å
Density
Minimum - Maximum-128.0 - 127.0
Average (Standard dev.)77.863910000000004 (±8.175553000000001)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-208208-72
Dimensions14401024145
Spacing10241440145
CellA: 10608.64 Å / B: 14918.399 Å / C: 1502.2 Å
α=β=γ: 90.0 °

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Supplemental data

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Sample components

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Entire : A549wt cell infected with WSNdeltaHA virus at 16 hpi.

EntireName: A549wt cell infected with WSNdeltaHA virus at 16 hpi.
Components
  • Cell: A549wt cell infected with WSNdeltaHA virus at 16 hpi.

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Supramolecule #1: A549wt cell infected with WSNdeltaHA virus at 16 hpi.

SupramoleculeName: A549wt cell infected with WSNdeltaHA virus at 16 hpi. / type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Homo sapiens (human) / Organ: Lung

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Experimental details

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Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statecell

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Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE
SectioningFocused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 / Focused ion beam - Current: 0.05 / Focused ion beam - Duration: 300 / Focused ion beam - Temperature: 95 K / Focused ion beam - Initial thickness: 1000 / Focused ion beam - Final thickness: 150
Focused ion beam - Details: The value given for _em_focused_ion_beam.instrument is TFS Aquilos 2. This is not in a list of allowed values {'OTHER', 'DB235'} so OTHER is written into the XML file.

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Electron microscopy

MicroscopeTFS KRIOS
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Average electron dose: 3.3 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: 3.0 µm / Nominal defocus min: 3.0 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionSoftware - Name: eTomo (ver. 5.1) / Number images used: 41
CTF correctionType: PHASE FLIPPING ONLY

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