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Yorodumi- EMDB-8733: The C-terminus of the herpes simplex virus pUL25 protein is requi... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-8733 | |||||||||||||||
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Title | The C-terminus of the herpes simplex virus pUL25 protein is required for release of viral genomes from capsids bound to nuclear pores. | |||||||||||||||
Map data | Virion with deletion of three C-terminal amino acids (578-580) of the UL25 protein | |||||||||||||||
Sample | herpes simplex virus != Human alphaherpesvirus 1 herpes simplex virus
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Biological species | Human alphaherpesvirus 1 (Herpes simplex virus type 1) | |||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 6.4 Å | |||||||||||||||
Authors | Huffman JB / Daniel GR / Falck-Pedersen E / Huet A / Smith GA / Conway JF / Homa FL | |||||||||||||||
Funding support | United States, 4 items
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Citation | Journal: J Virol / Year: 2017 Title: The C Terminus of the Herpes Simplex Virus UL25 Protein Is Required for Release of Viral Genomes from Capsids Bound to Nuclear Pores. Authors: Jamie B Huffman / Gina R Daniel / Erik Falck-Pedersen / Alexis Huet / Greg A Smith / James F Conway / Fred L Homa / Abstract: The herpes simplex virus (HSV) capsid is released into the cytoplasm after fusion of viral and host membranes, whereupon dynein-dependent trafficking along microtubules targets it to the nuclear ...The herpes simplex virus (HSV) capsid is released into the cytoplasm after fusion of viral and host membranes, whereupon dynein-dependent trafficking along microtubules targets it to the nuclear envelope. Binding of the capsid to the nuclear pore complex (NPC) is mediated by the capsid protein pUL25 and the capsid-tethered tegument protein pUL36. Temperature-sensitive mutants in both pUL25 and pUL36 dock at the NPC but fail to release DNA. The uncoating reaction has been difficult to study due to the rapid release of the genome once the capsid interacts with the nuclear pore. In this study, we describe the isolation and characterization of a truncation mutant of pUL25. Live-cell imaging and immunofluorescence studies demonstrated that the mutant was not impaired in penetration of the host cell or in trafficking of the capsid to the nuclear membrane. However, expression of viral proteins was absent or significantly delayed in cells infected with the pUL25 mutant virus. Transmission electron microscopy revealed capsids accumulated at nuclear pores that retained the viral genome for at least 4 h postinfection. In addition, cryoelectron microscopy (cryo-EM) reconstructions of virion capsids did not detect any obvious differences in the location or structural organization for the pUL25 or pUL36 proteins on the pUL25 mutant capsids. Further, in contrast to wild-type virus, the antiviral response mediated by the viral DNA-sensing cyclic guanine adenine synthase (cGAS) was severely compromised for the pUL25 mutant. These results demonstrate that the pUL25 capsid protein has a critical role in releasing viral DNA from NPC-bound capsids. Herpes simplex virus 1 (HSV-1) is the causative agent of several pathologies ranging in severity from the common cold sore to life-threatening encephalitic infection. Early steps in infection include release of the capsid into the cytoplasm, docking of the capsid at a nuclear pore, and release of the viral genome into the nucleus. A key knowledge gap is how the capsid engages the NPC and what triggers release of the viral genome into the nucleus. Here we show that the C-terminal region of the HSV-1 pUL25 protein is required for releasing the viral genome from capsids docked at nuclear pores. The significance of our research is in identifying pUL25 as a key viral factor for genome uncoating. pUL25 is found at each of the capsid vertices as part of the capsid vertex-specific component and implicates the importance of this complex for NPC binding and genome release. | |||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_8733.map.gz | 864.1 MB | EMDB map data format | |
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Header (meta data) | emd-8733-v30.xml emd-8733.xml | 13.2 KB 13.2 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_8733_fsc.xml | 12.8 KB | Display | FSC data file |
Images | emd_8733.png | 340.7 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-8733 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8733 | HTTPS FTP |
-Validation report
Summary document | emd_8733_validation.pdf.gz | 78.5 KB | Display | EMDB validaton report |
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Full document | emd_8733_full_validation.pdf.gz | 77.7 KB | Display | |
Data in XML | emd_8733_validation.xml.gz | 495 B | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8733 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8733 | HTTPS FTP |
-Related structure data
Related structure data | |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_8733.map.gz / Format: CCP4 / Size: 2.1 GB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Virion with deletion of three C-terminal amino acids (578-580) of the UL25 protein | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.8 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : herpes simplex virus
Entire | Name: herpes simplex virus |
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Components |
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-Supramolecule #1: Human alphaherpesvirus 1
Supramolecule | Name: Human alphaherpesvirus 1 / type: virus / ID: 1 / Parent: 0 / NCBI-ID: 10298 / Sci species name: Human alphaherpesvirus 1 / Virus type: VIRION / Virus isolate: OTHER / Virus enveloped: Yes / Virus empty: No |
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Host (natural) | Organism: Homo sapiens (human) |
Molecular weight | Theoretical: 144 MDa |
Virus shell | Shell ID: 1 / Name: capsid / Diameter: 1200.0 Å / T number (triangulation number): 16 |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 1 mg/mL | ||||||||||||
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Buffer | pH: 7.5 Component:
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Grid | Model: Quantifoil R2/1 / Material: COPPER / Mesh: 400 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Pretreatment - Type: GLOW DISCHARGE | ||||||||||||
Vitrification | Cryogen name: ETHANE-PROPANE / Chamber humidity: 100 % / Chamber temperature: 293 K / Instrument: FEI VITROBOT MARK III / Details: 8 second blot. | ||||||||||||
Details | Virion |
-Electron microscopy
Microscope | FEI POLARA 300 |
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Image recording | Film or detector model: FEI FALCON II (4k x 4k) / Digitization - Sampling interval: 14.0 µm / Digitization - Frames/image: 2-6 / Number grids imaged: 1 / Number real images: 4503 / Average exposure time: 1.3 sec. / Average electron dose: 10.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 50.0 µm / Calibrated defocus max: 5.0 µm / Calibrated defocus min: 1.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 5.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 59000 |
Sample stage | Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Tecnai Polara / Image courtesy: FEI Company |