Herpesviruses are enveloped viruses that are prevalent in the human population and are responsible for diverse pathologies, including cold sores, birth defects and cancers. They are characterized by a highly pressurized pseudo-icosahedral capsid—with triangulation number (T) equal to 16—encapsidating a tightly packed double-stranded DNA (dsDNA) genome1,2,3. A key process in the herpesvirus life cycle involves the recruitment of an ATP-driven terminase to a unique portal vertex to recognize, package and cleave concatemeric dsDNA, ultimately giving rise to a pressurized, genome-containing virion4,5. Although this process has been studied in dsDNA phages6,7,8,9—with which herpesviruses bear some similarities—a lack of high-resolution in situ structures of genome-packaging machinery has prevented the elucidation of how these multi-step reactions, which require close coordination among multiple actors, occur in an integrated environment. To better define the structural basis of genome packaging and organization in herpes simplex virus type 1 (HSV-1), we developed sequential localized classification and symmetry relaxation methods to process cryo-electron microscopy (cryo-EM) images of HSV-1 virions, which enabled us to decouple and reconstruct hetero-symmetric and asymmetric elements within the pseudo-icosahedral capsid. Here we present in situ structures of the unique portal vertex, genomic termini and ordered dsDNA coils in the capsid spooled around a disordered dsDNA core. We identify tentacle-like helices and a globular complex capping the portal vertex that is not observed in phages, indicative of herpesvirus-specific adaptations in the DNA-packaging process. Finally, our atomic models of portal vertex elements reveal how the fivefold-related capsid accommodates symmetry mismatch imparted by the dodecameric portal—a longstanding mystery in icosahedral viruses—and inform possible DNA-sequence recognition and headful-sensing pathways involved in genome packaging. This work showcases how to resolve symmetry-mismatched elements in a large eukaryotic virus and provides insights into the mechanisms of herpesvirus genome packaging.
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The five cryo-EM maps have been deposited in the Electron Microscopy Data Bank (EMDB) under accession numbers EMD-9860 (C5 portal vertex reconstruction), EMD-9861 (C1 portal vertex reconstruction), EMD-9862 (C12 portal reconstruction), EMD-9863 (C1 terminal DNA and portal vertex reconstruction) and EMD-9864 (C1 virion reconstruction). The atomic models for pUL6 and periportal capsid or CATC proteins have been deposited in the Protein Data Bank under accession numbers 6OD7 and 6ODM, respectively.
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We thank W. Liu for assistance in molecular dynamic flexible fitting. This research has been supported in part by grants from the National Key R&D Program of China (2017YFA0505300 and 2016YFA0400900) and the US National Institutes of Health (GM071940/DE025567/DE028583/AI094386). We acknowledge the use of instruments at the Electron Imaging Center for Nanomachines supported by UCLA and by instrumentation grants from NIH (1S10RR23057 and 1U24GM116792) and NSF (DBI-1338135 and DMR-1548924). We thank the Bioinformatics Center of the University of Science and Technology of China, School of Life Science, for providing supercomputing resources for this project.
Nature thanks Sarah Butcher, Venigalla Rao and the other anonymous reviewer(s) for their contribution to the peer review of this work.
The authors declare no competing interests.
Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Extended data figures and tables
Flow chart illustrates the identification and resolution of symmetry-mismatched structures of the unique portal vertex.
a, b, Resolution of reconstructions determined by gold-standard FSC at the 0.143 criterion. c–g, Density slices coloured by local resolution estimated from ResMap41.
a–c, pUL6 monomer coloured by domain for reference (a) and key (b) used to annotate a secondary structure and disorder prediction of pUL6 amino acid sequence (c) obtained from Phyre246.
a, b, C1 reconstruction of terminal DNA with surrounding portal colour-zoned by pUL6 domains and tentacle helices. c, Sequence of terminal DNA mapped onto our fitted terminal DNA model. d, Enlarged view of the trailing end of terminal DNA, where concatemeric cleavage occurs. e, Enlarged view of terminal the disordered leading end of DNA, which extends down through the portal aperture towards the interior of the capsid. f–h, Slab views of C1 density showing interaction of terminal DNA with tentacle helices (f), portal clip (g) and the portal aperture (h).
a–d, HSV-1 pUL6 and pUL6 homologues coloured analogously by pUL6 domain. e–h, HSV-1 pUL6 portal complex and homologues coloured in rainbow (red (N terminus) to blue (C terminus)). Respective insets illustrate the conserved left-handed corkscrew of stem helices in the portal channel beneath the clip.
a–c, Comparison of periportal and peripenton P1 MCPs (a) reveal conformational differences in their dimerization domains (b, c). d–f, Comparison of periportal and peripenton P6 MCPs (d) reveal conformational differences in their N-lassos (e, f). g–i, Comparison of periportal and peripenton Tri1s (g) reveal differences in a trunk loop where periportal Tri1 interfaces with tentacle helices (h) and a visible N-anchor helix in periportal Tri1 (i).
Structures of the HSV-1 virion and portal vertex. Depicts the global reconstructed features of the HSV-1 virion and portal vertex. Related to Figure 1.
pUL6 dodecameric portal. Depicts the atomic model of pUL6 protein and features of the dodecameric portal. Related to Figure 2.
Terminal DNA in the portal vertex channel. Depicts fitted model of terminal DNA and its interactions with structures of the DNA translocation channel. Related to Figures 2 and 3.
Capsid/CATC structures at the portal vertex. Depicts portal vertex-specific features of the capsid and CATC. Related to Figure 4.
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Liu, Y., Jih, J., Dai, X. et al. Cryo-EM structures of herpes simplex virus type 1 portal vertex and packaged genome. Nature 570, 257–261 (2019). https://doi.org/10.1038/s41586-019-1248-6
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