Structure of the herpes-simplex virus portal-vertex
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ABSTRACT:
SUBMITTER: David Bhella
PROVIDER: EMPIAR-10189 | biostudies-other |
REPOSITORIES: biostudies-other
Publications
Structure of the herpes simplex virus portal-vertex.
McElwee Marion M Vijayakrishnan Swetha S Rixon Frazer F Bhella David D
PLoS biology 20180620 6
Herpesviruses include many important human pathogens such as herpes simplex virus, cytomegalovirus, varicella-zoster virus, and the oncogenic Epstein-Barr virus and Kaposi sarcoma-associated herpesvirus. Herpes virions contain a large icosahedral capsid that has a portal at a unique 5-fold vertex, similar to that seen in the tailed bacteriophages. The portal is a molecular motor through which the viral genome enters the capsid during virion morphogenesis. The genome also exits the capsid through ...[more]
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Project description:Herpesviruses include many important human pathogens such as herpes simplex virus, cytomegalovirus, varicella-zoster virus, and the oncogenic Epstein-Barr virus and Kaposi sarcoma-associated herpesvirus. Herpes virions contain a large icosahedral capsid that has a portal at a unique 5-fold vertex, similar to that seen in the tailed bacteriophages. The portal is a molecular motor through which the viral genome enters the capsid during virion morphogenesis. The genome also exits the capsid through the portal-vertex when it is injected through the nuclear pore into the nucleus of a new host cell to initiate infection. Structural investigations of the herpesvirus portal-vertex have proven challenging, owing to the small size of the tail-like portal-vertex-associated tegument (PVAT) and the presence of the tegument layer that lays between the nucleocapsid and the viral envelope, obscuring the view of the portal-vertex. Here, we show the structure of the herpes simplex virus portal-vertex at subnanometer resolution, solved by electron cryomicroscopy (cryoEM) and single-particle 3D reconstruction. This led to a number of new discoveries, including the presence of two previously unknown portal-associated structures that occupy the sites normally taken by the penton and the Ta triplex. Our data revealed that the PVAT is composed of 10 copies of the C-terminal domain of pUL25, which are uniquely arranged as two tiers of star-shaped density. Our 3D reconstruction of the portal-vertex also shows that one end of the viral genome extends outside the portal in the manner described for some bacteriophages but not previously seen in any eukaryote viruses. Finally, we show that the viral genome is consistently packed in a highly ordered left-handed spool to form concentric shells of DNA. Our data provide new insights into the structure of a molecular machine critical to the biology of an important class of human pathogens.
| S-EPMC6028144 | biostudies-literature
Project description:The packaging of DNA into preformed capsids is a critical step during herpesvirus infection. For herpes simplex virus, this process requires the products of seven viral genes: the terminase proteins pUL15, pUL28, and pUL33; the capsid vertex-specific component (CVSC) proteins pUL17 and pUL25; and the portal proteins pUL6 and pUL32. The pUL6 portal dodecamer is anchored at one vertex of the capsid by interactions with the adjacent triplexes as well as helical density attributed to the pUL17 and pUL25 subunits of the CVSC. To define the roles and structures of the CVSC proteins in virus assembly and DNA packaging, we isolated a number of recombinant viruses expressing pUL25, pUL17, and pUL36 fused with green or red fluorescent proteins as well as viruses with specific deletions in the CVSC genes. Biochemical and structural studies of these mutants demonstrated that (i) four of the helices in the CVSC helix bundle can be attributed to two copies each of pUL36 and pUL25, (ii) pUL17 and pUL6 are required for capsid binding of the terminase complex in the nucleus, (iii) pUL17 is important for determining the site of the first cleavage reaction generating replicated genomes with termini derived from the long-arm component of the herpes simplex virus 1 (HSV-1) genome, (iv) pUL36 serves no direct role in cleavage/packaging, (v) cleavage and stable packaging of the viral genome involve an ordered interaction of the terminase complex and pUL25 with pUL17 at the portal vertex, and (vi) packaging of the viral genome results in a dramatic displacement of the portal.IMPORTANCE 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. A critical step during productive HSV-1 infection is the cleavage and packaging of replicated, concatemeric viral DNA into preformed capsids. A key knowledge gap is how the capsid engages the replicated viral genome and the subsequent packaging of a unit-length HSV genome. Here, biochemical and structural studies focused on the unique portal vertex of wild-type HSV and packaging mutants provide insights into the mechanism of HSV genome packaging. The significance of our research is in identifying the portal proteins pUL6 and pUL17 as key viral factors for engaging the terminase complex with the capsid and the subsequent cleavage, packaging, and stable incorporation of the viral genome in the HSV-1 capsid.
| S-EPMC7925205 | biostudies-literature
Project description: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) genome1-3. 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 phages6-9-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.
| S-EPMC6732574 | biostudies-literature
Project description:Herpes simplex viruses (HSVs) cause human oral and genital ulcer diseases. Patients with HSV-2 have a higher risk of acquiring a human immunodeficiency virus infection. HSV-2 is a member of the α-herpesvirinae subfamily that together with the β- and γ-herpesvirinae subfamilies forms the Herpesviridae family. Here, we report the cryo-electron microscopy structure of the HSV-2 C-capsid with capsid-vertex-specific component (CVSC) that was determined at 3.75 Å using a block-based reconstruction strategy. We present atomic models of multiple conformers for the capsid proteins (VP5, VP23, VP19C, and VP26) and CVSC. Comparison of the HSV-2 homologs yields information about structural similarities and differences between the three herpesviruses sub-families and we identify α-herpesvirus-specific structural features. The hetero-pentameric CVSC, consisting of a UL17 monomer, a UL25 dimer and a UL36 dimer, is bound tightly by a five-helix bundle that forms extensive networks of subunit contacts with surrounding capsid proteins, which reinforce capsid stability.
| S-EPMC6131487 | biostudies-literature
Project description:Herpes viruses are prevalent and well characterized human pathogens. Despite extensive study, much remains to be learned about the structure of the genome packaging and release machinery in the capsids of these large and complex double-stranded DNA viruses. However, such machinery is well characterized in tailed bacteriophage, which share a common evolutionary origin with herpesvirus. In tailed bacteriophage, the genome exits from the virus particle through a portal and is transferred into the host cell by a complex apparatus (i.e. the tail) located at the portal vertex. Here we use electron cryo-tomography of human herpes simplex type-1 (HSV-1) virions to reveal a previously unsuspected feature at the portal vertex, which extends across the HSV-1 tegument layer to form a connection between the capsid and the viral membrane. The location of this assembly suggests that it plays a role in genome release into the nucleus and is also important for virion architecture.
| S-EPMC3464221 | biostudies-literature
Project description:Using atomic force microscopy imaging and nanoindentation measurements, we investigated the effect of the minor capsid proteins pUL17 and pUL25 on the structural stability of icosahedral herpes simplex virus capsids. pUL17 and pUL25, which form the capsid vertex-specific component (CVSC), particularly contributed to capsid resilience along the 5-fold and 2-fold but not along the 3-fold icosahedral axes. Our detailed analyses, including quantitative mass spectrometry of the protein composition of the capsids, revealed that both pUL17 and pUL25 are required to stabilize the capsid shells at the vertices. This indicates that herpesviruses withstand the internal pressure that is generated during DNA genome packaging by locally reinforcing the mechanical sturdiness of the vertices, the most stressed part of the capsids.IMPORTANCE In this study, the structural, material properties of herpes simplex virus 1 were investigated. The capsid of herpes simplex virus is built up of a variety of proteins, and we scrutinized the influence of two of these proteins on the stability of the capsid. For this, we used a scanning force microscope that makes detailed, topographic images of the particles and that is able to perform mechanical deformation measurements. Using this approach, we revealed that both studied proteins play an essential role in viral stability. These new insights support us in forming a complete view on viral structure and furthermore could possibly help not only to develop specific antivirals but also to build protein shells with improved stability for drug delivery purposes.
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Project description:By electron microscopy and image analysis, we find that baculovirus-expressed UL6 is polymorphic, consisting of rings of 11-, 12-, 13-, and 14-fold symmetry. The 12-mer is likely to be the oligomer incorporated into procapsids: at a resolution of 16 A, it has an axial channel, peripheral flanges, and fits snugly into a vacant vertex site. Its architecture resembles those of bacteriophage portal/connector proteins.
| S-EPMC525097 | biostudies-literature
Project description:Resolving the nonicosahedral components in large icosahedral viruses remains a technical challenge in structural virology. We have used the emerging technique of Zernike phase-contrast electron cryomicroscopy to enhance the image contrast of ice-embedded herpes simplex virus type 1 capsids. Image reconstruction enabled us to retrieve the structure of the unique portal vertex in the context of the icosahedral capsid and, for the first time, show the subunit organization of a portal in a virus infecting eukaryotes. Our map unequivocally resolves the 12-subunit portal situated beneath one of the pentameric vertices, thus removing uncertainty over the location and stoichiometry of the herpesvirus portal.
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Project description:UnlabelledpU(L)34 and pU(L)31 of herpes simplex virus (HSV) comprise the nuclear egress complex (NEC) and are required for budding at the inner nuclear membrane. pU(L)31 also associates with capsids, suggesting it bridges the capsid and pU(L)34 in the nuclear membrane to initiate budding. Previous studies showed that capsid association of pU(L)31 was precluded in the absence of the C terminus of pU(L)25, which along with pU(L)17 comprises the capsid vertex-specific complex, or CVSC. The present studies show that the final 20 amino acids of pU(L)25 are required for pU(L)31 capsid association. Unexpectedly, in the complete absence of pU(L)25, or when pU(L)25 capsid binding was precluded by deletion of its first 50 amino acids, pU(L)31 still associated with capsids. Under these conditions, pU(L)31 was shown to coimmunoprecipitate weakly with pU(L)17. Based on these data, we hypothesize that the final 20 amino acids of pU(L)25 are required for pU(L)31 to associate with capsids. In the absence of pU(L)25 from the capsid, regions of capsid-associated pU(L)17 are bound by pU(L)31. Immunogold electron microscopy revealed that pU(L)31 could associate with multiple sites on a single capsid in the nucleus of infected cells. Electron tomography revealed that immunogold particles specific to pU(L)31 protein bind to densities at the vertices of the capsid, a location consistent with that of the CVSC. These data suggest that pU(L)31 loads onto CVSCs in the nucleus to eventually bind pU(L)34 located within the nuclear membrane to initiate capsid budding.ImportanceThis study is important because it localizes pU(L)1, a component previously known to be required for HSV capsids to bud through the inner nuclear membrane, to the vertex-specific complex of HSV capsids, which comprises the unique long region 25 (U(L)25) and U(L)17 gene products. It also shows this interaction is dependent on the C terminus of U(L)25. This information is vital for understanding how capsids bud through the inner nuclear membrane.
| S-EPMC3993549 | biostudies-literature
Project description:Cell entry of enveloped viruses requires specialized viral proteins that mediate fusion with the host membrane by substantial structural rearrangements from a metastable pre- to a stable postfusion conformation. This metastability renders the herpes simplex virus 1 (HSV-1) fusion glycoprotein B (gB) highly unstable such that it readily converts into the postfusion form, thereby precluding structural elucidation of the pharmacologically relevant prefusion conformation. By identification of conserved sequence signatures and molecular dynamics simulations, we devised a mutation that stabilized this form. Functionally locking gB allowed the structural determination of its membrane-embedded prefusion conformation at sub-nanometer resolution and enabled the unambiguous fit of all ectodomains. The resulting pseudo-atomic model reveals a notable conservation of conformational domain rearrangements during fusion between HSV-1 gB and the vesicular stomatitis virus glycoprotein G, despite their very distant phylogeny. In combination with our comparative sequence-structure analysis, these findings suggest common fusogenic domain rearrangements in all class III viral fusion proteins.
| S-EPMC7518877 | biostudies-literature