Most cited article - PubMed ID 30850609
Enterovirus particles expel capsid pentamers to enable genome release
UNLABELLED: Microviruses are single-stranded DNA viruses infecting bacteria, characterized by T = 1 shells made of single jelly-roll capsid proteins. To understand how microviruses infect their host cells, we have isolated and studied an unusually large microvirus, Ebor. Ebor belongs to the proposed "Tainavirinae" subfamily of Microviridae and infects the model Alphaproteobacterium Rhodobacter capsulatus. Using cryogenic electron microscopy, we show that the enlarged capsid of Ebor is the result of an extended C-terminus of the major capsid protein. The extra packaging space accommodates genes encoding a lytic enzyme and putative methylase, both absent in microviruses with shorter genomes. The capsid is decorated with protrusions at its 3-fold axes, which we show to recognize lipopolysaccharides on the host surface. Cryogenic electron tomography shows that during infection, Ebor attaches to the host cell via five such protrusions. This attachment brings a single pentameric capsomer into close contact with the cell membrane, creating a special vertex through which the genome is ejected. Both subtomogram averaging and single particle analysis identified two intermediates of capsid opening, showing that the interacting penton opens from its center via the separation of individual capsomer subunits. Structural comparison with the model Bullavirinae phage phiX174 suggests that this genome delivery mechanism may be widely present across Microviridae. IMPORTANCE: Tailless Microviridae bacteriophages are major components of the global virosphere. Notably, microviruses are prominent members of the mammalian gut virome, and certain compositions have been linked to serious health disorders; however, a molecular understanding of how they initiate infection of their host remains poorly characterized. We demonstrate that trimeric protrusions located at the corners of a single microvirus capsomer mediate host cell attachment. This interaction triggers opening of the capsomer, driven by separation of subunits from its center, much like flower petals open during blooming. This extensive opening explains how the genome translocation apparatus, along with the genome itself, is able to exit the capsid. "Penton blooming" likely represents a conserved mechanism shared by diverse viruses possessing similar capsid architectures.
- Keywords
- Microviridae, Rhodobacter, electron microscopy, structural biology, virion structure,
- MeSH
- Cryoelectron Microscopy MeSH
- Genome, Viral * MeSH
- Capsid ultrastructure MeSH
- Microviridae * genetics physiology ultrastructure MeSH
- Capsid Proteins genetics metabolism chemistry MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Capsid Proteins MeSH
Membrane penetration by non-enveloped viruses is diverse and generally not well understood. Enteroviruses, one of the largest groups of non-enveloped viruses, cause diseases ranging from the common cold to life-threatening encephalitis. Enteroviruses enter cells by receptor-mediated endocytosis. However, how enterovirus particles or RNA genomes cross the endosome membrane into the cytoplasm remains unknown. Here we used cryo-electron tomography of infected cells to show that endosomes containing enteroviruses deform, rupture, and release the virus particles into the cytoplasm. Blocking endosome acidification with bafilomycin A1 reduced the number of particles that released their genomes, but did not prevent them from reaching the cytoplasm. Inhibiting post-endocytic membrane remodeling with wiskostatin promoted abortive enterovirus genome release in endosomes. The rupture of endosomes also occurs in control cells and after the endocytosis of very low-density lipoprotein. In summary, our results show that cellular membrane remodeling disrupts enterovirus-containing endosomes and thus releases the virus particles into the cytoplasm to initiate infection. Since the studied enteroviruses employ different receptors for cell entry but are delivered into the cytoplasm by cell-mediated endosome disruption, it is likely that most if not all enteroviruses, and probably numerous other viruses from the family Picornaviridae, can utilize endosome rupture to infect cells.
- MeSH
- Cell Membrane ultrastructure virology MeSH
- Chlorocebus aethiops MeSH
- COS Cells MeSH
- Cytoplasm virology MeSH
- Cryoelectron Microscopy MeSH
- Endocytosis * MeSH
- Endosomes * pathology virology MeSH
- HeLa Cells MeSH
- Humans MeSH
- Macrolides pharmacology MeSH
- Picornaviridae Infections * virology MeSH
- Rhinovirus * genetics physiology MeSH
- Animals MeSH
- Check Tag
- Humans MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- bafilomycin A1 MeSH Browser
- Macrolides MeSH
Single-stranded DNA bacteriophages of the Microviridae family are major components of the global virosphere. Microviruses are highly abundant in aquatic ecosystems and are prominent members of the mammalian gut microbiome, where their diversity has been linked to various chronic health disorders. Despite the clear importance of microviruses, little is known about the molecular mechanism of host infection. Here, we have characterized an exceptionally large microvirus, Ebor, and provide crucial insights into long-standing mechanistic questions. Cryogenic electron microscopy of Ebor revealed a capsid with trimeric protrusions that recognise lipopolysaccharides on the host surface. Cryogenic electron tomography of the host cell colonized with virus particles demonstrated that the virus initially attaches to the cell via five such protrusions, located at the corners of a single pentamer. This interaction triggers a stargate mechanism of capsid opening along the 5-fold symmetry axis, enabling delivery of the virus genome. Despite variations in specific virus-host interactions among different Microviridae family viruses, structural data indicate that the stargate mechanism of infection is universally employed by all members of the family. Startlingly, our data reveal a mechanistic link for the opening of relatively small capsids made out of a single jelly-roll fold with the structurally unrelated giant viruses.
- Publication type
- Journal Article MeSH
- Preprint MeSH
Enterovirus 70 (EV70) is a human pathogen belonging to the family Picornaviridae. EV70 is transmitted by eye secretions and causes acute hemorrhagic conjunctivitis, a serious eye disease. Despite the severity of the disease caused by EV70, its structure is unknown. Here, we present the structures of the EV70 virion, altered particle, and empty capsid determined by cryo-electron microscopy. The capsid of EV70 is composed of the subunits VP1, VP2, VP3, and VP4. The partially collapsed hydrophobic pocket located in VP1 of the EV70 virion is not occupied by a pocket factor, which is commonly present in other enteroviruses. Nevertheless, we show that the pocket can be targeted by the antiviral compounds WIN51711 and pleconaril, which block virus infection. The inhibitors prevent genome release by stabilizing EV70 particles. Knowledge of the structures of complexes of EV70 with inhibitors will enable the development of capsid-binding therapeutics against this virus. IMPORTANCE Globally distributed enterovirus 70 (EV70) causes local outbreaks of acute hemorrhagic conjunctivitis. The discharge from infected eyes enables the high-efficiency transmission of EV70 in overcrowded areas with low hygienic standards. Currently, only symptomatic treatments are available. We determined the structures of EV70 in its native form, the genome release intermediate, and the empty capsid resulting from genome release. Furthermore, we elucidated the structures of EV70 in complex with two inhibitors that block virus infection, and we describe the mechanism of their binding to the virus capsid. These results enable the development of therapeutics against EV70.
- Keywords
- Picornavirales, Picornaviridae, acute hemorrhagic conjunctivitis, antiviral, canyon, capsid, enterovirus, human, inhibitor, jelly roll, protein, structure, virion, virion structure, virus,
- MeSH
- Conjunctivitis, Acute Hemorrhagic virology MeSH
- Antiviral Agents * pharmacology MeSH
- Cryoelectron Microscopy MeSH
- Capsid * ultrastructure MeSH
- Humans MeSH
- Enterovirus D, Human * drug effects ultrastructure MeSH
- Oxadiazoles pharmacology MeSH
- Oxazoles pharmacology MeSH
- Virion drug effects ultrastructure MeSH
- Capsid Proteins MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Antiviral Agents * MeSH
- Oxadiazoles MeSH
- Oxazoles MeSH
- pleconaril MeSH Browser
- Capsid Proteins MeSH
Coxsackievirus A6 (CV-A6) has recently overtaken enterovirus A71 and CV-A16 as the primary causative agent of hand, foot, and mouth disease worldwide. Virions of CV-A6 were not identified in previous structural studies, and it was speculated that the virus is unique among enteroviruses in using altered particles with expanded capsids to infect cells. In contrast, the virions of other enteroviruses are required for infection. Here we used cryo-electron microscopy (cryo-EM) to determine the structures of the CV-A6 virion, altered particle, and empty capsid. We show that the CV-A6 virion has features characteristic of virions of other enteroviruses, including a compact capsid, VP4 attached to the inner capsid surface, and fatty acid-like molecules occupying the hydrophobic pockets in VP1 subunits. Furthermore, we found that in a purified sample of CV-A6, the ratio of infectious units to virions is 1 to 500. Therefore, it is likely that virions of CV-A6 initiate infection, like those of other enteroviruses. Our results provide evidence that future vaccines against CV-A6 should target its virions instead of the antigenically distinct altered particles. Furthermore, the structure of the virion provides the basis for the rational development of capsid-binding inhibitors that block the genome release of CV-A6.
- MeSH
- Antigens, Viral MeSH
- Cryoelectron Microscopy MeSH
- Enterovirus * genetics MeSH
- Humans MeSH
- Hand, Foot and Mouth Disease * MeSH
- Antibodies, Viral MeSH
- Virion MeSH
- Capsid Proteins genetics MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- Antigens, Viral MeSH
- Antibodies, Viral MeSH
- Capsid Proteins MeSH
The Czech Republic, a part of the former Czechoslovakia, has been at the forefront of several research directions in virology, genetics and physiology [...].
- MeSH
- Virology * MeSH
- Publication type
- Research Support, Non-U.S. Gov't MeSH
- Editorial MeSH
- Geographicals
- Czech Republic MeSH
Most rhinoviruses, which are the leading cause of the common cold, utilize intercellular adhesion molecule-1 (ICAM-1) as a receptor to infect cells. To release their genomes, rhinoviruses convert to activated particles that contain pores in the capsid, lack minor capsid protein VP4, and have an altered genome organization. The binding of rhinoviruses to ICAM-1 promotes virus activation; however, the molecular details of the process remain unknown. Here, we present the structures of virion of rhinovirus 14 and its complex with ICAM-1 determined to resolutions of 2.6 and 2.4 Å, respectively. The cryo-electron microscopy reconstruction of rhinovirus 14 virions contains the resolved density of octanucleotide segments from the RNA genome that interact with VP2 subunits. We show that the binding of ICAM-1 to rhinovirus 14 is required to prime the virus for activation and genome release at acidic pH. Formation of the rhinovirus 14-ICAM-1 complex induces conformational changes to the rhinovirus 14 capsid, including translocation of the C termini of VP4 subunits, which become poised for release through pores that open in the capsids of activated particles. VP4 subunits with altered conformation block the RNA-VP2 interactions and expose patches of positively charged residues. The conformational changes to the capsid induce the redistribution of the virus genome by altering the capsid-RNA interactions. The restructuring of the rhinovirus 14 capsid and genome prepares the virions for conversion to activated particles. The high-resolution structure of rhinovirus 14 in complex with ICAM-1 explains how the binding of uncoating receptors enables enterovirus genome release.
- Keywords
- cryo-electron microscopy, genome release, receptor, structure, virus,
- MeSH
- Virus Activation physiology MeSH
- Cryoelectron Microscopy MeSH
- Enterovirus Infections metabolism virology MeSH
- Genome, Viral genetics MeSH
- HeLa Cells MeSH
- Capsid metabolism MeSH
- Nucleic Acid Conformation MeSH
- Protein Conformation MeSH
- Crystallography, X-Ray MeSH
- Humans MeSH
- Intercellular Adhesion Molecule-1 chemistry genetics metabolism MeSH
- Models, Molecular MeSH
- Rhinovirus genetics metabolism physiology MeSH
- RNA, Viral chemistry genetics metabolism MeSH
- Amino Acid Sequence MeSH
- Sequence Homology, Amino Acid MeSH
- Virus Uncoating physiology MeSH
- Protein Binding MeSH
- Virion genetics metabolism ultrastructure MeSH
- Capsid Proteins chemistry genetics metabolism MeSH
- Check Tag
- Humans MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- ICAM1 protein, human MeSH Browser
- Intercellular Adhesion Molecule-1 MeSH
- RNA, Viral MeSH
- Capsid Proteins MeSH
Infections of Kashmir bee virus (KBV) are lethal for honeybees and have been associated with colony collapse disorder. KBV and closely related viruses contribute to the ongoing decline in the number of honeybee colonies in North America, Europe, Australia, and other parts of the world. Despite the economic and ecological impact of KBV, its structure and infection process remain unknown. Here we present the structure of the virion of KBV determined to a resolution of 2.8 Å. We show that the exposure of KBV to acidic pH induces a reduction in inter-pentamer contacts within capsids and the reorganization of its RNA genome from a uniform distribution to regions of high and low density. Capsids of KBV crack into pieces at acidic pH, resulting in the formation of open particles lacking pentamers of capsid proteins. The large openings of capsids enable the rapid release of genomes and thus limit the probability of their degradation by RNases. The opening of capsids may be a shared mechanism for the genome release of viruses from the family Dicistroviridae ImportanceThe western honeybee (Apis mellifera) is indispensable for maintaining agricultural productivity as well as the abundance and diversity of wild flowering plants. However, bees suffer from environmental pollution, parasites, and pathogens, including viruses. Outbreaks of virus infections cause the deaths of individual honeybees as well as collapses of whole colonies. Kashmir bee virus has been associated with colony collapse disorder in the US, and no cure of the disease is currently available. Here we report the structure of an infectious particle of Kashmir bee virus and show how its protein capsid opens to release the genome. Our structural characterization of the infection process determined that therapeutic compounds stabilizing contacts between pentamers of capsid proteins could prevent the genome release of the virus.
- Publication type
- Journal Article MeSH
The family Iflaviridae includes economically important viruses of the western honeybee such as deformed wing virus, slow bee paralysis virus, and sacbrood virus. Iflaviruses have nonenveloped virions and capsids organized with icosahedral symmetry. The genome release of iflaviruses can be induced in vitro by exposure to acidic pH, implying that they enter cells by endocytosis. Genome release intermediates of iflaviruses have not been structurally characterized. Here, we show that conformational changes and expansion of iflavirus RNA genomes, which are induced by acidic pH, trigger the opening of iflavirus particles. Capsids of slow bee paralysis virus and sacbrood virus crack into pieces. In contrast, capsids of deformed wing virus are more flexible and open like flowers to release their genomes. The large openings in iflavirus particles enable the fast exit of genomes from capsids, which decreases the probability of genome degradation by the RNases present in endosomes.
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH