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Principal Investigator: Z Hong ZHOU
Organization: UNIVERSITY OF CALIFORNIA LOS ANGELES
Fiscal Year: 2023
Award: $456,167
Funding agency: National Institute of Allergy and Infectious Diseases
Double-stranded RNA (dsRNA) viruses comprise a large group of non-enveloped viruses characterized by their ability to transcribe their RNA within an intact capsid (i.e., endogenous RNA transcription), thus evading cellular antiviral responses to dsRNA. Among them, members of the Reoviridae family of dsRNA viruses are of significance in both public health and basic science, exemplified respectively by the gastroenteritis-causing rotavirus which is responsible for approximately half a million child deaths annually worldwide and the insect- killing cytoplasmic polyhedrosis virus (CPV) which was used historically as a model in the discovery of RNA capping. We have studied non-enveloped dsRNA viruses with single-layered (CPV), double-layered [mammalian reovirus (MRV) and aquareovirus (ARV)], and triple-layered [rhesus rotavirus (RRV), Bluetongue virus (BTV)] capsid. These viruses could also be classified based on the presence (such as CPV and reoviruses) or absence (such as BTV and RRV) of an mRNA-capping turret on the icosahedral vertices of their innermost shell. Results from the prior funding cycles have uncovered that BTV and CPV both use surface trimers bearing similarities to fusion proteins of enveloped viruses (e.g., flu, AIDS and COVID-19 viruses) for cell entry. We have also captured the asymmetrically attached transcriptional enzyme complex (TEC) at the quiescent, initiation and transcribing stages of CPV, BTV and RRV; and identified both conserved and diverse features among their structures and organizations of TEC and RNA capping. Our studies showed that, upon cell entry, these viruses sense different environmental cues for internal transcription activation; and in the case of CPV, sensing of SAM and ATP by the RNA-capping turret triggers a cascade of events: opening of the turret iris, detachment of the trimeric spike, and initiation of endogenous transcription. The need to conserve endogenous RNA transcription and the structural diversities uncovered in our prior studies have led to our overall hypothesis: genomes of dsRNA viruses have diverged substantially to allow incorporation of RNA segments encoding the distinct proteins required to interact with different host cells, giving rise to different genome and TEC organizations and variations to both RNA unwinding during transcription and RNA capping during release. The goal of this renewal application is to test this hypothesis with state-of-the-art cryogenic electron microscopy (cryoEM) and tomography (cryoET) by determining representative dsRNA viruses’ genome organizations during quiescence, unwinding and capping during transcription, and genome packing during assembly. We will model the genomes inside CPV, BTV, as well as dsRNA viruses with one and two dsRNA segments for comparison (Aim 1). Capping and cap-snatching during RNA transcription will then be investigated (Aim 2). Finally, we will visualize how different genomic RNA and capsid proteins assemble to form infectious virion particles (Aim 3). As demonstrated in our prior work, these studies will be complemented by structure-based mutagenesis for functional verification.
Terms: <0-11 years old><2019 novel corona virus><2019 novel coronavirus><2019-nCoV><5' Capped RNA><5' mRNA Cap Structure><AIDS Virus><Acquired Immune Deficiency Syndrome Virus><Acquired Immunodeficiency Syndrome Virus><Animals><Anti-Viral Response><Antiviral Response><Aqua reovirus><Aquareoviruses><Aquatic Reovirus><Avian Nephrosis Virus><Bacteria><Basic Research><Basic Science><Biochemical><Biologic Models><Biological Models><Blue Tongue Virus><Bluetongue virus><COVID-19 virus><COVID19 virus><Capsid><Capsid Proteins><Cell Body><Cells><Cessation of life><Child><Child Youth><Children (0-21)><Chimera Protein><Chimeric Proteins><Classification><CoV-2><CoV2><Coat Proteins><Communities><Complement><Complement Proteins><Complex><Cryo-electron Microscopy><Cryoelectron Microscopy><Cues><Cytoplasm><Cytoplasmic Polyhedrosis Viruses><DNA Molecular Biology><Death><Detection><Double Stranded RNA Virus><Double-Stranded RNA><Drug Delivery><Drug Delivery Systems><EC 2.7.7.48><Economics><Electron Cryomicroscopy><Electrons><Engineering><Environment><Enzyme Gene><Enzymes><Event><Family><Farm Animal><Flu virus><Funding><Fusion Protein><Gastroenteritis><Gene Transcription><General Population><General Public><Genetic Transcription><Genetics-Mutagenesis><Genome><Goals><Grant><Gumboro Disease Virus><HIV><Health Sciences><History><Human><Human Immunodeficiency Viruses><IBDV><In Situ><Infectious Bursal Agent><Infectious bursal disease virus><Insect Viruses><Insecta><Insects><Insects Invertebrates><Invaded><Ions><Iris><LAV-HTLV-III><Lead><Life Cycle><Life Cycle Stages><Link><Literature><Livestock><Lymphadenopathy-Associated Virus><Membrane Protein Gene><Membrane Proteins><Membrane-Associated Proteins><Messenger RNA><Model System><Modeling><Modern Man><Molecular Biology><Molecular Configuration><Molecular Conformation><Molecular Stereochemistry><Multienzyme Complexes><Mutagenesis><Mutagenesis Molecular Biology><N-terminal><NH2-terminal><NTPase><Negative Beta Particle><Negatrons><Non-Polyadenylated RNA><Nucleoside Triphosphate Phosphohydrolase><Nucleosidetriphosphatase><Ovine Catarrhal Fever Virus><Pattern><Pb element><Phase><Plants><Polymerase><Proteins><Public Health><Publishing><RNA><RNA Caps><RNA Expression><RNA Gene Products><RNA Replicase><RNA replication><RNA-Dependent RNA Polymerase><RNA-Directed RNA Polymerase><Recording of previous events><Reoviridae><Reovirus><Respiratory Enteric Orphan Viruses><Rhesus><Ribonucleic Acid><Role><Rotavirus><SARS corona virus 2><SARS-CO-V2><SARS-COVID-2><SARS-CoV-2><SARS-CoV2><SARS-associated corona virus 2><SARS-associated coronavirus 2><SARS-coronavirus-2><SARS-related corona virus 2><SARS-related coronavirus 2><SARSCoV2><Severe Acute Respiratory Coronavirus 2><Severe Acute Respiratory Distress Syndrome CoV 2><Severe Acute Respiratory Distress Syndrome Corona Virus 2><Severe Acute Respiratory Distress Syndrome Coronavirus 2><Severe Acute Respiratory Syndrome CoV 2><Severe Acute Respiratory Syndrome-associated coronavirus 2><Severe Acute Respiratory Syndrome-related coronavirus 2><Severe acute respiratory syndrome associated corona virus 2><Severe acute respiratory syndrome coronavirus 2><Severe acute respiratory syndrome related corona virus 2><Structure><Surface><Surface Proteins><System><Systematics><T vaginalis><T. vaginalis><Techniques><Technology><Testing><Transcription><Transcription Activation><Transcriptional Activation><Trichomonas vaginalis><Vaccines><Variant><Variation><Viral><Viral Coat Proteins><Viral Genome><Viral Outer Coat Protein><Virion><Virus><Virus Assembly><Virus Particle><Virus-HIV><Visualization><Work><Wuhan coronavirus><anti-viral development><anti-viral drug development><anti-viral therapeutic development><anti-viral therapy development><antiviral development><antiviral drug development><antiviral therapeutic development><antiviral therapy development><comparative><conformation><conformational><conformational state><conformationally><conformations><coronavirus disease 2019 virus><coronavirus disease-19 virus><cryo-EM><cryoEM><cryogenic electron microscopy><cytoplasmic polyhedrosis virus><develop a vaccine><develop vaccines><developing anti-viral agent><developing anti-viral drug><developing anti-viral therapeutic><developing anti-viral therapy><developing antiviral agent><developing antiviral drug><developing antiviral therapeutic><developing antiviral therapy><development of a vaccine><dsRNA><dsRNA Virus><economic><electron tomography><endosome membrane><enzyme complex><fungus><genomic RNA><hCoV19><heavy metal Pb><heavy metal lead><histories><insight><interest><kids><life course><mRNA><mRNA capping><member><nCoV2><nanometer resolution><novel><nucleoside triphosphatase><particle><pathogen><reconstruction><replicator><social><social role><vaccine development><viral assembly><virus core><virus genome><youngster>