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Cell Reports. Medicine Oct 2023The complex interplay between microbiota and immunity is important to human health. To explore how altered adaptive immunity influences the microbiome, we characterize...
The complex interplay between microbiota and immunity is important to human health. To explore how altered adaptive immunity influences the microbiome, we characterize skin, nares, and gut microbiota of patients with recombination-activating gene (RAG) deficiency-a rare genetically defined inborn error of immunity (IEI) that results in a broad spectrum of clinical phenotypes. Integrating de novo assembly of metagenomes from RAG-deficient patients with reference genome catalogs provides an expansive multi-kingdom view of microbial diversity. RAG-deficient patient microbiomes exhibit inter-individual variation, including expansion of opportunistic pathogens (e.g., Corynebacterium bovis, Haemophilus influenzae), and a relative loss of body site specificity. We identify 35 and 27 bacterial species derived from skin/nares and gut microbiomes, respectively, which are distinct to RAG-deficient patients compared to healthy individuals. Underscoring IEI patients as potential reservoirs for viral persistence and evolution, we further characterize the colonization of eukaryotic RNA viruses (e.g., Coronavirus 229E, Norovirus GII) in this patient population.
Topics: Humans; Microbiota; Gastrointestinal Microbiome; Skin; Metagenome
PubMed: 37757827
DOI: 10.1016/j.xcrm.2023.101205 -
Journal of Virology Jul 2023Adeno-associated virus (AAV) is a nonenveloped single-stranded DNA (ssDNA) icosahedral T=1 virus being developed as a vector for clinical gene delivery systems....
Adeno-associated virus (AAV) is a nonenveloped single-stranded DNA (ssDNA) icosahedral T=1 virus being developed as a vector for clinical gene delivery systems. Currently, there are approximately 160 AAV clinical trials, with AAV2 being the most widely studied serotype. To further understand the AAV gene delivery system, this study investigates the role of viral protein (VP) symmetry interactions on capsid assembly, genome packaging, stability, and infectivity. A total of 25 (seven 2-fold, nine 3-fold, and nine 5-fold symmetry interface) AAV2 VP variants were studied. Six 2-fold and two 5-fold variants did not assemble capsids based on native immunoblots and anti-AAV2 enzyme-linked immunosorbent assays (ELISAs). Seven of the 3-fold and seven of the 5-fold variants that assembled capsids were less stable, while the only 2-fold variant that assembled had ~2°C higher thermal stability () than recombinant wild-type AAV2 (wtAAV2). Three of the 3-fold variants (AAV2-R432A, AAV2-L510A, and N511R) had an approximately 3-log defect in genome packaging. Consistent with previous reports of the 5-fold axes, the region of the capsid is important for VP1u externalization and genome ejection, and one 5-fold variant (R404A) had a significant defect in viral infectivity. The structures of wtAAV2 packaged with a transgene (AAV2-full) and without a transgene (AAV2-empty) and one 5-fold variant (AAV2-R404A) were determined by cryo-electron microscopy and three dimensional (3D)-image reconstruction to 2.8, 2.9, and 3.6 Å resolution, respectively. These structures revealed the role of stabilizing interactions on the assembly, stability, packaging, and infectivity of the virus capsid. This study provides insight into the structural characterization and functional implications of the rational design of AAV vectors. Adeno-associated viruses (AAVs) have been shown to be useful vectors for gene therapy applications. Consequently, AAV has been approved as a biologic for the treatment of several monogenic disorders, and many additional clinical trials are ongoing. These successes have generated significant interest in all aspects of the basic biology of AAV. However, to date, there are limited data available on the importance of the capsid viral protein (VP) symmetry-related interactions required to assemble and maintain the stability of the AAV capsids and the infectivity of the AAV capsids. Characterizing the residue type and interactions at these symmetry-driven assembly interfaces of AAV2 has provided the foundation for understanding their role in AAV vectors (serotypes and engineered chimeras) and has determined the residues or regions of the capsid that can or cannot tolerate alterations.
Topics: Capsid; Dependovirus; Serogroup; Cryoelectron Microscopy; Capsid Proteins; Parvovirinae; Viral Proteins; Genetic Vectors; Virus Assembly
PubMed: 37310260
DOI: 10.1128/jvi.01772-22 -
ELife Dec 2023Nucleotide and force-dependent mechanisms control how the viral genome of lambda bacteriophage is inserted into capsids.
Nucleotide and force-dependent mechanisms control how the viral genome of lambda bacteriophage is inserted into capsids.
Topics: DNA, Viral; Bacteriophage lambda; Capsid; Genome, Viral; Nucleotides; Virus Assembly
PubMed: 38095555
DOI: 10.7554/eLife.94128 -
Spatial and functional arrangement of Ebola virus polymerase inside phase-separated viral factories.Nature Communications Jul 2023Ebola virus (EBOV) infection induces the formation of membrane-less, cytoplasmic compartments termed viral factories, in which multiple viral proteins gather and...
Ebola virus (EBOV) infection induces the formation of membrane-less, cytoplasmic compartments termed viral factories, in which multiple viral proteins gather and coordinate viral transcription, replication, and assembly. Key to viral factory function is the recruitment of EBOV polymerase, a multifunctional machine that mediates transcription and replication of the viral RNA genome. We show that intracellularly reconstituted EBOV viral factories are biomolecular condensates, with composition-dependent internal exchange dynamics that likely facilitates viral replication. Within the viral factory, we found the EBOV polymerase clusters into foci. The distance between these foci increases when viral replication is enabled. In addition to the typical droplet-like viral factories, we report the formation of network-like viral factories during EBOV infection. Unlike droplet-like viral factories, network-like factories are inactive for EBOV nucleocapsid assembly. This unique view of EBOV propagation suggests a form-to-function relationship that describes how physical properties and internal structures of biomolecular condensates influence viral biogenesis.
Topics: Humans; Ebolavirus; Hemorrhagic Fever, Ebola; Viral Replication Compartments; Transcription, Genetic; Virus Replication; Nucleotidyltransferases
PubMed: 37443171
DOI: 10.1038/s41467-023-39821-7 -
Viruses Aug 2023Rotaviruses (RVs) are non-enveloped multilayered dsRNA viruses that are major etiologic agents of diarrheal disease in humans and in the young in a large number of... (Review)
Review
Rotaviruses (RVs) are non-enveloped multilayered dsRNA viruses that are major etiologic agents of diarrheal disease in humans and in the young in a large number of animal species. The viral particle is composed of three different protein layers that enclose the segmented dsRNA genome and the transcriptional complexes. Each layer defines a unique subparticle that is associated with a different phase of the replication cycle. Thus, while single- and double-layered particles are associated with the intracellular processes of selective packaging, genome replication, and transcription, the viral machinery necessary for entry is located in the third layer. This modular nature of its particle allows rotaviruses to control its replication cycle by the disassembly and assembly of its structural proteins. In this review, we examine the significant advances in structural, molecular, and cellular RV biology that have contributed during the last few years to illuminating the intricate details of the RV particle disassembly and assembly processes.
Topics: Animals; Humans; Rotavirus; RNA, Double-Stranded; Virion
PubMed: 37632092
DOI: 10.3390/v15081750 -
Proceedings of the National Academy of... Jul 2023HIV-1 assembly occurs at the inner leaflet of the plasma membrane (PM) in highly ordered membrane microdomains. The size and stability of membrane microdomains is...
HIV-1 assembly occurs at the inner leaflet of the plasma membrane (PM) in highly ordered membrane microdomains. The size and stability of membrane microdomains is regulated by activity of the sphingomyelin hydrolase neutral sphingomyelinase 2 (nSMase2) that is localized primarily to the inner leaflet of the PM. In this study, we demonstrate that pharmacological inhibition or depletion of nSMase2 in HIV-1-producer cells results in a block in the processing of the major viral structural polyprotein Gag and the production of morphologically aberrant, immature HIV-1 particles with severely impaired infectivity. We find that disruption of nSMase2 also severely inhibits the maturation and infectivity of other primate lentiviruses HIV-2 and simian immunodeficiency virus, has a modest or no effect on nonprimate lentiviruses equine infectious anemia virus and feline immunodeficiency virus, and has no effect on the gammaretrovirus murine leukemia virus. These studies demonstrate a key role for nSMase2 in HIV-1 particle morphogenesis and maturation.
Topics: Animals; Cats; Horses; Mice; HIV-1; Sphingomyelin Phosphodiesterase; Virus Assembly; Lentivirus; Infectious Anemia Virus, Equine
PubMed: 37406093
DOI: 10.1073/pnas.2219475120 -
Seminars in Liver Disease Aug 2023First discovered over 40 years ago, the hepatitis delta virus (HDV) is a unique RNA virus, requiring hepatitis B virus (HBV) antigens for its assembly, replication, and... (Review)
Review
First discovered over 40 years ago, the hepatitis delta virus (HDV) is a unique RNA virus, requiring hepatitis B virus (HBV) antigens for its assembly, replication, and transmission. HBV and HDV can be acquired at the same time (coinfection) or HDV infection can occur in persons with chronic HBV (superinfection). Screening guidelines for HDV are inconsistent. While some guidelines recommend universal screening for all people with HBV, others recommend risk-based screening. Estimates of the global HDV prevalence range from 4.5 to 14.6% among persons with HBV; thus, there may be up to 72 million individuals with HDV worldwide. HDV is the most severe form of viral hepatitis. Compared to HBV monoinfection, HDV coinfection increases the risk of cirrhosis, hepatocellular carcinoma, hepatic decompensation, mortality, and necessity for liver transplant. Despite the severity of HDV, there are few treatment options. Pegylated interferon (off-label use) has long been the only available treatment, although bulevirtide is conditionally approved in some European countries. There are many potential treatments in development, but as yet, there are few effective and safe therapies for HDV infection. In conclusion, given the severity of HDV disease and the paucity of treatments, there is a great unmet need for HDV therapies.
Topics: Humans; Hepatitis B; Coinfection; Liver Neoplasms; Hepatitis Delta Virus; Hepatitis D; Hepatitis B virus
PubMed: 37473778
DOI: 10.1055/a-2133-8614 -
Nature Communications Aug 2023The human endogenous retrovirus K (HERV-K) is the most recently acquired endogenous retrovirus in the human genome and is activated and expressed in many cancers and...
The human endogenous retrovirus K (HERV-K) is the most recently acquired endogenous retrovirus in the human genome and is activated and expressed in many cancers and amyotrophic lateral sclerosis. We present the immature HERV-K capsid structure at 3.2 Å resolution determined from native virus-like particles using cryo-electron tomography and subtomogram averaging. The structure shows a hexamer unit oligomerized through a 6-helix bundle, which is stabilized by a small molecule analogous to IP6 in immature HIV-1 capsid. The HERV-K immature lattice is assembled via highly conserved dimer and trimer interfaces, as detailed through all-atom molecular dynamics simulations and supported by mutational studies. A large conformational change mediated by the linker between the N-terminal and the C-terminal domains of CA occurs during HERV-K maturation. Comparison between HERV-K and other retroviral immature capsid structures reveals a highly conserved mechanism for the assembly and maturation of retroviruses across genera and evolutionary time.
Topics: Humans; Endogenous Retroviruses; Amyotrophic Lateral Sclerosis; Biological Evolution; Capsid; Capsid Proteins
PubMed: 37620323
DOI: 10.1038/s41467-023-40786-w -
Nature Communications Nov 2023For successful infection of host cells and virion production, enveloped viruses, including Zika virus (ZIKV), extensively rely on cellular lipids. However, how virus...
For successful infection of host cells and virion production, enveloped viruses, including Zika virus (ZIKV), extensively rely on cellular lipids. However, how virus protein-lipid interactions contribute to the viral life cycle remains unclear. Here, we employ a chemo-proteomics approach with a bifunctional cholesterol probe and show that cholesterol is closely associated with the ZIKV structural protein prM. Bioinformatic analyses, reverse genetics alongside with photoaffinity labeling assays, and atomistic molecular dynamics simulations identified two functional cholesterol binding motifs within the prM transmembrane domain. Loss of prM-cholesterol association has a bipartite effect reducing ZIKV entry and leading to assembly defects. We propose a model in which membrane-resident M facilitates cholesterol-supported lipid exchange during endosomal entry and, together with cholesterol, creates a platform promoting virion assembly. In summary, we identify a bifunctional role of prM in the ZIKV life cycle by mediating viral entry and virus assembly in a cholesterol-dependent manner.
Topics: Humans; Zika Virus; Zika Virus Infection; Virus Internalization; Virus Replication; Viral Proteins; Lipids
PubMed: 37957166
DOI: 10.1038/s41467-023-42985-x -
Viruses Jun 2023Inflammasome activation is exclusively involved in sensing activation of innate immunity and inflammatory response during viral infection. Accumulating evidence suggests... (Review)
Review
Inflammasome activation is exclusively involved in sensing activation of innate immunity and inflammatory response during viral infection. Accumulating evidence suggests that the manipulation of inflammasome assembly or its interaction with viral proteins are critical factors in viral pathogenesis. Results from pilot clinical trials show encouraging results of NLRP3 inflammasome suppression in reducing mortality and morbidity in SARS-CoV-2-infected patients. In this article, we summarize the up-to-date understanding of inflammasomes, including NLRP3, AIM2, NLRP1, NLRP6, and NLRC4 in various viral infections, with particular focus on RNA viruses such as SARS-CoV-2, HIV, IAV, and Zika virus and DNA viruses such as herpes simplex virus 1. We also discuss the current achievement of the mechanisms involved in viral infection-induced inflammatory response, host defense, and possible therapeutic solutions.
Topics: Humans; Inflammasomes; NLR Family, Pyrin Domain-Containing 3 Protein; COVID-19; SARS-CoV-2; Immunity, Innate; Virus Diseases; Zika Virus; Zika Virus Infection
PubMed: 37515138
DOI: 10.3390/v15071451