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Cell Reports. Medicine May 2024Stimulator of interferon genes (STING)-dependent signaling is requisite for effective anti-microbial and anti-tumor activity. STING signaling is commonly defective in...
Stimulator of interferon genes (STING)-dependent signaling is requisite for effective anti-microbial and anti-tumor activity. STING signaling is commonly defective in cancer cells, which enables tumor cells to evade the immunosurveillance system. We evaluate here whether intrinsic STING signaling in such tumor cells could be reconstituted by creating recombinant herpes simplex viruses (rHSVs) that express components of the STING signaling pathway. We observe that rHSVs expressing STING and/or cGAS replicate inefficiently yet retain in vivo anti-tumor activity, independent of oncolytic activity requisite on the trans-activation of extrinsic STING signaling in phagocytes by engulfed microbial dsDNA species. Accordingly, the in vivo effects of virotherapy could be simulated by nanoparticles incorporating non-coding dsDNA species, which comparably elicit the trans-activation of phagocytes and augment the efficacy of established cancer treatments including checkpoint inhibition and radiation therapy. Our results help elucidate mechanisms of virotherapeutic anti-tumor activity as well as provide alternate strategies to treat cancer.
Topics: Animals; Phagocytes; Humans; Mice; DNA; Membrane Proteins; Signal Transduction; Nucleotidyltransferases; Cell Line, Tumor; Neoplasms; Simplexvirus; Mice, Inbred C57BL; Oncolytic Virotherapy
PubMed: 38677283
DOI: 10.1016/j.xcrm.2024.101528 -
Chimia Apr 2024RNA, widely recognized as an information-carrier molecule, is capable of catalyzing essential biological processes through ribozymes. Despite their ubiquity, specific...
RNA, widely recognized as an information-carrier molecule, is capable of catalyzing essential biological processes through ribozymes. Despite their ubiquity, specific functions in a biological context and phenotypes based on the ribozymes' activity are often unknown. Here, we present the discovery of a subgroup of minimal HDV-like ribozymes, which reside 3' to viral tRNAs and appear to cleave the 3'-trailers of viral premature tRNA transcripts. This proposed tRNA-processing function is unprecedented for any ribozymes, thus, we designate this subgroup as theta ribozymes. Most theta ribozymes were identified in Caudoviricetes bacteriophages, the main constituent (>90%) of the mammalian gut virome. Intriguingly, our findings further suggest the involvement of theta ribozymes in the transition of certain bacteriophages between distinct genetic codes, thus possibly contributing to the phage lysis trigger. Our discovery expands the limited repertoire of biological functions attributed to HDV-like ribozymes and provides insights into the fascinating world of RNA catalysis.
Topics: RNA, Catalytic; RNA, Viral; RNA, Transfer; Bacteriophages; Hepatitis Delta Virus
PubMed: 38676609
DOI: 10.2533/chimia.2024.200 -
Viruses Apr 2024Virus-encoded replicases often generate aberrant RNA genomes, known as defective viral genomes (DVGs). When co-infected with a helper virus providing necessary proteins,...
Virus-encoded replicases often generate aberrant RNA genomes, known as defective viral genomes (DVGs). When co-infected with a helper virus providing necessary proteins, DVGs can multiply and spread. While DVGs depend on the helper virus for propagation, they can in some cases disrupt infectious virus replication, impact immune responses, and affect viral persistence or evolution. Understanding the dynamics of DVGs alongside standard viral genomes during infection remains unclear. To address this, we conducted a long-term experimental evolution of two betacoronaviruses, the human coronavirus OC43 (HCoV-OC43) and the murine hepatitis virus (MHV), in cell culture at both high and low multiplicities of infection (MOI). We then performed RNA-seq at regular time intervals, reconstructed DVGs, and analyzed their accumulation dynamics. Our findings indicate that DVGs evolved to exhibit greater diversity and abundance, with deletions and insertions being the most common types. Notably, some high MOI deletions showed very limited temporary existence, while others became prevalent over time. We observed differences in DVG abundance between high and low MOI conditions in HCoV-OC43 samples. The size distribution of HCoV-OC43 genomes with deletions differed between high and low MOI passages. In low MOI lineages, short and long DVGs were the most common, with an additional cluster in high MOI lineages which became more prevalent along evolutionary time. MHV also showed variations in DVG size distribution at different MOI conditions, though they were less pronounced compared to HCoV-OC43, suggesting a more random distribution of DVG sizes. We identified hotspot regions for deletions that evolved at a high MOI, primarily within cistrons encoding structural and accessory proteins. In conclusion, our study illustrates the widespread formation of DVGs during betacoronavirus evolution, influenced by MOI and cell- and virus-specific factors.
Topics: Genome, Viral; Animals; Humans; Virus Replication; Defective Viruses; Murine hepatitis virus; Evolution, Molecular; Coronavirus OC43, Human; Mice; RNA, Viral; Cell Line
PubMed: 38675984
DOI: 10.3390/v16040644 -
Viruses Apr 2024The prevalence of hepatitis B and delta viruses (HBV/HDV) among people who use drugs (PWUD) remains largely unknown. In the context of one Philadelphia-based harm...
The prevalence of hepatitis B and delta viruses (HBV/HDV) among people who use drugs (PWUD) remains largely unknown. In the context of one Philadelphia-based harm reduction organization (HRO), this study aimed to assess HBV/HDV prevalence and facilitate linkage to care. Participants completed a demographic HBV/HDV risk factor survey and were screened for HBV and reflexively for HDV if positive for HBV surface antigen or isolated core antibody. Fisher's exact tests and regression were used to understand relationships between risks and HBV blood markers. Of the 498 participants, 126 (25.3%) did not have hepatitis B immunity, 52.6% had been vaccinated against HBV, and 17.9% had recovered from a past infection. Eleven (2.2%) participants tested positive for isolated HBV core antibody, 10 (2.0%) for HBV surface antigen, and one (0.2%) for HDV antibody. History of incarceration was associated with current HBV infection, while transactional sex and experience of homelessness were predictive of previous exposure. This study found high rates of current and past HBV infection, and a 10% HBV/HDV co-infection rate. Despite availability of vaccine, one quarter of participants remained vulnerable to infection. Findings demonstrate the need to improve low-threshold HBV/HDV screening, vaccination, and linkage to care among PWUD. The study also identified gaps in the HBV/HDV care cascade, including lack of point-of-care diagnostics and lack of support for HROs to provide HBV services.
Topics: Humans; Female; Male; Philadelphia; Hepatitis B; Adult; Middle Aged; Mass Screening; Hepatitis D; Prevalence; Drug Users; Risk Factors; Young Adult; Hepatitis Delta Virus; Hepatitis B virus; Hepatitis B Surface Antigens
PubMed: 38675969
DOI: 10.3390/v16040628 -
Viruses Apr 2024In a survey of mycoviruses in species that cause sugarcane Pokkah boeng disease, twelve strains from three species (, , and ) were found to contain Fusarium sacchari...
In a survey of mycoviruses in species that cause sugarcane Pokkah boeng disease, twelve strains from three species (, , and ) were found to contain Fusarium sacchari hypovirus 1 (FsHV1), which we reported previously. The genomes of these variants range from 13,966 to 13,983 nucleotides, with 98.6% to 99.9% nucleotide sequence identity and 98.70% to 99.9% protein sequence similarity. Phylogenetic analysis placed these FsHV1 variants within the cluster of . Intriguingly, no clear correlation was found between the geographic origin and host specificity of these viral variants. Additionally, six out of the twelve variants displayed segmental deletions of 1.5 to 1.8 kilobases, suggesting the existence of defective viral dsRNA. The presence of defective viral dsRNA led to a two-thirds reduction in the dsRNA of the wild-type viral genome, yet a tenfold increase in the total viral dsRNA content. To standardize virulence across natural strains, all FsHV1 strains were transferred into a single, virus-free recipient strain, FZ06-VF, via mycelial fusion. Strains of carrying FsHV1 exhibited suppressed pigment synthesis, diminished microspore production, and a marked decrease in virulence. Inoculation tests revealed varying capacities among different FsHV1 variants to modulate fungal virulence, with the strain harboring the FsHV1-FSA1 showing the lowest virulence, with a disease severity index (DSI) of 3.33, and the FsHV1-FS1 the highest (DSI = 17.66). The identification of highly virulent FsHV1 variants holds promise for the development of biocontrol agents for Pokkah boeng management.
Topics: Fusarium; Virulence; Plant Diseases; Phylogeny; Genome, Viral; Fungal Viruses; Saccharum; RNA, Viral; Host Specificity
PubMed: 38675949
DOI: 10.3390/v16040608 -
Viruses Mar 2024The administration route affects the biodistribution of a gene transfer vector and the expression of a transgene. A simian adenovirus 1 vector carrying firefly...
The administration route affects the biodistribution of a gene transfer vector and the expression of a transgene. A simian adenovirus 1 vector carrying firefly luciferase and GFP reporter genes (SAdV1-GFluc) were constructed, and its biodistribution was investigated in a mouse model by bioluminescence imaging and virus DNA tracking with real-time PCR. Luciferase activity and virus DNA were mainly found in the liver and spleen after the intravenous administration of SAdV1-GFluc. The results of flow cytometry illustrated that macrophages in the liver and spleen as well as hepatocytes were the target cells. Repeated inoculation was noneffective because of the stimulated serum neutralizing antibodies (NAbs) against SAdV-1. A transient, local expression of low-level luciferase was detected after intragastric administration, and the administration could be repeated without compromising the expression of the reporter gene. Intranasal administration led to a moderate, constant expression of a transgene in the whole respiratory tract and could be repeated one more time without a significant increase in the NAb titer. An immunohistochemistry assay showed that respiratory epithelial cells and macrophages in the lungs were transduced. High luciferase activity was restricted at the injection site and sustained for a week after intramuscular administration. A compromised transgene expression was observed after a repeated injection. When these mice were intramuscularly injected for a third time with the human adenovirus 5 (HAdV-5) vector carrying a luciferase gene, the luciferase activity recovered and reached the initial level, suggesting that the sequential use of SAdV-1 and HAdV-5 vectors was practicable. In short, the intranasal inoculation or intramuscular injection may be the preferred administration routes for the novel SAdV-1 vector in vaccine development.
Topics: Animals; Genetic Vectors; Mice; Genes, Reporter; Adenoviruses, Simian; Tissue Distribution; Green Fluorescent Proteins; Humans; Transgenes; Virus Replication; Luciferases, Firefly; Mice, Inbred BALB C; Female; Transduction, Genetic; Models, Animal; Spleen; Liver; Antibodies, Neutralizing; Gene Expression; Injections, Intramuscular; Administration, Intranasal
PubMed: 38675893
DOI: 10.3390/v16040550 -
Viruses Mar 2024Individuals chronically infected with hepatitis B virus (HBV) and hepatitis Delta virus (HDV) present an increased risk of developing cirrhosis and hepatocellular...
Individuals chronically infected with hepatitis B virus (HBV) and hepatitis Delta virus (HDV) present an increased risk of developing cirrhosis and hepatocellular carcinoma in comparison to HBV mono-infected individuals. Although HDV only replicates in individuals coinfected or superinfected with HBV, there is currently no in vitro model that can stably express both viruses simultaneously, mimicking the chronic infections seen in HBV/HDV patients. Here, we present the HepG2BD cell line as a novel in vitro culture system for long-term replication of HBV and HDV. HepG2BD cells derive from HepG2.2.15 cells in which a 2 kb HDV cDNA sequence was inserted into the adeno-associated virus safe harbor integration site 1 (AAVS1) using CRISPR-Cas9. A Tet-Off promoter was placed 5' of the genomic HDV sequence for reliable initiation/repression of viral replication and secretion. HBV and HDV replication were then thoroughly characterized. Of note, non-dividing cells adopt a hepatocyte-like morphology associated with an increased production of both HDV and HBV virions. Finally, HDV seems to negatively interfere with HBV in this model system. Altogether, HepG2BD cells will be instrumental to evaluate, in vitro, the fundamental HBV-HDV interplay during simultaneous chronic replication as well as for antivirals screening targeting both viruses.
Topics: Hepatitis Delta Virus; Humans; Virus Replication; Hepatitis B virus; Hep G2 Cells; Hepatocytes; Hepatitis D; CRISPR-Cas Systems; Dependovirus; Coinfection
PubMed: 38675875
DOI: 10.3390/v16040532 -
Viruses Mar 2024Replicating RNA, including self-amplifying RNA (saRNA) and trans-amplifying RNA (taRNA), holds great potential for advancing the next generation of RNA-based vaccines.... (Review)
Review
Replicating RNA, including self-amplifying RNA (saRNA) and trans-amplifying RNA (taRNA), holds great potential for advancing the next generation of RNA-based vaccines. Unlike transcribed mRNA found in most current RNA vaccines, saRNA or taRNA can be massively replicated within cells in the presence of RNA-amplifying enzymes known as replicases. We recently demonstrated that this property could enhance immune responses with minimal injected RNA amounts. In saRNA-based vaccines, replicase and antigens are encoded on the same mRNA molecule, resulting in very long RNA sequences, which poses significant challenges in production, delivery, and stability. In taRNA-based vaccines, these challenges can be overcome by splitting the replication system into two parts: one that encodes replicase and the other that encodes a short antigen-encoding RNA called transreplicon. Here, we review the identification and use of transreplicon RNA in alphavirus research, with a focus on the development of novel taRNA technology as a state-of-the art vaccine platform. Additionally, we discuss remaining challenges essential to the clinical application and highlight the potential benefits related to the unique properties of this future vaccine platform.
Topics: Animals; Humans; Alphavirus; Alphavirus Infections; RNA, Viral; Vaccine Development; Viral Vaccines; Virus Replication
PubMed: 38675846
DOI: 10.3390/v16040503 -
PLoS Biology Apr 2024The signature feature of all plant viruses is the encoding of movement proteins (MPs) that supports the movement of the viral genome into adjacent cells and through the...
The signature feature of all plant viruses is the encoding of movement proteins (MPs) that supports the movement of the viral genome into adjacent cells and through the vascular system. The recent discovery of umbravirus-like viruses (ULVs), some of which only encode replication-associated proteins, suggested that they, as with umbraviruses that lack encoded capsid proteins (CPs) and silencing suppressors, would require association with a helper virus to complete an infection cycle. We examined the infection properties of 2 ULVs: citrus yellow vein associated virus 1 (CY1), which only encodes replication proteins, and closely related CY2 from hemp, which encodes an additional protein (ORF5CY2) that was assumed to be an MP. We report that both CY1 and CY2 can independently infect the model plant Nicotiana benthamiana in a phloem-limited fashion when delivered by agroinfiltration. Unlike encoded MPs, ORF5CY2 was dispensable for infection of CY2, but was associated with faster symptom development. Examination of ORF5CY2 revealed features more similar to luteoviruses/poleroviruses/sobemovirus CPs than to 30K class MPs, which all share a similar single jelly-roll domain. In addition, only CY2-infected plants contained virus-like particles (VLPs) associated with CY2 RNA and ORF5CY2. CY1 RNA and a defective (D)-RNA that arises during infection interacted with host protein phloem protein 2 (PP2) in vitro and in vivo, and formed a high molecular weight complex with sap proteins in vitro that was partially resistant to RNase treatment. When CY1 was used as a virus-induced gene silencing (VIGS) vector to target PP2 transcripts, CY1 accumulation was reduced in systemic leaves, supporting the usage of PP2 for systemic movement. ULVs are therefore the first plant viruses encoding replication and CPs but no MPs, and whose systemic movement relies on a host MP. This explains the lack of discernable helper viruses in many ULV-infected plants and evokes comparisons with the initial viruses transferred into plants that must have similarly required host proteins for movement.
Topics: Nicotiana; Plant Diseases; Plant Viral Movement Proteins; RNA Viruses; Plant Viruses; Capsid Proteins; RNA, Viral; Genome, Viral; Phloem
PubMed: 38662792
DOI: 10.1371/journal.pbio.3002600 -
Virus Research Jul 2024Although all herpesviruses utilize a highly conserved replication machinery to amplify their viral genomes, different members may have unique strategies to modulate the...
Although all herpesviruses utilize a highly conserved replication machinery to amplify their viral genomes, different members may have unique strategies to modulate the assembly of their replication components. Herein, we characterize the subcellular localization of seven essential replication proteins of varicella-zoster virus (VZV) and show that several viral replication enzymes such as the DNA polymerase subunit ORF28, when expressed alone, are localized in the cytoplasm. The nuclear import of ORF28 can be mediated by the viral DNA polymerase processivity factor ORF16. Besides, ORF16 could markedly enhance the protein abundance of ORF28. Noteworthily, an ORF16 mutant that is defective in nuclear transport still retained the ability to enhance ORF28 abundance. The low abundance of ORF28 in transfected cells was due to its rapid degradation mediated by the ubiquitin-proteasome system. We additionally reveal that radicicol, an inhibitor of the chaperone Hsp90, could disrupt the interaction between ORF16 and ORF28, thereby affecting the nuclear entry and protein abundance of ORF28. Collectively, our findings imply that the cytoplasmic retention and rapid degradation of ORF28 may be a key regulatory mechanism for VZV to prevent untimely viral DNA replication, and suggest that Hsp90 is required for the interaction between ORF16 and ORF28.
Topics: Herpesvirus 3, Human; Humans; Viral Proteins; Virus Replication; Active Transport, Cell Nucleus; DNA-Directed DNA Polymerase; Cell Nucleus; Cytoplasm; Cell Line; DNA Replication
PubMed: 38643859
DOI: 10.1016/j.virusres.2024.199379