Deep sequencing identified miR-193b-3p as a positive regulator of autophagy targeting Akt3 in Ctenopharyngodon idella CIK cells during GCRV infection.

Recent research has highlighted complex and close interaction between miRNAs, autophagy, and viral infection. In this study, we observed the autophagy status in CIK cells infected with GCRV at various time points. We found that GCRV consistently induced cellar autophagy from 0 h to 12 h post infection. Subsequently, we performed deep sequencing on CIK cells infected with GCRV at 0 h and 12 h respectively, identifying 38 DEMs and predicting 9581 target genes. With the functional enrichment analyses of GO and KEGG, we identified 35 autophagy-related target genes of these DEMs, among which akt3 was pinpointed as the most central hub gene using module assay of the PPI network. Then employing the miRanda and Targetscan programs for prediction, and verification through a double fluorescent enzyme system and qPCR method, we confirmed that miR-193 b-3p could target the 3'-UTR of grass carp akt3, reducing its gene expression. Ultimately, we illustrated that grass carp miR-193 b-3p could promote autophagy in CIK cells. Above results collectively indicated that miRNAs might play a critical role in autophagy of grass carp during GCRV infection and contributed significantly to antiviral immunity by targeting autophagy-related genes. This study may provide new insights into the intricate mechanisms involved in virus, autophagy, and miRNAs.

IL-6/STAT3 axis is hijacked by GCRV to facilitate viral replication via suppressing type â IFN signaling.

Grass carp reovirus (GCRV) infections and hemorrhagic disease (GCHD) outbreaks are typically seasonally periodic and temperature-dependent, yet the molecular mechanism remains unclear. Herein, we depicted that temperature-dependent IL-6/STAT3 axis was exploited by GCRV to facilitate viral replication via suppressing type Ⅰ IFN signaling. Combined multi-omics analysis and qPCR identified IL-6, STAT3, and IRF3 as potential effector molecules mediating GCRV infection. Deploying GCRV challenge at 18 °C and 28 °C as models of resistant and permissive infections and switched to the corresponding temperatures as temperature stress models, we illustrated that IL-6 and STAT3 expression, genome level of GCRV, and phosphorylation of STAT3 were temperature dependent and regulated by temperature stress. Further research revealed that activating IL-6/STAT3 axis enhanced GCRV replication and suppressed the expression of IFNs, whereas blocking the axis impaired viral replication. Mechanistically, grass carp STAT3 inhibited IRF3 nuclear translocation via interacting with it, thus down-regulating IFNs expression, restraining transcriptional activation of the IFN promoter, and facilitating GCRV replication. Overall, our work sheds light on an immune evasion mechanism whereby GCRV facilitates viral replication by hijacking IL-6/STAT3 axis to down-regulate IFNs expression, thus providing a valuable reference for targeted prevention and therapy of GCRV.

Evaluating a combination treatment of NK cells and reovirus against bladder cancer cells using an in vitro assay to simulate intravesical therapy.

Intravesical treatment using either reovirus or natural killer (NK) cells serves as an efficient strategy for the treatment of bladder cancer cells (BCCs); however, corresponding monotherapies have often shown modest cytotoxicity. The potential of a locoregional combination using high-dose reovirus and NK cell therapy in an intravesical approach has not yet been studied. In this study, we evaluated the effectiveness of reoviruses and expanded NK cells (eNK) as potential strategies for the treatment of bladder cancer. The anti-tumor effects of mono-treatment with reovirus type 3 Dearing strain (RC402 and RP116) and in combination with interleukin (IL)-18/-21-pretreated eNK cells were investigated on BCC lines (5637, HT-1376, and 253J-BV) using intravesical therapy to simulate in vitro model. RP116 and IL-18/-21-pretreated eNK cells exhibited effective cytotoxicity against grade 1 carcinoma (5637 cells) when used alone, but not against HT-1376 (grade 2 carcinoma) and 253J-BV cells (derived from a metastatic site). Notably, combining RP116 with IL-18/-21-pretreated eNK cells displayed effective cytotoxicity against both HT-1376 and 253J-BV cells. Our findings underscore the potential of a combination therapy using reoviruses and NK cells as a promising strategy for treating bladder cancer.

Neutralizing Antibodies Impair the Oncolytic Efficacy of Reovirus but Permit Effective Combination with T cell-Based Immunotherapies.

Reovirus type 3 Dearing (Reo), manufactured for clinical application as pelareorep, is an attractive anticancer agent under evaluation in multiple phase 2 clinical trials for the treatment of solid tumors. It elicits its anticancer efficacy by inducing both oncolysis and intratumoral T-cell influx. Because most people have been preexposed to Reo, neutralizing antibodies (NAb) are prevalent in patients with cancer and might present a barrier to effective Reo therapy. Here, we tested serum of patients with cancer and healthy controls (n = 100) and confirmed that Reo NAbs are present in >80% of individuals. To investigate the effect of NAbs on both the oncolytic and the immunostimulatory efficacy of Reo, we established an experimental mouse model with Reo preexposure. The presence of preexposure-induced NAbs reduced Reo tumor infection and prevented Reo-mediated control of tumor growth after intratumoral Reo administration. In B cell-deficient mice, the lack of NAbs provided enhanced tumor growth control after Reo monotherapy, indicating that NAbs limit the oncolytic capacity of Reo. In immunocompetent mice, intratumoral T-cell influx was not affected by the presence of preexposure-induced NAbs and consequently, combinatorial immunotherapy strategies comprising Reo and T-cell engagers or checkpoint inhibitors remained effective in these settings, also after a clinically applied regimen of multiple intravenous pelareorep administrations. Altogether, our data indicate that NAbs hamper the oncolytic efficacy of Reo, but not its immunotherapeutic capacity. Given the high prevalence of seropositivity for Reo in patients with cancer, our data strongly advocate for the application of Reo as part of T cell-based immunotherapeutic strategies.

Grass carp superoxide dismutases exert antioxidant function and inhibit autophagy to promote grass carp reovirus (GCRV) replication.

Superoxide dismutases (SODs) are potent antioxidants crucial for neutralizing reactive oxygen species (ROS) and protecting organisms from oxidative damage. In this study, we successfully cloned and analyzed two SOD genes, CiSOD1 and CiSOD2, from grass carp (Ctenopharyngodon idellus). CiSOD1 consists of two CuZn signature motifs and two conserved cysteine residues, while CiSOD2 contains a single Mn signature motif. The expression of CiSODs was found to be ubiquitous across all examined tissues, with their expression levels significantly altered after stimulation by grass carp reovirus (GCRV) or pathogen-associated molecular patterns (PAMPs). CiSOD1 was observed to be uniformly distributed in the cytoplasm, whereas CiSOD2 localized in the mitochondria. Escherichia coli transformed with both CiSODs demonstrated enhanced host resistance to H2O2 and heavy metals. Additionally, purified recombinant CiSOD proteins effectively protected DNA against oxidative damage. Furthermore, overexpression of CiSODs in fish cells reduced intracellular ROS, inhibited autophagy, and then resulted in the promotion of GCRV replication. Knockdown of CiSODs showed opposite trends. Notably, these roles of CiSODs in autophagy and GCRV replication were reversed upon treatment with an autophagy inducer. In summary, our findings suggest that grass carp SODs play an important role in decreasing intracellular ROS levels, inhibiting autophagy, and subsequently promoting GCRV replication.

Bmp4 in Zebrafish Enhances Antiviral Innate Immunity through p38 MAPK (Mitogen-Activated Protein Kinases) Pathway.

Bone morphogenetic proteins (BMPs) are a group of structurally and functionally related signaling molecules that comprise a subfamily, belonging to the TGF-ß superfamily. Most BMPs play roles in the regulation of embryonic development, stem cell differentiation, tumor growth and some cardiovascular and cerebrovascular diseases. Although evidence is emerging for the antiviral immunity of a few BMPs, more BMPs are needed to determine whether this function is universal. Here, we identified the zebrafish bmp4 ortholog, whose expression is up-regulated through challenge with grass carp reovirus (GCRV) or its mimic poly(I:C). The overexpression of bmp4 in epithelioma papulosum cyprini (EPC) cells significantly decreased the viral titer of GCRV-infected cells. Moreover, compared to wild-type zebrafish, viral load and mortality were significantly increased in both larvae and adults of bmp4-/- mutant zebrafish infected with GCRV virus. We further demonstrated that Bmp4 promotes the phosphorylation of Tbk1 and Irf3 through the p38 MAPK pathway, thereby inducing the production of type I IFNs in response to virus infection. These data suggest that Bmp4 plays an important role in the host defense against virus infection. Our study expands the understanding of BMP protein functions and opens up new targets for the control of viral infection.

Black carp RIOK3 suppresses MDA5-mediated IFN signaling in the antiviral innate immunity.

In mammals, right open reading frame kinase 3 (RIOK3) is related with cancer development and immune regulation. To explore the role of teleost RIOK3 in the antiviral innate immunity, the homolog of RIOK3 (bcRIOK3) from black carp (Mylopharyngodon piceus) has been cloned and characterized in this study. Sequence analysis revealed that bcRIOK3 is conserved in vertebrates. The transcription of bcRIOK3 varied in host cells in response to the stimulation of spring viremia of carp virus (SVCV), poly (I:C), and LPS. Immunoblotting (IB) and immunofluorescence (IF) assays identified bcRIOK3 as a cytoplasmic protein with a molecular weight of ∼60 kDa. It was interesting that bcRIOK3 knockdown led to the decreased basal mRNA levels of IFNa, IFNb and Viperin; however, triggered obviously higher mRNA levels of the above genes after viral infection and enhanced host resistance to SVCV. Like its mammalian counterpart, bcRIOK3 overexpression in EPC cells showed a significant inhibitory effect on black carp MDA5 (bcMDA5)-mediated transcription of interferon promoters and antiviral activity. Co-immunoprecipitation and immunofluorescent assays identified the association between bcRIOK3 and bcMDA5. Further analysis revealed that bcRIOK3 enhanced the K48-linked ubiquitination and proteasome-dependent degradation of bcMDA5, and it weakened the oligomerization of bcMDA5 under poly (I:C) stimulation. In summary, our data conclude that RIOK3 dampens MDA5-mediated IFN signaling by promoting its degradation in black carp, which provide new insights into the regulation of IFN signaling in teleost.

The Identification of Viral Pathogens in a Physostegia virginiana Plant Using High-Throughput RNA Sequencing.

Physostegia virginiana is an important ornamental and cut-flower plant in China. Its commonly used method of clonal propagation leads to virus accumulation in this plant. However, which viruses can infect the Physostegia virginiana plant remains to be illuminated. In this work, five viral pathogens in a Physostegia virginiana plant with virus-like symptoms of yellow, shriveled, and curled leaves were identified using RNA-seq, bioinformatics, and molecular biological techniques. These techniques allowed us to identify five viruses comprising one known alfalfa mosaic virus (AMV) and four novel viruses. The novel viruses include a virus belonging to the genus Fabavirus, temporarily named Physostegia virginiana crinkle-associated virus 1 (PVCaV1); two viruses belonging to the genus Caulimovirus, temporarily named Physostegia virginiana caulimovirus 1 and 2 (PVCV1 and PVCV2); and a virus belonging to the genus Fijivirus, temporarily named Physostegia virginiana fijivirus (PVFV). The genome sequences of PVCaV1, PVCV1, and PVCV2, and the partial genome sequence of PVFV were identified. Genome organizations and genetic evolutionary relationships of all four novel viruses were analyzed. PVCaV1 has a relatively close evolutionary relationship with five analyzed fabiviruses. PVCV1 and PVCV2 have separately a closest evolutionary relationship with lamium leaf distortion-associated virus (LLDAV) and figwort mosaic virus (FMV), and PVFV has a close evolutionary relationship with the five analyzed fijiviruses. Additionally, PVCaV1 can infect Nicotiana benthamiana plants via friction inoculation. The findings enrich our understanding of Physostegia virginiana viruses and contribute to the prevention and control of Physostegia virginiana viral diseases.

Reovirus infection of tumor cells reduces the expression of NKG2D ligands, leading to impaired NK-cell cytotoxicity and functionality.

In recent years, reoviruses have been of major interest in immunotherapy because of their oncolytic properties. Preclinical and clinical trials, in which reovirus was used for the treatment of melanoma and glioblastoma, have paved the way for future clinical use of reovirus. However, little is known about how reovirus infection affects the tumor microenvironment and immune response towards infected tumor cells. Studies have shown that reovirus can directly stimulate natural killer (NK) cells, but how reovirus affects cellular ligands on tumor cells, which are ultimately key to tumor recognition and elimination by NK cells, has not been investigated. We tested how reovirus infection affects the binding of the NK Group-2 member D (NKG2D) receptor, which is a dominant mediator of NK cell anti-tumor activity. Using models of human-derived melanoma and glioblastoma tumors, we demonstrated that NKG2D ligands are downregulated in tumor cells post-reovirus-infection due to the impaired translation of these ligands in reovirus-infected cells. Moreover, we showed that downregulation of NKG2D ligands significantly impaired the binding of NKG2D to infected tumor cells. We further demonstrated that reduced recognition of NKG2D ligands significantly alters NK cell anti-tumor cytotoxicity in human primary NK cells and in the NK cell line NK-92. Thus, this study provides novel insights into reovirus-host interactions and could lead to the development of novel reovirus-based therapeutics that enhance the anti-tumor immune response.

Reovirus combined with a STING agonist enhances anti-tumor immunity in a mouse model of colorectal cancer.

Reovirus, a naturally occurring oncolytic virus, initiates the lysis of tumor cells while simultaneously releasing tumor antigens or proapoptotic cytokines in the tumor microenvironment to augment anticancer immunity. However, reovirus has developed a strategy to evade antiviral immunity via its inhibitory effect on interferon production, which negatively affects the induction of antitumor immune responses. The mammalian adaptor protein Stimulator of Interferon Genes (STING) was identified as a key regulator that orchestrates immune responses by sensing cytosolic DNA derived from pathogens or tumors, resulting in the production of type I interferon. Recent studies reported the role of STING in innate immune responses to RNA viruses leading to the restriction of RNA virus replication. In the current study, we found that reovirus had a reciprocal reaction with a STING agonist regarding type I interferon responses in vitro; however, we found that the combination of reovirus and STING agonist enhanced anti-tumor immunity by enhancing cytotoxic T cell trafficking into tumors, leading to significant tumor regression and survival benefit in a syngeneic colorectal cancer model. Our data indicate the combination of reovirus and a STING agonist to enhance inflammation in the tumor microenvironment might be a strategy to improve oncolytic reovirus immunotherapy.

Modulation of Reoviral Cytolysis (I): Combination Therapeutics.

Patients with stage IV gastric cancer suffer from dismal outcomes, a challenge especially in many Asian populations and for which new therapeutic options are needed. To explore this issue, we used oncolytic reovirus in combination with currently used chemotherapeutic drugs (irinotecan, paclitaxel, and docetaxel) for the treatment of gastric and other gastrointestinal cancer cells in vitro and in a mouse model. Cell viability in vitro was quantified by WST-1 assays in human cancer cell lines treated with reovirus and/or chemotherapeutic agents. The expression of reovirus protein and caspase activity was determined by flow cytometry. For in vivo studies, athymic mice received intratumoral injections of reovirus in combination with irinotecan or paclitaxel, after which tumor size was monitored. In contrast to expectations, we found that reoviral oncolysis was only poorly correlated with Ras pathway activation. Even so, the combination of reovirus with chemotherapeutic agents showed synergistic cytopathic effects in vitro, plus enhanced reovirus replication and apoptosis. In vivo experiments showed that reovirus alone can reduce tumor size and that the combination of reovirus with chemotherapeutic agents enhances this effect. Thus, we find that oncolytic reovirus therapy is effective against gastric cancer. Moreover, the combination of reovirus and chemotherapeutic agents synergistically enhanced cytotoxicity in human gastric cancer cell lines in vitro and in vivo. Our data support the use of reovirus in combination with chemotherapy in further clinical trials, and highlight the need for better biomarkers for reoviral oncolytic responsiveness.

Modulation of Reoviral Cytolysis (II): Cellular Stemness.

Oncolytic viruses (OVs) are an emerging cancer therapeutic that are intended to act by selectively targeting and lysing cancerous cells and by stimulating anti-tumour immune responses, while leaving normal cells mainly unaffected. Reovirus is a well-studied OV that is undergoing advanced clinical trials and has received FDA approval in selected circumstances. However, the mechanisms governing reoviral selectivity are not well characterised despite many years of effort, including those in our accompanying paper where we characterize pathways that do not consistently modulate reoviral cytolysis. We have earlier shown that reovirus is capable of infecting and lysing both certain types of cancer cells and also cancer stem cells, and here we demonstrate its ability to also infect and kill healthy pluripotent stem cells (PSCs). This led us to hypothesize that pathways responsible for stemness may constitute a novel route for the modulation of reoviral tropism. We find that reovirus is capable of killing both murine and human embryonic and induced pluripotent stem cells. Differentiation of PSCs alters the cells' reoviral-permissive state to a resistant one. In a breast cancer cell line that was resistant to reoviral oncolysis, induction of pluripotency programming rendered the cells permissive to cytolysis. Bioinformatic analysis indicates that expression of the Yamanaka pluripotency factors may be associated with regulating reoviral selectivity. Mechanistic insights from these studies will be useful for the advancement of reoviral oncolytic therapy.

Nonstructural protein NS17 of grass carp reovirus Honghu strain promotes virus infection by mediating cell-cell fusion and apoptosis.

Fusion-associated small transmembrane (FAST) proteins can promote cell fusion, alter membrane permeability and trigger apoptosis to promote virus proliferation in orthoreoviruses. However, it is unknown whether FAST proteins perform these functions in aquareoviruses (AqRVs). Non-structural protein 17 (NS17) carried by grass carp reovirus Honghu strain (GCRV-HH196) belongs to the FAST protein family, and we preliminarily explored its relevance to virus infection. NS17 has similar domains to FAST protein NS16 of GCRV-873, comprising a transmembrane domain, a polybasic cluster, a hydrophobic patch and a polyproline motif. It was observed in the cytoplasm and the cell membrane. Overexpression of NS17 enhanced the efficiency of cell-cell fusion induced by GCRV-HH196 and promoted virus replication. Overexpression of NS17 also led to DNA fragmentation and reactive oxygen species (ROS) accumulation, and it triggered apoptosis. The findings illuminate the functions of NS17 in GCRV infection, and provide a reference for the development of novel antiviral strategies.

Transcriptome Analysis of the Spleen Provides Insight into the Immune Regulation of GCRV Resistance in Grass Carp (Ctenopharyngodon idella).

Grass carp (Ctenopharyngodon idella) is one of the most economically important fish in China, and its production is commonly lost due to GCRV infection. To understand the molecular mechanism of GCRV resistance in grass carp, we compared the spleen transcriptome of the GCRV-resistant and susceptible individuals under GCRV infection (Res-Sus) and the GCRV-resistant individuals under different conditions of injection with GCRV and PBS (Res-Ctl). A total of 87.56 GB of clean data were obtained from 12 transcriptomic libraries of spleen tissues. A total of 379 DEGs (156 upregulated genes and 223 downregulated genes) were identified in the comparison group Res-Ctl. A total of 1207 DEGs (633 upregulated genes and 574 downregulated genes) were identified in the comparison group Res-Sus. And 54 DEGs were shared including immune-related genes of stc2 (stanniocalcin 2), plxna1 (plexin A1), ifnα (interferon alpha), cxcl 11 (C-X-C motif chemokine ligand 11), ngfr (nerve growth factor receptor), mx (MX dynamin-like GTPase), crim1 (cysteine-rich transmembrane BMP regulator 1), plxnb2 (plexin B2), and slit2 (slit guidance ligand 2). KEGG pathway analysis revealed significant differences in the expression of genes mainly involved in immune system and signal transduction, including antigen processing and presentation, Toll-like receptor signaling pathway, natural killer cell-mediated cytotoxicity, and Hippo signaling pathway. This study investigates the immune mechanism of the resistance to GCRV infection in grass carp and provides useful information for the development of methods to control the spread of the GCRV infection.

Metabolomics in rare minnow (Gobiocypris rarus) after infection by attenuated and virulent grass carp reovirus genotype â ¡.

Grass carp reovirus genotype Ⅱ (GCRV Ⅱ) causes hemorrhagic disease in a variety fish, seriously affecting the aquaculture industry in China. However, the pathogenesis of GCRV Ⅱ is unclear. Rare minnow is an ideal model organism to study the pathogenesis of GCRV Ⅱ. Herein, we applied liquid chromatography-tandem mass spectrometry metabolomics to investigate metabolic responses in the spleen and hepatopancreas of rare minnow injected with virulent GCRV Ⅱ isolate DY197 and attenuated isolate QJ205. Results indicated that marked metabolic changes were identified in both the spleen and hepatopancreas after GCRV Ⅱ infection, and the virulent DY197 strain induced more significantly different metabolites (SDMs) than the attenuated QJ205 strain. Moreover, most SDMs were downregulated in the spleen and tend to be upregulated in hepatopancreas. The Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed that tissue-specific metabolic responses were identified after viruses infection, and the virulent DY197 strain induced more SDMs involved in amino acid metabolism in the spleen, especially the tryptophan metabolism, cysteine and methionine metabolism, which were essential for immune regulation in host; Meanwhile, nucleotide metabolism, protein synthesis and metabolism related pathways were enriched in the hepatopancreas by both virulent and attenuated strains. Our findings revealed the large scale metabolic alterations in rare minnow in response to attenuated and virulent GCRV Ⅱ infection, which will lead to a better understanding of the pathogenesis of viruses and host-pathogens interactions.

Deciphering molecular mechanisms stabilizing the reovirus-binding complex.

Mammalian orthoreoviruses (reoviruses) serve as potential triggers of celiac disease and have oncolytic properties, making these viruses potential cancer therapeutics. Primary attachment of reovirus to host cells is mainly mediated by the trimeric viral protein, σ1, which engages cell-surface glycans, followed by high-affinity binding to junctional adhesion molecule-A (JAM-A). This multistep process is thought to be accompanied by major conformational changes in σ1, but direct evidence is lacking. By combining biophysical, molecular, and simulation approaches, we define how viral capsid protein mechanics influence virus-binding capacity and infectivity. Single-virus force spectroscopy experiments corroborated by in silico simulations show that GM2 increases the affinity of σ1 for JAM-A by providing a more stable contact interface. We demonstrate that conformational changes in σ1 that lead to an extended rigid conformation also significantly increase avidity for JAM-A. Although its associated lower flexibility impairs multivalent cell attachment, our findings suggest that diminished σ1 flexibility enhances infectivity, indicating that fine-tuning of σ1 conformational changes is required to successfully initiate infection. Understanding properties underlying the nanomechanics of viral attachment proteins offers perspectives in the development of antiviral drugs and improved oncolytic vectors.

Oncolytic Reoviruses: Can These Emerging Zoonotic Reoviruses Be Tamed and Utilized?

Orthoreovirus is a nonenveloped double-stranded RNA virus under the Reoviridae family. This group of viruses, especially mammalian orthoreovirus (MRV), are reported with great therapeutic values due to their oncolytic effects. In this review, the life cycle and oncolytic effect of MRV and a few emerging reoviruses were summarized. This article also highlights the challenges and strategies of utilizing MRV and the emerging reoviruses, avian orthoreovirus (ARV) and pteropine orthoreovirus (PRV), as oncolytic viruses (OVs). Besides, the emergence of potential ARV and PRV as OVs were discussed in comparison to MRV. Finally, the risk of reovirus as zoonosis or reverse zoonosis (zooanthroponosis) were debated, and concerns were raised in this article, which warrant continue surveillance of reovirus (MRV, ARV, and PRV) in animals, humans, and the environment.

An aquatic virus exploits the IL6-STAT3-HSP90 signaling axis to promote viral entry.

Viral seasonality in the aquaculture industry is an important scientific issue for decades. While the molecular mechanisms underpinning the temperature-dependent pathogenesis of aquatic viral diseases remain largely unknown. Here we report that temperature-dependent activation of IL6-STAT3 signaling was exploited by grass carp reovirus (GCRV) to promote viral entry via increasing the expression of heat shock protein 90 (HSP90). Deploying GCRV infection as a model system, we discovered that GCRV induces the IL6-STAT3-HSP90 signaling activation to achieve temperature-dependent viral entry. Further biochemical and microscopic analyses revealed that the major capsid protein VP7 of GCRV interacted with HSP90 and relevant membrane-associated proteins to boost viral entry. Accordingly, exogenous expression of either IL6, HSP90, or VP7 in cells increased GCRV entry in a dose-dependent manner. Interestingly, other viruses (e.g., koi herpesvirus, Rhabdovirus carpio, Chinese giant salamander iridovirus) infecting ectothermic vertebrates have evolved a similar mechanism to promote their infection. This work delineates a molecular mechanism by which an aquatic viral pathogen exploits the host temperature-related immune response to promote its entry and replication, instructing us on new ways to develop targeted preventives and therapeutics for aquaculture viral diseases.

A Fijivirus Major Viroplasm Protein Shows RNA-Stimulated ATPase Activity by Adopting Pentameric and Hexameric Assemblies of Dimers.

Fijiviruses replicate and package their genomes within viroplasms in a process involving RNA-RNA and RNA-protein interactions. Here, we demonstrate that the 24 C-terminal residues (C-arm) of the P9-1 major viroplasm protein of the mal de Río Cuarto virus (MRCV) are required for its multimerization and the formation of viroplasm-like structures. Using an integrative structural approach, the C-arm was found to be dispensable for P9-1 dimer assembly but essential for the formation of pentamers and hexamers of dimers (decamers and dodecamers), which favored RNA binding. Although both P9-1 and P9-1ΔC-arm catalyzed ATP with similar activities, an RNA-stimulated ATPase activity was only detected in the full-length protein, indicating a C-arm-mediated interaction between the ATP catalytic site and the allosteric RNA binding sites in the (do)decameric assemblies. A stronger preference to bind phosphate moieties in the decamer was predicted, suggesting that the allosteric modulation of ATPase activity by RNA is favored in this structural conformation. Our work reveals the structural versatility of a fijivirus major viroplasm protein and provides clues to its mechanism of action. IMPORTANCE The mal de Río Cuarto virus (MRCV) causes an important maize disease in Argentina. MRCV replicates in several species of Gramineae plants and planthopper vectors. The viral factories, also called viroplasms, have been studied in detail in animal reovirids. This work reveals that a major viroplasm protein of MRCV forms previously unidentified structural arrangements and provides evidence that it may simultaneously adopt two distinct quaternary assemblies. Furthermore, our work uncovers an allosteric communication between the ATP and RNA binding sites that is favored in the multimeric arrangements. Our results contribute to the understanding of plant reovirids viroplasm structure and function and pave the way for the design of antiviral strategies for disease control.

Oncolytic viruses as emerging therapy against cancers including Oncovirus-induced cancers.

There are several human viruses with known potential for causing cancers including, Hepatitis B virus, Hepatitis C virus, Epstein-Barr virus, Kaposi's sarcoma herpesvirus, Human T-cell lymphotropic virus, Human papillomavirus, and Merkel cell polyomavirus. Cancer is the second leading cause of death that affects humans worldwide, especially in developing countries. Surgery, chemotherapy, and radiotherapy can cure about 60% of humans with cancer but recurrent and metastatic diseases remain a major reason for death. In recent years, understanding the molecular characteristics of cancer cells has led to the improvement of therapeutic strategies using novel emerging therapies. Oncolytic viruses with the potential of lysing cancer cells defined the field of oncolytic virology, hence becoming a biotechnology tool rather than just a cause of disease. This study mainly focused on targeting cell proliferation and death pathways in human tumor-inducing viruses by developing innovative therapies for cancer patients based on the natural oncolytic properties of reovirus. To kill tumor cells efficiently and reduce the chance of recurrence both the direct ability of reovirus infection to lyse the tumor cells and the stimulation of a potent host immune response are applied. Hence, bioengineered stem cells can be used as smart carriers to improve the efficacy of oncolytic reovirus and safety profiles.