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Clinical data from over two decades, involving more than 3000 treated patients, demonstrate that adeno‐associated virus (AAV) gene therapy is a safe, effective, and well‐tolerated therapeutic method. Clinical trials using AAV‐mediated gene delivery to accessible tissues have led to successful treatments for numerous monogenic disorders and advancements in tissue engineering. Although the US Food and Drug Administration (FDA) has approved AAV for clinical use, systemic administration remains a significant challenge. In this review, we delve into AAV biology, focusing on current manufacturing technologies and transgene engineering strategies. We examine the use of AAVs in ongoing clinical trials for ocular, neurological, and hematological disorders, as well as cancers. By discussing recent advancements and current challenges in the field, we aim to provide valuable insights for researchers and clinicians navigating the evolving landscape of AAV‐based gene therapy. [ABSTRACT FROM AUTHOR]

Abstract In the increasing demand for virus vaccines, large-scale production of safe, efficient, and economical viral antigens has become a significant challenge. High-cell-density manufacturing processes are the most commonly used to produce vaccine antigens and protein drugs. However, the cellular stress response in large-scale cell culture may directly affect host cell growth and metabolism, reducing antigen production and increasing production costs. This study provided a novel strategy of the antioxidant auxiliary system (AAS) to supply molecular hydrogen (H2) into the cell culture media via proton exchange membrane (PEM) electrolysis. Integrated with a high-density cell bioreactor, the AAS aims to alleviate cellular stress response and increase viral vaccine production. In the results, the AAS stably maintained H2 concentration in media even in the high-air exposure tiding cell bioreactor. H2 treatment was shown safe to cell culture and effectively alleviated oxidative stress. In two established virus cultures models, bovine epidemic fever virus (BEFV) and porcine circovirus virus type 2 (PCV-2), were employed to verify the efficacy of AAS. The virus yield was increased by 3.7 and 2.5 folds in BEFV and PCV-2 respectively. In conclusion, the AAS-connected bioreactor effectively alleviated cellular oxidative stress and enhanced virus production in high-density cell culture.

In the increasing demand for virus vaccines, large-scale production of safe, efficient, and economical viral antigens has become a significant challenge. High-cell-density manufacturing processes are the most commonly used to produce vaccine antigens and protein drugs. However, the cellular stress response in large-scale cell culture may directly affect host cell growth and metabolism, reducing antigen production and increasing production costs. This study provided a novel strategy of the antioxidant auxiliary system (AAS) to supply molecular hydrogen (H2) into the cell culture media via proton exchange membrane (PEM) electrolysis. Integrated with a high-density cell bioreactor, the AAS aims to alleviate cellular stress response and increase viral vaccine production. In the results, the AAS stably maintained H2 concentration in media even in the high-air exposure tiding cell bioreactor. H2 treatment was shown safe to cell culture and effectively alleviated oxidative stress. In two established virus cultures models, bovine epidemic fever virus (BEFV) and porcine circovirus virus type 2 (PCV-2), were employed to verify the efficacy of AAS. The virus yield was increased by 3.7 and 2.5 folds in BEFV and PCV-2 respectively. In conclusion, the AAS-connected bioreactor effectively alleviated cellular oxidative stress and enhanced virus production in high-density cell culture. [ABSTRACT FROM AUTHOR]

Abstract Although we have gained insight into the biological and biochemical effects of natural sunlight exposure on prokaryotes, little is known about sunlight exposure on natural virus communities. To address this question, an investigation of the effects of sunlight and dark treatments on viral communities, viral production and decay rates in Kenting coastal waters was conducted. The average rate of net viral production in the sunlight and dark treatment was 0.010 and 0.018 × 106 viruses mL–1 h− 1, respectively. Furthermore, averaged value for viral decay in the sunlight treatment was 0.016 × 106 viruses mL− 1 h− 1, a significant decrease (ca. 60%=((0.83 − 0.33)/(0.83 × 100%)) was observed in sunlight conditions, whereas no significant changes occurred in dark conditions. The gross viral production under sunlight conditions was slightly higher, however, non-significantly higher than that in the dark treatment. As a result, we suggest that sunlight damages a large portion of the natural viral community, affecting the role viruses play in food webs.

Although we have gained insight into the biological and biochemical effects of natural sunlight exposure on prokaryotes, little is known about sunlight exposure on natural virus communities. To address this question, an investigation of the effects of sunlight and dark treatments on viral communities, viral production and decay rates in Kenting coastal waters was conducted. The average rate of net viral production in the sunlight and dark treatment was 0.010 and 0.018 × 106 viruses mL–1 h− 1, respectively. Furthermore, averaged value for viral decay in the sunlight treatment was 0.016 × 106 viruses mL− 1 h− 1, a significant decrease (ca. 60%=((0.83 − 0.33)/(0.83 × 100%)) was observed in sunlight conditions, whereas no significant changes occurred in dark conditions. The gross viral production under sunlight conditions was slightly higher, however, non-significantly higher than that in the dark treatment. As a result, we suggest that sunlight damages a large portion of the natural viral community, affecting the role viruses play in food webs. [ABSTRACT FROM AUTHOR]

Numerous viruses manipulate host factors for viral production. We demonstrated that human enterovirus A71 (EVA71), a primary causative agent for hand, foot, and mouth disease (HFMD), increased the level of the DNA damage response (DDR) marker γ-H2AX. DDR is primarily mediated by the ataxia telangiectasia mutated (ATM), ATM and Rad3-related (ATR), or DNA-dependent protein kinase (DNA-PK) pathways. Upregulation of γ-H2AX by EVA71 was dependent on the ATR but not the ATM or DNA-PK pathway. As a nuclear factor, there is no previous evidence of cytoplasmic distribution of γ-H2AX. However, the present findings demonstrated that EVA71 encouraged the localization of γ-H2AX to the cytoplasm. Of note, γ-H2AX formed a complex with structural protein VP3, non-structural protein 3D, and the viral genome. Treatment with an inhibitor or CRISPR/Cas9 technology to decrease or silence the expression of γ-H2AX decreased viral genome replication in host cells; this effect was accompanied by decreased viral protein expression and virions. In animal experiments, caffeine was used to inhibit DDR; the results revealed that caffeine protected neonatal mice from death after infection with EVA71, laying the foundation for new therapeutic applications of caffeine. More importantly, in children with HFMD, γ-H2AX was upregulated in peripheral blood lymphocytes. The consistent in vitro and in vivo data on γ-H2AX from this study suggested that caffeine or other inhibitors of DDR might be novel therapeutic agents for HFMD.

The transactive response DNA-binding protein (TARDBP/TDP-43) is known to stabilize the anti-HIV-1 factor, histone deacetylase 6 (HDAC6). TDP-43 has been reported to determine cell permissivity to HIV-1 fusion and infection acting on tubulin-deacetylase HDAC6. Here, we studied the functional involvement of TDP-43 in the late stages of the HIV-1 viral cycle. The overexpression of TDP-43, in virus-producing cells, stabilized HDAC6 (i.e., mRNA and protein) and triggered the autophagic clearance of HIV-1 Pr55Gag and Vif proteins. These events inhibited viral particle production and impaired virion infectiveness, observing a reduction in the amount of Pr55Gag and Vif proteins incorporated into virions. A nuclear localization signal (NLS)-TDP-43 mutant was not able to control HIV-1 viral production and infection. Likewise, specific TDP-43-knockdown reduced HDAC6 expression (i.e., mRNA and protein) and increased the expression level of HIV-1 Vif and Pr55Gag proteins and α-tubulin acetylation. Thus, TDP-43 silencing favored virion production and enhanced virus infectious capacity, thereby increasing the amount of Vif and Pr55Gag proteins incorporated into virions. Noteworthy, there was a direct relationship between the content of Vif and Pr55Gag proteins in virions and their infection capacity. Therefore, for TDP-43, the TDP-43/HDAC6 axis could be considered a key factor to control HIV-1 viral production and virus infectiveness. [ABSTRACT FROM AUTHOR]

Viral dynamics are the result of the balance between the rates of viral production and decay. Here, we have carried out independent measurements of viral production and decay rates in different depths of the southern East China Sea in summer (August and October 2021). In this study, the prevalence of viral abundance at the surface waters (14.2~27.6 × 105 viruses mL−1) was significantly higher than the bottom of the euphotic zone (2.9~12.6 × 105 viruses mL−1). As for viruses to bacteria ratio (VBR) values, we found a wide variability both at the surface (1.4 to 3.2) and bottom of the euphotic zone (2.1 to 16.2). The results of our study showed that at all stations examined, in the southern East China Sea, the values of gross viral production (GVP) were significantly higher in the sunlit surfaces compared to the bottom of the euphotic zone. In particular, our analysis indicates that no significant viral decay rates (VD) were observed in some regions at the bottom of the euphotic zone. Here, we also provide a budget for viral abundance and net viral production in different regions in the southern East China Sea. The GVP or VD is not applicable in our case to explain VBR is high at bottom of the euphotic zone. The mechanisms underlying VBR uncoupling, viral production, and viral loss in marine systems are still being investigated. [ABSTRACT FROM AUTHOR]

Although we have gained insight into the biological and biochemical effects of natural sunlight exposure on prokaryotes, little is known about sunlight exposure on natural virus communities. To address this question, an investigation of the effects of sunlight and dark treatments on viral communities, viral production and decay rates in Kenting coastal waters was conducted. The average rate of net viral production in the sunlight and dark treatment was 0.010 and 0.018 x 106 viruses mL-1 h-1, respectively. Furthermore, averaged value for viral decay in the sunlight treatment was 0.016 x 106 viruses mL-1 h-1, a significant decrease (ca. 60%=((0.83 - 0.33)/(0.83 x 100%)) was observed in sunlight conditions, whereas no significant changes occurred in dark conditions. The gross viral production under sunlight conditions was slightly higher, however, non-significantly higher than that in the dark treatment. As a result, we suggest that sunlight damages a large portion of the natural viral community, affecting the role viruses play in food webs. [ABSTRACT FROM AUTHOR]

Recombinant adeno-associated virus (rAAV) vectors are an effective and well-established tool in the growing gene therapy field, with five FDA-approved AAV-mediated gene therapies already on the market and numerous more in clinical trials. However, manufacturing rAAV vectors is an expensive, timely, and labor-intensive process that limits the commercial use of AAV-mediated gene therapies. To address this limitation, we screened producer cells for genes that could be targeted to increase rAAV yield. Specifically, we performed a CRISPR-based genome-wide knockout screen in HEK 293 cells using an antibody specific to intact AAV2 capsids coupled with flow cytometry to identify genes that modulate rAAV production. We discovered that the knockout of a group of heparan sulfate biosynthesis genes previously implicated in rAAV infectivity decreased rAAV production. Additionally, we identified several vesicular trafficking proteins for which knockout in HEK 293 cells increased rAAV yields. Our findings provide evidence that host proteins associated with viral infection may have also been co-opted for viral assembly and that the genetic makeup of viral producer cells can be manipulated to increase particle yield., Competing Interests: DECLARATION OF INTERESTS: B.L.D. serves on the advisory board of Latus Biosciences, Patch Bio, Spirovant Biosciences, Resilience, and Carbon Biosciences and has sponsored research unrelated to this work from Roche, Latus, and Spirovant. Authors have filed a patent related to this manuscript through the Children’s Hospital of Philadelphia.

The transactive response DNA-binding protein (TARDBP/TDP-43) is known to stabilize the anti-HIV-1 factor, histone deacetylase 6 (HDAC6). TDP-43 has been reported to determine cell permissivity to HIV-1 fusion and infection acting on tubulin-deacetylase HDAC6. Here, we studied the functional involvement of TDP-43 in the late stages of the HIV-1 viral cycle. The overexpression of TDP-43, in virus-producing cells, stabilized HDAC6 (i.e., mRNA and protein) and triggered the autophagic clearance of HIV-1 Pr55Gag and Vif proteins. These events inhibited viral particle production and impaired virion infectiveness, observing a reduction in the amount of Pr55Gag and Vif proteins incorporated into virions. A nuclear localization signal (NLS)-TDP-43 mutant was not able to control HIV-1 viral production and infection. Likewise, specific TDP-43-knockdown reduced HDAC6 expression (i.e., mRNA and protein) and increased the expression level of HIV-1 Vif and Pr55Gag proteins and α-tubulin acetylation. Thus, TDP-43 silencing favored virion production and enhanced virus infectious capacity, thereby increasing the amount of Vif and Pr55Gag proteins incorporated into virions. Noteworthy, there was a direct relationship between the content of Vif and Pr55Gag proteins in virions and their infection capacity. Therefore, for TDP-43, the TDP-43/HDAC6 axis could be considered a key factor to control HIV-1 viral production and virus infectiveness.

Viral dynamics are the result of the balance between the rates of viral production and decay. Here, we have carried out independent measurements of viral production and decay rates in different depths of the southern East China Sea in summer (August and October 2021). In this study, the prevalence of viral abundance at the surface waters (14.2~27.6 × 105 viruses mL−1) was significantly higher than the bottom of the euphotic zone (2.9~12.6 × 105 viruses mL−1). As for viruses to bacteria ratio (VBR) values, we found a wide variability both at the surface (1.4 to 3.2) and bottom of the euphotic zone (2.1 to 16.2). The results of our study showed that at all stations examined, in the southern East China Sea, the values of gross viral production (GVP) were significantly higher in the sunlit surfaces compared to the bottom of the euphotic zone. In particular, our analysis indicates that no significant viral decay rates (VD) were observed in some regions at the bottom of the euphotic zone. Here, we also provide a budget for viral abundance and net viral production in different regions in the southern East China Sea. The GVP or VD is not applicable in our case to explain VBR is high at bottom of the euphotic zone. The mechanisms underlying VBR uncoupling, viral production, and viral loss in marine systems are still being investigated.

In spite of the fact that the interactions between environmental parameters and prokaryotic and viral abundance have been explored in various aquatic environments, only a few independent estimates of viral production and decay in the estuarine region have been explored. In this study, data were analyzed for viral and prokaryotic abundance, viral production, and viral decay in a subtropical Danshui estuary in summer 2021. Prokaryotic abundance varied from 2.4 ± 0.6 × 105 to 12 ± 2.3 × 105 cells mL−1, and viral abundance ranged from 2.3 ± 0.9 × 105 to 6.9 ± 1.3 × 105 viruses mL−1 during the study period. Viral abundance was significantly correlated with prokaryotic abundance and chlorophyll a concentration. Furthermore, studies of changes in viral to prokaryotic abundance ratio (VPR) ranged from 0.42 ± 0.11 to 2.0 ± 0.25. Viral decay values were 2.1 ± 0.5 and 2.1 ± 0.3 × 104 virus mL−1h−1, and non-significant differences were observed between the inner estuary and coastal water region. Viral decay almost balanced gross viral production in this study. The dilution experiments revealed non-significant net viral production in July; thus, a lower VPR might be explained in this estuarine environment. [ABSTRACT FROM AUTHOR]

In freshwater environments, limited data exist on the impact of mortality forces (viruses and heterotrophic nanoflagellates) on bacterial growth efficiency (BGE, index of bacterial carbon metabolism) compared to resource availability. An investigation to determine the relative influence of viral lysis and flagellate predation (top-down forces) on BGE was conducted in a mesotrophic freshwater system (Lake Goule, France) with time and space. Viral abundance was significantly (p < 0.001) related to bacterial abundance by a power law function with an exponent less than 1, emphasizing that the increases in host population (bacteria) together with viruses were not proportionate. A lytic viral strategy was evident throughout the study period, with high lysis of the bacterial population (up to 60%) supported by viral production rates. Viral processes (lysis and production) that were influenced by bacterial production and heterotrophic nanoflagellate abundance had a positive impact on BGE. Estimates of BGE were variable (9.9–45.5%) due to uncoupling between two metabolic parameters—namely bacterial production and respiration. The existence of a synergistic relationship between viruses and flagellates with bacteria in Lake Goule highlighted the decisive impact of top-down agents in sustaining the bacterial carbon metabolism of non-infected population through the nature of vital resources released via mortality processes. [ABSTRACT FROM AUTHOR]

Viral processes in the coastal and offshore regions of the Eastern Arabian Sea (west coast of India) and their contribution of viral lysis to the organic carbon and nitrogen pool was examined. Water samples were collected during the southwest monsoon season at different depths (up to 1000 m) from four transects, with each transect consisting of two sampling points (S1–S8). Abundances of viruses and prokaryotes together with viral mediated prokaryotic mortality (up to 49.7%) were significantly (p < 0.001) higher in eutrophic coastal stations, whereas high percent lysogeny (up to 93%) was observed in the offshore regions. High viral-mediated carbon (Mean ± SD = 67.47 ± 2.0 μM C L−1 d−1) and nitrogen (Mean ± SD = 13.49 ± 14.0 μM N L−1 d−1) release was evident in the surface coastal waters compared to offshore regions. The percentage contributions of carbon and nitrogen released by viral lysis to the total dissolved organic carbon and nitrogen pool were estimated to be 7.4% and 3.9%, respectively, in the coastal surface waters. Our findings suggest that the contribution of viral lysis to DOM production through viral shunt could be crucial for the cycling of major biogeochemical elements and functioning of the studied tropical ecosystem. [ABSTRACT FROM AUTHOR]

Kaposi's sarcoma-associated herpesvirus (KSHV) is an etiologic agent of Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman disease. In studies of KSHV, efficient virus production and isolation are essential. Reactivation of KSHV can be initiated by treating latently infected cells with chemicals, such as 12-O-tetradecanoyl-phorbol-13-acetate and sodium butyrate. These chemicals have been used as tools to induce lytic replication and viral production in KSHV-producing cell lines. Dimethyl sulfoxide (DMSO) is an organosulfur compound that is frequently used as an aprotic solvent similar to water. In experiments exploring signaling pathways in KSHV-infected cells, DMSO treatment alone as a vehicle affected the lytic gene expression of KSHV. However, to the best of our knowledge, the effects of DMSO on KSHV-producing cells have not yet been reported. Therefore, in this study, we investigated whether DMSO could be used as a reagent to enhance viral production during lytic replication in KSHV-producing cells and assessed the underlying mechanisms. The effects of DMSO on KSHV production were analyzed in iSLK BAC16 cells, which have been widely used for recombinant KSHV production. We found that the production of KSHV virions was significantly increased by treatment with DMSO during the induction of lytic replication. Mechanistically, lytic genes of KSHV were enhanced by DMSO treatment, which was correlated with virion production. Additionally, DMSO induced the phosphorylation of JNK during lytic replication, and inhibition of JNK abolished the effects of DMSO on lytic replication and virion production. Our findings showed that additional treatment with DMSO during the induction of lytic replication significantly improved the yield of KSHV production. [ABSTRACT FROM AUTHOR]

Viral production is a key parameter for assessing virus-mediated biogeochemical cycles. One widely used method for the determination of viral production, called the virus reduction assay, reduces viral abundance, while maintaining bacterial abundance, using 0.2-μm pore-size filters. Viral production is estimated from the increase of viral abundance during incubation. We hypothesized that small-cell-sized bacterial communities can pass through 0.2-μm filters and drive viral production, representing a missing fraction of viral production that is missed by the virus reduction assay. Coastal seawater was filtered through 0.2-μm filters and diluted with virus-free seawater. Viral production in the <0.2-μm filtrate was estimated from changes in viral abundance determined through flow cytometry. We found that viruses were produced in the <0.2-μm communities, which were strongly enriched with low nucleic acid content bacteria. Estimated viral production in the <0.2-μm filtrates accounted for up to 43% of total viral production and 10% of dissolved organic carbon production mediated by viral lysis of bacterial cells. By not considering viral production in these <0.2-μm communities, the virus reduction assay may underestimate viral production. Virus–bacteria interactions in <0.2-μm communities may represent a significant and overlooked role of viruses in marine food webs and carbon fluxes. [ABSTRACT FROM AUTHOR]

Kaposi’s sarcoma-associated herpesvirus (KSHV) is an etiologic agent of Kaposi’s sarcoma, primary effusion lymphoma, and multicentric Castleman disease. In studies of KSHV, efficient virus production and isolation are essential. Reactivation of KSHV can be initiated by treating latently infected cells with chemicals, such as 12-O-tetradecanoyl-phorbol-13-acetate and sodium butyrate. These chemicals have been used as tools to induce lytic replication and viral production in KSHV-producing cell lines. Dimethyl sulfoxide (DMSO) is an organosulfur compound that is frequently used as an aprotic solvent similar to water. In experiments exploring signaling pathways in KSHV-infected cells, DMSO treatment alone as a vehicle affected the lytic gene expression of KSHV. However, to the best of our knowledge, the effects of DMSO on KSHV-producing cells have not yet been reported. Therefore, in this study, we investigated whether DMSO could be used as a reagent to enhance viral production during lytic replication in KSHV-producing cells and assessed the underlying mechanisms. The effects of DMSO on KSHV production were analyzed in iSLK BAC16 cells, which have been widely used for recombinant KSHV production. We found that the production of KSHV virions was significantly increased by treatment with DMSO during the induction of lytic replication. Mechanistically, lytic genes of KSHV were enhanced by DMSO treatment, which was correlated with virion production. Additionally, DMSO induced the phosphorylation of JNK during lytic replication, and inhibition of JNK abolished the effects of DMSO on lytic replication and virion production. Our findings showed that additional treatment with DMSO during the induction of lytic replication significantly improved the yield of KSHV production.

Viral production is a key parameter for assessing virus-mediated biogeochemical cycles. One widely used method for the determination of viral production, called the virus reduction assay, reduces viral abundance, while maintaining bacterial abundance, using 0.2-μm pore-size filters. Viral production is estimated from the increase of viral abundance during incubation. We hypothesized that small-cell-sized bacterial communities can pass through 0.2-μm filters and drive viral production, representing a missing fraction of viral production that is missed by the virus reduction assay. Coastal seawater was filtered through 0.2-μm filters and diluted with virus-free seawater. Viral production in the

In freshwater environments, limited data exist on the impact of mortality forces (viruses and heterotrophic nanoflagellates) on bacterial growth efficiency (BGE, index of bacterial carbon metabolism) compared to resource availability. An investigation to determine the relative influence of viral lysis and flagellate predation (top-down forces) on BGE was conducted in a mesotrophic freshwater system (Lake Goule, France) with time and space. Viral abundance was significantly (p < 0.001) related to bacterial abundance by a power law function with an exponent less than 1, emphasizing that the increases in host population (bacteria) together with viruses were not proportionate. A lytic viral strategy was evident throughout the study period, with high lysis of the bacterial population (up to 60%) supported by viral production rates. Viral processes (lysis and production) that were influenced by bacterial production and heterotrophic nanoflagellate abundance had a positive impact on BGE. Estimates of BGE were variable (9.9–45.5%) due to uncoupling between two metabolic parameters—namely bacterial production and respiration. The existence of a synergistic relationship between viruses and flagellates with bacteria in Lake Goule highlighted the decisive impact of top-down agents in sustaining the bacterial carbon metabolism of non-infected population through the nature of vital resources released via mortality processes.

Viral processes in the coastal and offshore regions of the Eastern Arabian Sea (west coast of India) and their contribution of viral lysis to the organic carbon and nitrogen pool was examined. Water samples were collected during the southwest monsoon season at different depths (up to 1000 m) from four transects, with each transect consisting of two sampling points (S1–S8). Abundances of viruses and prokaryotes together with viral mediated prokaryotic mortality (up to 49.7%) were significantly (p < 0.001) higher in eutrophic coastal stations, whereas high percent lysogeny (up to 93%) was observed in the offshore regions. High viral-mediated carbon (Mean ± SD = 67.47 ± 2.0 μM C L−1 d−1) and nitrogen (Mean ± SD = 13.49 ± 14.0 μM N L−1 d−1) release was evident in the surface coastal waters compared to offshore regions. The percentage contributions of carbon and nitrogen released by viral lysis to the total dissolved organic carbon and nitrogen pool were estimated to be 7.4% and 3.9%, respectively, in the coastal surface waters. Our findings suggest that the contribution of viral lysis to DOM production through viral shunt could be crucial for the cycling of major biogeochemical elements and functioning of the studied tropical ecosystem.

In spite of the fact that the interactions between environmental parameters and prokaryotic and viral abundance have been explored in various aquatic environments, only a few independent estimates of viral production and decay in the estuarine region have been explored. In this study, data were analyzed for viral and prokaryotic abundance, viral production, and viral decay in a subtropical Danshui estuary in summer 2021. Prokaryotic abundance varied from 2.4 ± 0.6 × 105 to 12 ± 2.3 × 105 cells mL−1, and viral abundance ranged from 2.3 ± 0.9 × 105 to 6.9 ± 1.3 × 105 viruses mL−1 during the study period. Viral abundance was significantly correlated with prokaryotic abundance and chlorophyll a concentration. Furthermore, studies of changes in viral to prokaryotic abundance ratio (VPR) ranged from 0.42 ± 0.11 to 2.0 ± 0.25. Viral decay values were 2.1 ± 0.5 and 2.1 ± 0.3 × 104 virus mL−1h−1, and non-significant differences were observed between the inner estuary and coastal water region. Viral decay almost balanced gross viral production in this study. The dilution experiments revealed non-significant net viral production in July; thus, a lower VPR might be explained in this estuarine environment.

FAM46C is a multiple myeloma (MM) tumor suppressor whose function is only starting to be elucidated. We recently showed that in MM cells FAM46C triggers apoptosis by inhibiting autophagy and altering intracellular trafficking and protein secretion. To date, both a physiological characterization of FAM46C role and an assessment of FAM46C-induced phenotypes outside of MM are lacking. Preliminary reports suggested an involvement of FAM46C with regulation of viral replication, but this was never confirmed. Here, we show that FAM46C is an interferon-stimulated gene and that the expression of wild-type FAM46C in HEK-293T cells, but not of its most frequently found mutant variants, inhibits the production of both HIV-1-derived and HIV-1 lentiviruses. We demonstrate that this effect does not require transcriptional regulation and does not depend on inhibition of either global or virus-specific translation but rather mostly relies on FAM46C-induced deregulation of autophagy, a pathway that we show to be required for efficient lentiviral particle production. These studies not only provide new insights on the physiological role of the FAM46C protein but also could help in implementing more efficient antiviral strategies on one side and lentiviral particle production approaches on the other. IMPORTANCE FAM46C role has been thoroughly investigated in MM, but studies characterizing its role outside of the tumoral environment are still lacking. Despite the success of antiretroviral therapy in suppressing HIV load to undetectable levels, there is currently no HIV cure, and treatment is lifelong. Indeed, HIV continues to be a major global public health issue. Here, we show that FAM46C expression in HEK-293T cells inhibits the production of both HIV and HIV-derived lentiviruses. We also demonstrate that such inhibitory effect relies, at least in part, on the well-established regulatory role that FAM46C exerts on autophagy. Deciphering the molecular mechanism underlying this regulation will not only facilitate the understanding of FAM46C physiological role but also give new insights on the interplay between HIV and the cellular environment., Competing Interests: The authors declare no conflict of interest.

Numerous viruses manipulate host factors for viral production. We demonstrated that human enterovirus A71 (EVA71), a primary causative agent for hand, foot, and mouth disease (HFMD), increased the level of the DNA damage response (DDR) marker γ-H2AX. DDR is primarily mediated by the ataxia telangiectasia mutated (ATM), ATM and Rad3-related (ATR), or DNA-dependent protein kinase (DNA-PK) pathways. Upregulation of γ-H2AX by EVA71 was dependent on the ATR but not the ATM or DNA-PK pathway. As a nuclear factor, there is no previous evidence of cytoplasmic distribution of γ-H2AX. However, the present findings demonstrated that EVA71 encouraged the localization of γ-H2AX to the cytoplasm. Of note, γ-H2AX formed a complex with structural protein VP3, non-structural protein 3D, and the viral genome. Treatment with an inhibitor or CRISPR/Cas9 technology to decrease or silence the expression of γ-H2AX decreased viral genome replication in host cells; this effect was accompanied by decreased viral protein expression and virions. In animal experiments, caffeine was used to inhibit DDR; the results revealed that caffeine protected neonatal mice from death after infection with EVA71, laying the foundation for new therapeutic applications of caffeine. More importantly, in children with HFMD, γ-H2AX was upregulated in peripheral blood lymphocytes. The consistent in vitro and in vivo data on γ-H2AX from this study suggested that caffeine or other inhibitors of DDR might be novel therapeutic agents for HFMD.