Interferon-Induced HERC5 Inhibits Ebola Virus Particle Production and Is Antagonized by Ebola Glycoprotein.
Ontology highlight
ABSTRACT:
SUBMITTER: Paparisto E
PROVIDER: S-EPMC8472148 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:Ebola virus (EBOV), a member of the mononegaviral family Filoviridae, causes severe disease associated with high lethality in humans. Despite enormous progress in development of EBOV medical countermeasures, no anti-EBOV treatment has been approved. We designed an immunotoxin in which a single-chain variable region fragment of the EBOV glycoprotein-specific monoclonal antibody 6D8 was fused to the effector domains of Pseudomonas aeruginosa exotoxin A (PE38). This immunotoxin, 6D8-PE38, bound specifically to cells expressing EBOV glycoproteins. Importantly, 6D8-PE38 targeted EBOV-infected cells, as evidenced by inhibition of infectious EBOV production from infected cells, including primary human macrophages. The data presented here provide a proof of concept for immunotoxin-based targeted killing of infected cells as a potential antiviral intervention for Ebola virus disease.
| S-EPMC7790382 | biostudies-literature
Project description:Ebola virus outbreaks, such as the 2014 Makona epidemic in West Africa, are episodic and deadly. Filovirus antivirals are currently not clinically available. Our findings suggest interferon gamma, an FDA-approved drug, may serve as a novel and effective prophylactic or treatment option. Using mouse-adapted Ebola virus, we found that murine interferon gamma administered 24 hours before or after infection robustly protects lethally-challenged mice and reduces morbidity and serum viral titers. Furthermore, we demonstrated that interferon gamma profoundly inhibits Ebola virus infection of macrophages, an early cellular target of infection. As early as six hours following in vitro infection, Ebola virus RNA levels in interferon gamma-treated macrophages were lower than in infected, untreated cells. Addition of the protein synthesis inhibitor, cycloheximide, to interferon gamma-treated macrophages did not further reduce viral RNA levels, suggesting that interferon gamma blocks life cycle events that require protein synthesis such as virus replication. Microarray studies with interferon gamma-treated human macrophages identified more than 160 interferon-stimulated genes. Ectopic expression of a select group of these genes inhibited Ebola virus infection. These studies provide new potential avenues for antiviral targeting as these genes that have not previously appreciated to inhibit negative strand RNA viruses and specifically Ebola virus infection. As treatment of interferon gamma robustly protects mice from lethal Ebola virus infection, we propose that interferon gamma should be further evaluated for its efficacy as a prophylactic and/or therapeutic strategy against filoviruses. Use of this FDA-approved drug could rapidly be deployed during future outbreaks.
| S-EPMC4643030 | biostudies-literature
Project description:Type I interferon (IFN) inhibits virus replication by activating multiple antiviral mechanisms and pathways. It has long been recognized that type I IFNs can potently block HIV-1 replication in vitro; as such, HIV-1 has been used as a system to identify and characterize IFN-induced antiviral proteins responsible for this block. IFN-induced HERC5 contains an amino-terminal Regulator of Chromosome Condensation 1 (RCC1)-like domain and a carboxyl-terminal Homologous to the E6-AP Carboxyl Terminus (HECT) domain. HERC5 is the main cellular E3 ligase that conjugates the IFN-induced protein ISG15 to proteins. This E3 ligase activity was previously shown to inhibit the replication of evolutionarily diverse viruses, including HIV-1. The contribution of the RCC1-like domain to the antiviral activity of HERC5 was previously unknown.In this study, we showed that HERC5 inhibits HIV-1 particle production by a second distinct mechanism that targets the nuclear export of Rev/RRE-dependent RNA. Unexpectedly, the E3 ligase activity of HERC5 was not required for this inhibition. Instead, this activity required the amino-terminal RCC1-like domain of HERC5. Inhibition correlated with a reduction in intracellular RanGTP protein levels and/or the ability of RanGTP to interact with RanBP1. Inhibition also correlated with altered subcellular localization of HIV-1 Rev. In addition, we demonstrated that positive evolutionary selection is operating on HERC5. We identified a region in the RCC1-like domain that exhibits an exceptionally high probability of having evolved under positive selection and showed that this region is required for HERC5-mediated inhibition of nuclear export.We have identified a second distinct mechanism by which HERC5 inhibits HIV-1 replication and demonstrate that HERC5 is evolving under strong positive selection. Together, our findings contribute to a growing body of evidence suggesting that HERC5 is a novel host restriction factor.
| S-EPMC4021598 | biostudies-other
Project description:Type 1 interferon (IFN) inhibits the release of HIV-1 virus particles via poorly defined mechanisms. Here, we show that IFNalpha induces retention of viral particles on the surface of fibroblasts, T cells, or primary lymphocytes infected with HIV-1 lacking the Vpu protein. Retained particles are tethered to cell surfaces, can be endocytosed, appear fully assembled, exhibit mature morphology, and can be detached by protease. Strikingly, expression of the HIV-1 Vpu protein attenuates the ability of human cells to adhere to, and thereby retain, nascent HIV-1 particles upon IFNalpha treatment. Vpu also counteracts the IFNalpha-induced retention of virus-like particles assembled from the Ebola virus matrix protein. Furthermore, levels of IFNalpha that suppress replication of Vpu-defective HIV-1 have little effect on wild-type HIV-1. Thus, we propose that HIV-1 expresses Vpu to counteract an IFNalpha-induced, general host defense that inhibits dissemination of enveloped virions from the surface of infected cells.
| S-EPMC3793644 | biostudies-literature
Project description:Mammalian cells employ numerous innate cellular mechanisms to inhibit viral replication and spread. Tetherin, also known as Bst-2 or CD317, is a recently identified, IFN-induced, cellular response factor that blocks release of HIV-1 and other retroviruses from infected cells. The means by which tetherin retains retroviruses on the cell surface, as well as the mechanism used by the HIV-1 accessory protein Vpu to antagonize tetherin function and promote HIV-1 release, are unknown. Here, we document that tetherin functions as a broadly acting antiviral factor by demonstrating that both human and murine tetherin potently inhibit the release of the filovirus, Ebola, from the surface of cells. Expression of the Ebola glycoprotein (GP) antagonized the antiviral effect of human and murine tetherin and facilitated budding of Ebola particles, as did the HIV-1 Vpu protein. Conversely, Ebola GP could substitute for Vpu to promote HIV-1 virion release from tetherin-expressing cells, demonstrating a common cellular target for these divergent viral proteins. Ebola GP efficiently coimmunoprecipitated with tetherin, suggesting that the viral glycoprotein directly interferes with this host antiviral factor. These results demonstrate that tetherin is a cellular antiviral factor that restricts budding of structurally diverse enveloped viruses. Additionally, Ebola has evolved a highly effective strategy to combat this antiviral response elicited in the host during infection.
| S-EPMC2650360 | biostudies-literature
Project description:Membrane-associated RING-CH-type 8 (MARCH8) strongly blocks human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) incorporation into virions by downregulating its cell surface expression, but the mechanism is still unclear. We now report that MARCH8 also blocks the Ebola virus (EBOV) glycoprotein (GP) incorporation via surface downregulation. To understand how these viral fusion proteins are downregulated, we investigated the effects of MARCH8 on EBOV GP maturation and externalization via the conventional secretion pathway. MARCH8 interacted with EBOV GP and furin when detected by immunoprecipitation and retained the GP/furin complex in the Golgi when their location was tracked by a bimolecular fluorescence complementation (BiFC) assay. MARCH8 did not reduce the GP expression or affect the GP modification by high-mannose N-glycans in the endoplasmic reticulum (ER), but it inhibited the formation of complex N-glycans on the GP in the Golgi. Additionally, the GP O-glycosylation and furin-mediated proteolytic cleavage were also inhibited. Moreover, we identified a novel furin cleavage site on EBOV GP and found that only those fully glycosylated GPs were processed by furin and incorporated into virions. Furthermore, the GP shedding and secretion were all blocked by MARCH8. MARCH8 also blocked the furin-mediated cleavage of HIV-1 Env (gp160) and the highly pathogenic avian influenza virus H5N1 hemagglutinin (HA). We conclude that MARCH8 has a very broad antiviral activity by prohibiting different viral fusion proteins from glycosylation and proteolytic cleavage in the Golgi, which inhibits their transport from the Golgi to the plasma membrane and incorporation into virions.IMPORTANCE Enveloped viruses express three classes of fusion proteins that are required for their entry into host cells via mediating virus and cell membrane fusion. Class I fusion proteins are produced from influenza viruses, retroviruses, Ebola viruses, and coronaviruses. They are first synthesized as a type I transmembrane polypeptide precursor that is subsequently glycosylated and oligomerized. Most of these precursors are cleaved en route to the plasma membrane by a cellular protease furin in the late secretory pathway, generating the trimeric N-terminal receptor-binding and C-terminal fusion subunits. Here, we show that a cellular protein, MARCH8, specifically inhibits the furin-mediated cleavage of EBOV GP, HIV-1 Env, and H5N1 HA. Further analyses uncovered that MARCH8 blocked the EBOV GP glycosylation in the Golgi and inhibited its transport from the Golgi to the plasma membrane. Thus, MARCH8 has a very broad antiviral activity by specifically inactivating different viral fusion proteins.
| S-EPMC7492737 | biostudies-literature
Project description:Treatment of human cells with Type 1 interferons restricts HIV replication. Here we report that the tripartite motif protein TRIM22 is a key mediator. We used transcriptional profiling to identify cellular genes that were induced by interferon treatment and identified TRIM22 as one of the most strongly up-regulated genes. We confirmed, as in previous studies, that TRIM22 over-expression inhibited HIV replication. To assess the role of TRIM22 expressed under natural inducing conditions, we compared the effects of interferon in cells depleted for TRIM22 using RNAi and found that HIV particle release was significantly increased in the knockdown, implying that TRIM22 acts as a natural antiviral effector. Further studies showed that TRIM22 inhibited budding of virus-like particles containing Gag only, indicating that Gag was the target of TRIM22. TRIM22 did not block the release of MLV or EIAV Gag particles. Inhibition was associated with diffuse cytoplasmic staining of HIV Gag rather than accumulation at the plasma membrane, suggesting TRIM22 disrupts proper trafficking. Mutational analyses of TRIM22 showed that the catalytic amino acids Cys15 and Cys18 of the RING domain are required for TRIM22 antiviral activity. These data disclose a pathway by which Type 1 interferons obstruct HIV replication.
| S-EPMC2279259 | biostudies-literature
Project description:UnlabelledThe Ebola virus (EBOV) surface glycoprotein (GP1,2) mediates host cell attachment and fusion and is the primary target for host neutralizing antibodies. Expression of GP1,2 at high levels disrupts normal cell physiology, and EBOV uses an RNA-editing mechanism to regulate expression of the GP gene. In this study, we demonstrate that high levels of GP1,2 expression impair production and release of EBOV virus-like particles (VLPs) as well as infectivity of GP1,2-pseudotyped viruses. We further show that this effect is mediated through two mechanisms. First, high levels of GP1,2 expression reduce synthesis of other proteins needed for virus assembly. Second, viruses containing high levels of GP1,2 are intrinsically less infectious, possibly due to impaired receptor binding or endosomal processing. Importantly, proteolysis can rescue the infectivity of high-GP1,2-containing viruses. Taken together, our findings indicate that GP1,2 expression levels have a profound effect on factors that contribute to virus fitness and that RNA editing may be an important mechanism employed by EBOV to regulate GP1,2 expression in order to optimize virus production and infectivity.ImportanceThe Ebola virus (EBOV), as well as other members of the Filoviridae family, causes severe hemorrhagic fever that is highly lethal, with up to 90% mortality. The EBOV surface glycoprotein (GP1,2) plays important roles in virus infection and pathogenesis, and its expression is tightly regulated by an RNA-editing mechanism during virus replication. Our study demonstrates that the level of GP1,2 expression profoundly affects virus particle production and release and uncovers a new mechanism by which Ebola virus infectivity is regulated by the level of GP1,2 expression. These findings extend our understanding of EBOV infection and replication in adaptation of host environments, which will aid the development of countermeasures against EBOV infection.
| S-EPMC4300637 | biostudies-literature
Project description:Ebola virus (EBOV) protein VP24 carries out at least two critical functions. It promotes condensation of viral nucleocapsids, which is crucial for infectious virus production, and it suppresses interferon (IFN) signaling, which requires interaction with the NPI-1 subfamily of importin-α (IMPA) nuclear transport proteins. Interestingly, over-expressed IMPA leads to VP24 nuclear accumulation and a carboxy-terminus nuclear export signal (NES) has been reported, suggesting that VP24 may undergo nuclear trafficking. For the first time, we demonstrate that NPI-1 IMPA overexpression leads to the nuclear accumulation of VP24 during EBOV infection. To assess the functional impact of nuclear trafficking, we generated tetracistronic minigenomes encoding VP24 nuclear import and/or export signal mutants. The minigenomes, which also encode Renilla luciferase and viral proteins VP40 and GP, were used to generate transcription and replication competent virus-like particles (trVLPs) that can be used to assess EBOV RNA synthesis, gene expression, entry and viral particle production. With this system, we confirmed that NES or IMPA binding site mutations altered VP24 nuclear localization, demonstrating functional trafficking signals. While these mutations minimally affected transcription and replication, the trVLPs exhibited impaired infectivity and formation of shortened nucleocapsids for the IMPA binding mutant. For the NES mutants, infectivity was reduced approximately 1000-fold. The NES mutant could still suppress IFN signaling but failed to promote nucleocapsid formation. To determine whether VP24 nuclear export is required for infectivity, the residues surrounding the wildtype NES were mutated to alanine or the VP24 NES was replaced with the Protein Kinase A Inhibitor NES. While nuclear export remained intact for these mutants, infectivity was severely impaired. These data demonstrate that VP24 undergoes nuclear trafficking and illuminates a separate and critical role for the NES and surrounding sequences in infectivity and nucleocapsid assembly.
| S-EPMC10980025 | biostudies-literature
Project description: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.
| S-EPMC10434054 | biostudies-literature