Ebola virus-like particle-induced activation of NF-κB and Erk signaling in human dendritic cells requires the glycoprotein mucin domain

Martinez, Osvaldo; Valmas, Charalampos; Basler, Christopher F.

doi:10.1016/j.virol.2007.03.020

Cited by 63 publications

(74 citation statements)

References 59 publications

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“…In this study, we demonstrated that IFN treatment and/or expression of human ISG15 significantly inhibited release of Ebola VP40-WT VLPs. It is well documented that Ebola virus VP40 can bud indepen- dently as a VLP from mammalian cells (40,46,47,61,65). Efficient release of VP40 VLPs is known to depend on functional L-domain(s) of VP40, and at least two host proteins, tsg101 and Nedd4 (36,48,51,61,62,66).…”

Section: Discussionmentioning

confidence: 99%

ISG15 inhibits Ebola VP40 VLP budding in an L-domain-dependent manner by blocking Nedd4 ligase activity

Okumura

Pitha

Harty

2008

Proc. Natl. Acad. Sci. U.S.A.

252

253

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Ebola virus budding is mediated by the VP40 matrix protein. VP40 can bud from mammalian cells independent of other viral proteins, and efficient release of VP40 virus-like particles (VLPs) requires interactions with host proteins such as tsg101 and Nedd4, an E3 ubiquitin ligase. Ubiquitin itself is thought to be exploited by Ebola virus to facilitate efficient virus egress. Disruption of VP40 function and thus virus budding remains an attractive target for the development of novel antiviral therapies. Here, we investigate the effect of ISG15 protein on the release of Ebola VP40 VLPs. ISG15 is an IFN-inducible, ubiquitinlike protein expressed after bacterial or viral infection. Our results show that expression of free ISG15, or the ISGylation system (UbE1L and UbcH8), inhibits budding of Ebola virus VP40 VLPs. Addressing the molecular mechanism of this inhibition, we show that ISG15 interacts with Nedd4 ubiquitin ligase and inhibits ubiquitination of VP40. Furthermore, the L-domain deletion mutant of VP40 (⌬PT/PY), which does not interact with Nedd4, was insensitive to ISG15-mediated inhibition of VLP release. These data provide evidence of antiviral activity of ISG15 against Ebola virus and suggest a mechanism of action involving disruption of Nedd4 function and subsequent ubiquitination of VP40.Ebola virus ͉ interferon ͉ innate immunity ͉ ubiquitin T he IFN pathway activates hundreds of cellular IFNstimulated genes (ISGs), some of which have direct antiviral activity (1-5). For example, ISG15 was one of the first recognized ISG proteins whose expression was up-regulated not only by IFN but also by viral infection, LPS treatment, and retinoic acid (6-8). ISG15 has high homology to ubiquitin and can be detected in cells in both free and conjugated forms (9, 10). ISG15 conjugation (ISGylation) to proteins uses cascades of enzymatic reactions similar to those used in protein ubiquitination pathways (11). Some of these enzymes, like ubiquitin E1-like protein (UBE1L) (12, 13), are unique for ISG15, whereas two E2 enzymes, UbcH8 and UbcH6, are also used in the ubiquitination pathway (14)(15)(16) The observation that ISG15 conjugation targets many components of the antiviral signaling pathway suggests that ISG15 may play a role in the innate antiviral response (12,17). To this effect, it was shown that the NS1 protein of the Influenza B virus inhibits ISGylation (12), and ISG15 expression decreased Sindbis virus replication and provided protection against lethal infection (12, 18). Also, ISG15-null mice showed an increase susceptibility to both DNA-and RNA-containing viruses in vivo, including influenza, herpes simplex, and Sindbis viruses (19). Recently, inhibition of HIV-1 virion release in IFN-treated cells was shown to be mediated by ISG15 (20). Several reports have also linked induction and expression of ISG15 to inhibition of important viral pathogens of fish (21,22). Together, these studies suggest that ISG15 has a potent and broad-based antiviral effect; however, the mechanism of action of ISG15 remains to be d...

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Section: Discussionmentioning

confidence: 99%

ISG15 inhibits Ebola VP40 VLP budding in an L-domain-dependent manner by blocking Nedd4 ligase activity

Okumura

Pitha

Harty

2008

Proc. Natl. Acad. Sci. U.S.A.

252

253

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“…The identification of viral antigen in the brains of NiV encephalitis patients postmortem indicates the spread of the virus past the presumed respiratory route of entry (13,27). DCs have also been known to provide privileged havens for viruses and have been indicated in the systemic spread of Ebola virus and Dengue virus (9,21,33,38,44). Additionally, relapse and late-onset encephalitis from Nipah virus infections suggest viral persistence associated with longterm immune evasion (58).…”

Section: Discussionmentioning

confidence: 99%

Novel Nipah Virus Immune-Antagonism Strategy Revealed by Experimental and Computational Study

Seto

Qiao

Guenzel

et al. 2010

J Virol

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Nipah virus is an emerging pathogen that causes severe disease in humans. It expresses several antagonist proteins that subvert the immune response and that may contribute to its pathogenicity. Studies of its biology are difficult due to its high pathogenicity and requirement for biosafety level 4 containment. We integrated experimental and computational methods to elucidate the effects of Nipah virus immune antagonists. Individual Nipah virus immune antagonists (phosphoprotein and V and W proteins) were expressed from recombinant Newcastle disease viruses, and the responses of infected human monocyte-derived dendritic cells were determined. We developed an ordinary differential equation model of the infectious process that that produced results with a high degree of correlation with these experimental results. In order to simulate the effects of wild-type virus, the model was extended to incorporate published experimental data on the time trajectories of immune-antagonist production. These data showed that the RNA-editing mechanism utilized by the wild-type Nipah virus to produce immune antagonists leads to a delay in the production of the most effective immune antagonists, V and W. Model simulations indicated that this delay caused a disconnection between attenuation of the antiviral response and suppression of inflammation. While the antiviral cytokines were efficiently suppressed at early time points, some early inflammatory cytokine production occurred, which would be expected to increase vascular permeability and promote virus spread and pathogenesis. These results suggest that Nipah virus has evolved a unique immune-antagonist strategy that benefits from controlled expression of multiple antagonist proteins with various potencies.

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“…Monoclonal antibodies against Zaire EBOV VP35 and against Zaire EBOV NP were generated in collaboration with the Mount Sinai Hybridoma Center and have been described previously (6,27). A monoclonal antihemagglutinin (anti-HA) antibody was purchased from Sigma (St. Louis, MO).…”

Section: Antibodiesmentioning

confidence: 99%

“…Sequences encoding Zaire EBOV L amino acids 1 to 505 (HA-L amino acids 1 to 505) were amplified from plasmid pTM1-L by PCR (33), cloned with an amino-terminal HA tag, and cloned into pCAGGS. The expression plasmid for Zaire EBOV NP, pcDNA3 EBOV NP, has been described previously (27).…”

Section: Antibodiesmentioning

confidence: 99%

Basic Residues within the Ebolavirus VP35 Protein Are Required for Its Viral Polymerase Cofactor Function

Prins

Binning

Shabman

et al. 2010

J Virol

Self Cite

139

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The ebolavirus (EBOV) VP35 protein binds to double-stranded RNA (dsRNA), inhibits host alpha/beta interferon (IFN-␣/␤) production, and is an essential component of the viral polymerase complex. Structural studies of the VP35 C-terminal IFN inhibitory domain (IID) identified specific structural features, including a central basic patch and a hydrophobic pocket, that are important for dsRNA binding and IFN inhibition. Several other conserved basic residues bordering the central basic patch and a separate cluster of basic residues, called the first basic patch, were also identified. Functional analysis of alanine substitution mutants indicates that basic residues outside the central basic patch are not required for dsRNA binding or for IFN inhibition. However, minigenome assays, which assess viral RNA polymerase complex function, identified these other basic residues to be critical for viral RNA synthesis. Of these, a subset located within the first basic patch is important for VP35-nucleoprotein (NP) interaction, as evidenced by the inability of alanine substitution mutants to coimmunoprecipitate with NP. Therefore, first basic patch residues are likely critical for replication complex formation through interactions with NP. Coimmunoprecipitation studies further demonstrate that the VP35 IID is sufficient to interact with NP and that dsRNA can modulate VP35 IID interactions with NP. Other basic residue mutations that disrupt the VP35 polymerase cofactor function do not affect interaction with NP or with the amino terminus of the viral polymerase. Collectively, these results highlight the importance of conserved basic residues from the EBOV VP35 C-terminal IID and validate the VP35 IID as a potential therapeutic target.

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Ebola virus-like particle-induced activation of NF-κB and Erk signaling in human dendritic cells requires the glycoprotein mucin domain

Cited by 63 publications

References 59 publications

ISG15 inhibits Ebola VP40 VLP budding in an L-domain-dependent manner by blocking Nedd4 ligase activity

ISG15 inhibits Ebola VP40 VLP budding in an L-domain-dependent manner by blocking Nedd4 ligase activity

Novel Nipah Virus Immune-Antagonism Strategy Revealed by Experimental and Computational Study

Basic Residues within the Ebolavirus VP35 Protein Are Required for Its Viral Polymerase Cofactor Function

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