Nipah Virus V Protein Binding Alters MDA5 Helicase Folding Dynamics

ABSTRACT Paramyxovirus V proteins are known antagonists of the RIG-I-like receptor (RLR)-mediated interferon induction pathway, interacting with and inhibiting the RLR MDA5. We report interactions between the Nipah virus V protein and both RIG-I regulatory protein TRIM25 and RIG-I. We also observed interactions between these host proteins and the V proteins of measles virus, Sendai virus, and parainfluenza virus. These interactions are mediated by the conserved C-terminal domain of the V protein, which binds to the tandem caspase activation and recruitment domains (CARDs) of RIG-I (the region of TRIM25 ubiquitination) and to the SPRY domain of TRIM25, which mediates TRIM25 interaction with the RIG-I CARDs. Furthermore, we show that V interaction with TRIM25 and RIG-I prevents TRIM25-mediated ubiquitination of RIG-I and disrupts downstream RIG-I signaling to the mitochondrial antiviral signaling protein. This is a novel mechanism for innate immune inhibition by paramyxovirus V proteins, distinct from other known V protein functions such as MDA5 and STAT1 antagonism. IMPORTANCE The host RIG-I signaling pathway is a key early obstacle to paramyxovirus infection, as it results in rapid induction of an antiviral response. This study shows that paramyxovirus V proteins interact with and inhibit the activation of RIG-I, thereby interrupting the antiviral signaling pathway and facilitating virus replication.

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Sendai Virus V Protein Inhibits the Secretion of Interleukin-1β by Preventing NLRP3 Inflammasome Assembly

Journal of Virology ◽

10.1128/jvi.00842-18 ◽

2018 ◽

Vol 92 (19) ◽

Cited By ~ 7

Author(s):

Takayuki Komatsu ◽

Yukie Tanaka ◽

Yoshinori Kitagawa ◽

Naoki Koide ◽

Yoshikazu Naiki ◽

...

Keyword(s):

Nlrp3 Inflammasome ◽

Sendai Virus ◽

Nipah Virus ◽

Innate Immune ◽

Parainfluenza Virus ◽

Inflammasome Activation ◽

V Protein ◽

Nlrp3 Inflammasome Activation ◽

Human Parainfluenza Virus

ABSTRACT Inflammasomes play a key role in host innate immune responses to viral infection by caspase-1 (Casp-1) activation to facilitate interleukin-1β (IL-1β) secretion, which contributes to the host antiviral defense. The NLRP3 inflammasome consists of the cytoplasmic sensor molecule NLRP3, adaptor protein ASC, and effector protein pro-caspase-1 (pro-Casp-1). NLRP3 and ASC promote pro-Casp-1 cleavage, leading to IL-1β maturation and secretion. However, as a countermeasure, viral pathogens have evolved virulence factors to antagonize inflammasome pathways. Here we report that V gene knockout Sendai virus [SeV V(−)] induced markedly greater amounts of IL-1β than wild-type SeV in infected THP1 macrophages. Deficiency of NLRP3 in cells inhibited SeV V(−)-induced IL-1β secretion, indicating an essential role for NLRP3 in SeV V(−)-induced IL-1β activation. Moreover, SeV V protein inhibited the assembly of NLRP3 inflammasomes, including NLRP3-dependent ASC oligomerization, NLRP3-ASC association, NLRP3 self-oligomerization, and intermolecular interactions between NLRP3 molecules. Furthermore, a high correlation between the NLRP3-binding capacity of V protein and the ability to block inflammasome complex assembly was observed. Therefore, SeV V protein likely inhibits NLRP3 self-oligomerization by interacting with NLRP3 and inhibiting subsequent recruitment of ASC to block NLRP3-dependent ASC oligomerization, in turn blocking full activation of the NLRP3 inflammasome and thus blocking IL-1β secretion. Notably, the inhibitory action of SeV V protein on NLRP3 inflammasome activation is shared by other paramyxovirus V proteins, such as Nipah virus and human parainfluenza virus type 2. We thus reveal a mechanism by which paramyxovirus inhibits inflammatory responses by inhibiting NLRP3 inflammasome complex assembly and IL-1β activation. IMPORTANCE The present study demonstrates that the V protein of SeV, Nipah virus, and human parainfluenza virus type 2 interacts with NLRP3 to inhibit NLRP3 inflammasome activation, potentially suggesting a novel strategy by which viruses evade the host innate immune response. As all members of the Paramyxovirinae subfamily carry similar V genes, this new finding may also lead to identification of novel therapeutic targets for paramyxovirus infection and related diseases.

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Identification of the Nuclear Export Signal and STAT-Binding Domains of the Nipah Virus V Protein Reveals Mechanisms Underlying Interferon Evasion

Journal of Virology ◽

10.1128/jvi.78.10.5358-5367.2004 ◽

2004 ◽

Vol 78 (10) ◽

pp. 5358-5367 ◽

Cited By ~ 92

Author(s):

Jason J. Rodriguez ◽

Cristian D. Cruz ◽

Curt M. Horvath

Keyword(s):

Amino Acids ◽

Protein Interactions ◽

Nuclear Export ◽

Nipah Virus ◽

Nuclear Export Signal ◽

Hendra Virus ◽

Nuclear Accumulation ◽

V Protein ◽

Binding Domains ◽

Export Signal

ABSTRACT The V proteins of Nipah virus and Hendra virus have been demonstrated to bind to cellular STAT1 and STAT2 proteins to form high-molecular-weight complexes that inhibit interferon (IFN)-induced antiviral transcription by preventing STAT nuclear accumulation. Analysis of the Nipah virus V protein has revealed a region between amino acids 174 and 192 that functions as a CRM1-dependent nuclear export signal (NES). This peptide is sufficient to complement an export-defective human immunodeficiency virus Rev protein, and deletion and substitution mutagenesis revealed that this peptide is necessary for both V protein shuttling and cytoplasmic retention of STAT1 and STAT2 proteins. However, the NES is not required for V-dependent IFN signaling inhibition. IFN signaling is blocked primarily by interaction between Nipah virus V residues 100 to 160 and STAT1 residues 509 to 712. Interaction with STAT2 requires a larger Nipah virus V segment between amino acids 100 and 300, but deletion of residues 230 to 237 greatly reduced STAT2 coprecipitation. Further, V protein interactions with cellular STAT1 is a prerequisite for STAT2 binding, and sequential immunoprecipitations demonstrate that V, STAT1, and STAT2 can form a tripartite complex. These findings characterize essential regions for Henipavirus V proteins that represent potential targets for therapeutic intervention.

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Possible role of the Nipah virus V protein in the regulation of the interferon beta induction by interacting with UBX domain-containing protein1

Scientific Reports ◽

10.1038/s41598-018-25815-9 ◽

2018 ◽

Vol 8 (1) ◽

Cited By ~ 4

Author(s):

Shotaro Uchida ◽

Ryo Horie ◽

Hiroki Sato ◽

Chieko Kai ◽

Misako Yoneda

Keyword(s):

Interferon Beta ◽

Nipah Virus ◽

V Protein

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A single amino acid substitution in the V protein of Nipah virus alters its ability to block interferon signalling in cells from different species

Journal of General Virology ◽

10.1099/vir.0.82261-0 ◽

2006 ◽

Vol 87 (12) ◽

pp. 3649-3653 ◽

Cited By ~ 28

Author(s):

Kathrin Hagmaier ◽

Nicola Stock ◽

Steve Goodbourn ◽

Lin-Fa Wang ◽

Richard Randall

Keyword(s):

Amino Acid ◽

Amino Acid Substitution ◽

Molecular Basis ◽

Nipah Virus ◽

Human Cells ◽

Single Amino Acid Substitution ◽

Single Amino Acid ◽

V Protein ◽

Major Constraint ◽

Interferon Signalling

The V protein of the paramyxovirus Nipah virus (NiV) has been shown to antagonize the interferon (IFN) response in human cells via sequestration of STAT1 and STAT2. This study describes a mutant of the NiV V protein, referred to as V(AAHL), that is unable to antagonize IFN signalling and demonstrates that a single amino acid substitution is responsible for its inactivity. The molecular basis for this was identified as a failure to interact with STAT1 and STAT2. It was also shown that NiV V, but not V(AAHL), was functional as an IFN antagonist in human, monkey, rabbit, dog, horse, pig and bat cells, which suggests that the ability of NiV to block IFN signalling is not a major constraint that prevents this virus from crossing species barriers.

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Nuclear Localization of the Nipah Virus W Protein Allows for Inhibition of both Virus- and Toll-Like Receptor 3-Triggered Signaling Pathways

Journal of Virology ◽

10.1128/jvi.79.10.6078-6088.2005 ◽

2005 ◽

Vol 79 (10) ◽

pp. 6078-6088 ◽

Cited By ~ 119

Author(s):

Megan L. Shaw ◽

Washington B. Cardenas ◽

Dmitriy Zamarin ◽

Peter Palese ◽

Christopher F. Basler

Keyword(s):

Nuclear Localization ◽

Nipah Virus ◽

Toll Like Receptor ◽

Promoter Activation ◽

Double Stranded Rna ◽

V Protein ◽

Terminal Domain ◽

P Gene ◽

Localization Signal ◽

Stimulation Of

ABSTRACT The Nipah virus V and W proteins, which are encoded by the P gene via RNA editing, have a common N-terminal domain but unique C-terminal domains. They localize to the cytoplasm and nucleus, respectively, and have both been shown to function as inhibitors of JAK/STAT signaling. Here we report that V and W proteins also block virus activation of the beta interferon (IFN-β) promoter and the IFN regulatory factor 3 (IRF3)-responsive IFN-stimulated gene 54 promoter. Surprisingly, only W protein shows strong inhibition of promoter activation in response to stimulation of Toll-like receptor 3 (TLR3) by extracellular double-stranded RNA. This activity is dependent on the nuclear localization of W protein. Within the unique C-terminal domain of W protein, we have identified a nuclear localization signal (NLS) that requires basic residues at positions 439, 440, and 442. This NLS is responsible for mediating the preferential interaction of W protein with karyopherin-α 3 and karyopherin-α 4. Nuclear localization of W protein therefore enables it to target both virus and TLR3 pathways, whereas the cytoplasmic V protein is restricted to inhibiting the virus pathway. We propose that this discrepancy is in part due to the V protein being less able to block signaling in response to the kinase, TBK-1, whereas both V and W can prevent promoter activation in response to IKKε. We demonstrate that, when the TLR3 pathway is stimulated, the levels of phosphorylated IRF3 are reduced in the presence of W protein but not V protein, confirming the differential effects of these proteins and illustrating that W protein-mediated inhibition is due to a loss of active IRF3.

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Newcastle Disease Virus (NDV)-Based Assay Demonstrates Interferon-Antagonist Activity for the NDV V Protein and the Nipah Virus V, W, and C Proteins

Journal of Virology ◽

10.1128/jvi.77.2.1501-1511.2003 ◽

2003 ◽

Vol 77 (2) ◽

pp. 1501-1511 ◽

Cited By ~ 272

Author(s):

Man-Seong Park ◽

Megan L. Shaw ◽

Jorge Muñoz-Jordan ◽

Jerome F. Cros ◽

Takaaki Nakaya ◽

...

Keyword(s):

Newcastle Disease Virus ◽

Disease Virus ◽

Newcastle Disease ◽

Nipah Virus ◽

Terminal Region ◽

Ns1 Protein ◽

Antagonist Activity ◽

V Protein ◽

Amino Terminal ◽

C Proteins

ABSTRACT We have generated a recombinant Newcastle disease virus (NDV) that expresses the green fluorescence protein (GFP) in infected chicken embryo fibroblasts (CEFs). This virus is interferon (IFN) sensitive, and pretreatment of cells with chicken alpha/beta IFN (IFN-α/β) completely blocks viral GFP expression. Prior transfection of plasmid DNA induces an IFN response in CEFs and blocks NDV-GFP replication. However, transfection of known inhibitors of the IFN-α/β system, including the influenza A virus NS1 protein and the Ebola virus VP35 protein, restores NDV-GFP replication. We therefore conclude that the NDV-GFP virus could be used to screen proteins expressed from plasmids for the ability to counteract the host cell IFN response. Using this system, we show that expression of the NDV V protein or the Nipah virus V, W, or C proteins rescues NDV-GFP replication in the face of the transfection-induced IFN response. The V and W proteins of Nipah virus, a highly lethal pathogen in humans, also block activation of an IFN-inducible promoter in primate cells. Interestingly, the amino-terminal region of the Nipah virus V protein, which is identical to the amino terminus of Nipah virus W, is sufficient to exert the IFN-antagonist activity. In contrast, the anti-IFN activity of the NDV V protein appears to be located in the carboxy-terminal region of the protein, a region implicated in the IFN-antagonist activity exhibited by the V proteins of mumps virus and human parainfluenza virus type 2.

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Studies on the metabolic fate of azuletil sodium. V: Protein binding and uptake by erythrocytes.

Drug Metabolism and Pharmacokinetics ◽

10.2133/dmpk.5.375 ◽

1990 ◽

Vol 5 (3) ◽

pp. 375-382

Author(s):

Masao SATO ◽

Hiroaki SUZAKA ◽

Akira TOMIYAMA ◽

Hiroshi MIYAZAKI

Keyword(s):

Protein Binding ◽

Metabolic Fate ◽

V Protein

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Ensemble description of the intrinsically disordered N-terminal domain of the Nipah virus P/V protein from combined NMR and SAXS

Scientific Reports ◽

10.1038/s41598-020-76522-3 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Marco Schiavina ◽

Edoardo Salladini ◽

Maria Grazia Murrali ◽

Giancarlo Tria ◽

Isabella C. Felli ◽

...

Keyword(s):

Nmr Spectroscopy ◽

Chemical Shifts ◽

Nipah Virus ◽

Conformational Ensemble ◽

Genome Replication ◽

Nmr Studies ◽

V Protein ◽

Intrinsically Disordered ◽

Terminal Domain ◽

Starting Point

Abstract Using SAXS and NMR spectroscopy, we herein provide a high-resolution description of the intrinsically disordered N-terminal domain (PNT, aa 1–406) shared by the Nipah virus (NiV) phosphoprotein (P) and V protein, two key players in viral genome replication and in evasion of the host innate immune response, respectively. The use of multidimensional NMR spectroscopy allowed us to assign as much as 91% of the residues of this intrinsically disordered domain whose size constitutes a technical challenge for NMR studies. Chemical shifts and nuclear relaxation measurements provide the picture of a highly flexible protein. The combination of SAXS and NMR information enabled the description of the conformational ensemble of the protein in solution. The present results, beyond providing an overall description of the conformational behavior of this intrinsically disordered region, also constitute an asset for obtaining atomistic information in future interaction studies with viral and/or cellular partners. The present study can thus be regarded as the starting point towards the design of inhibitors that by targeting crucial protein–protein interactions involving PNT might be instrumental to combat this deadly virus.

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Henipavirus V Protein Association with Polo-Like Kinase Reveals Functional Overlap with STAT1 Binding and Interferon Evasion

Journal of Virology ◽

10.1128/jvi.00409-08 ◽

2008 ◽

Vol 82 (13) ◽

pp. 6259-6271 ◽

Cited By ~ 21

Author(s):

Louise E. Ludlow ◽

Michael K. Lo ◽

Jason J. Rodriguez ◽

Paul A. Rota ◽

Curt M. Horvath

Keyword(s):

Protein Interactions ◽

Rna Synthesis ◽

Nipah Virus ◽

Hendra Virus ◽

Host Protein ◽

Binding Motif ◽

Emerging Viruses ◽

V Protein ◽

P Protein ◽

Synthetic Capacity

ABSTRACT Emerging viruses in the paramyxovirus genus Henipavirus evade host antiviral responses via protein interactions between the viral V and W proteins and cellular STAT1 and STAT2 and the cytosolic RNA sensor MDA5. Polo-like kinase (PLK1) is identified as being an additional cellular partner that can bind to Nipah virus P, V, and W proteins. For both Nipah virus and Hendra virus, contact between the V protein and the PLK1 polo box domain is required for V protein phosphorylation. Results indicate that PLK1 is engaged by Nipah virus V protein amino acids 100 to 160, previously identified as being the STAT1 binding domain responsible for host interferon (IFN) signaling evasion, via a Thr-Ser-Ser-Pro motif surrounding residue 130. A distinct Ser-Thr-Pro motif surrounding residue 199 mediates the PLK1 interaction with Hendra virus V protein. Select mutations in the motif surrounding residue 130 also influenced STAT1 binding and innate immune interference, and data indicate that the V:PLK1 and V:STAT complexes are V mediated yet independent of one another. The effects of STAT1/PLK1 binding motif mutations on the function the Nipah virus P protein in directing RNA synthesis were tested. Remarkably, mutations that selectively disrupt the STAT or PLK1 interaction site have no effects on Nipah virus P protein-mediated viral RNA synthesis. Therefore, mutations targeting V protein-mediated IFN evasion will not alter the RNA synthetic capacity of the virus, supporting an attenuation strategy based on disrupting host protein interactions.

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