Discovery of Novel Antiviral Agents Directed Against the Influenza A Virus Nucleoprotein

Aida, Yoko; Sasaki, Yutaka; Hagiwara, Kyoji

doi:10.5772/33573

Cited by 4 publications

(4 citation statements)

References 118 publications

(98 reference statements)

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“…Viral nucleoprotein (NP) plays multiple roles in viral lifecycle, including nuclear import/export of viral ribonucleoprotein (vRNP) and transcription/replication of the viral genome [ 8 , 9 , 10 ]. Targeting at the unconventional nuclear localization signal (NLS) of NP by mycalamide analogs results in a reduction of viral replication [ 11 ].…”

Section: Introductionmentioning

confidence: 99%

NXT1, a Novel Influenza A NP Binding Protein, Promotes the Nuclear Export of NP via a CRM1-Dependent Pathway

Chutiwitoonchai

Aida

2016

Viruses

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Influenza remains a serious worldwide public health problem. After infection, viral genomic RNA is replicated in the nucleus and packed into viral ribonucleoprotein, which will then be exported to the cytoplasm via a cellular chromosome region maintenance 1 (CRM1)-dependent pathway for further assembly and budding. However, the nuclear export mechanism of influenza virus remains controversial. Here, we identify cellular nuclear transport factor 2 (NTF2)-like export protein 1 (NXT1) as a novel binding partner of nucleoprotein (NP) that stimulates NP-mediated nuclear export via the CRM1-dependent pathway. NXT1-knockdown cells exhibit decreased viral replication kinetics and nuclear accumulated viral RNA and NP. By contrast, NXT1 overexpression promotes nuclear export of NP in a CRM1-dependent manner. Pull-down assays suggest the formation of an NXT1, NP, and CRM1 complex, and demonstrate that NXT1 binds to the C-terminal region of NP. These findings reveal a distinct mechanism for nuclear export of the influenza virus and identify the NXT1/NP interaction as a potential target for antiviral drug development.

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Section: Introductionmentioning

confidence: 99%

NXT1, a Novel Influenza A NP Binding Protein, Promotes the Nuclear Export of NP via a CRM1-Dependent Pathway

Chutiwitoonchai

Aida

2016

Viruses

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“…The multiple functions of NP mean that it has attracted interest as a novel target for antiviral drugs [ 21 ]. A recent study shows that viral replication is inhibited by small-molecule compounds that target NP, including nuclear localization signal (NLS)-binding compounds (i.e., mycalamide analogs) [ 22 ] and NP salt bridge inhibitors (i.e., compound 3) [ 23 ].…”

Section: Introductionmentioning

confidence: 99%

A Novel Antiviral Target Structure Involved in the RNA Binding, Dimerization, and Nuclear Export Functions of the Influenza A Virus Nucleoprotein

et al. 2015

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Developing antiviral therapies for influenza A virus (IAV) infection is an ongoing process because of the rapid rate of antigenic mutation and the emergence of drug-resistant viruses. The ideal strategy is to develop drugs that target well-conserved, functionally restricted, and unique surface structures without affecting host cell function. We recently identified the antiviral compound, RK424, by screening a library of 50,000 compounds using cell-based infection assays. RK424 showed potent antiviral activity against many different subtypes of IAV in vitro and partially protected mice from a lethal dose of A/WSN/1933 (H1N1) virus in vivo. Here, we show that RK424 inhibits viral ribonucleoprotein complex (vRNP) activity, causing the viral nucleoprotein (NP) to accumulate in the cell nucleus. In silico docking analysis revealed that RK424 bound to a small pocket in the viral NP. This pocket was surrounded by three functionally important domains: the RNA binding groove, the NP dimer interface, and nuclear export signal (NES) 3, indicating that it may be involved in the RNA binding, oligomerization, and nuclear export functions of NP. The accuracy of this binding model was confirmed in a NP-RK424 binding assay incorporating photo-cross-linked RK424 affinity beads and in a plaque assay evaluating the structure-activity relationship of RK424. Surface plasmon resonance (SPR) and pull-down assays showed that RK424 inhibited both the NP-RNA and NP-NP interactions, whereas size exclusion chromatography showed that RK424 disrupted viral RNA-induced NP oligomerization. In addition, in vitro nuclear export assays confirmed that RK424 inhibited nuclear export of NP. The amino acid residues comprising the NP pocket play a crucial role in viral replication and are highly conserved in more than 7,000 NP sequences from avian, human, and swine influenza viruses. Furthermore, we found that the NP pocket has a surface structure different from that of the pocket in host molecules. Taken together, these results describe a promising new approach to developing influenza virus drugs that target a novel pocket structure within NP.

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“…NP must also interact with cellular nucleolin, a major component of the nucleolar compartment, to facilitate the interaction between vRNP and the cellular nuclear export machinery (Terrier et al, 2016). In addition to its role in vRNP nuclear export, NP plays other important roles in the viral life cycle, including vRNP nuclear import and transcription/replication of the viral genome (Aida et al, 2012;Cianci et al, 2013;Portela and Digard, 2002;Sasaki et al, 2013). These functional studies support NP as a promising novel target for antiviral drug development.…”

Section: Introductionmentioning

confidence: 93%

Inhibition of CRM1-mediated nuclear export of influenza A nucleoprotein and nuclear export protein as a novel target for antiviral drug development

et al. 2017

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An anti-influenza compound, DP2392-E10 based on inhibition of the nuclear export function of the viral nucleoprotein-nuclear export signal 3 (NP-NES3) domain was successfully identified by our previous high-throughput screening system. Here, we demonstrated that DP2392-E10 exerts its antiviral effect by inhibiting replication of a broad range of influenza A subtypes. In regard to the molecular mechanism, we revealed that DP2392-E10 inhibits nuclear export of both viral NP and nuclear export protein (NEP). More specifically, in vitro pull-down assays revealed that DP2392-E10 directly binds cellular CRM1, which mediates nuclear export of NP and NEP. In silico docking suggested that DP2392-E10 binds at a region close to the HEAT9 and HEAT10 domains of CRM1. Together, these results indicate that the CRM1-mediated nuclear export function of influenza virus represents a new potential target for antiviral drug development, and also provide a core structure for a novel class of inhibitors that target this function.

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Discovery of Novel Antiviral Agents Directed Against the Influenza A Virus Nucleoprotein

Cited by 4 publications

References 118 publications

NXT1, a Novel Influenza A NP Binding Protein, Promotes the Nuclear Export of NP via a CRM1-Dependent Pathway

NXT1, a Novel Influenza A NP Binding Protein, Promotes the Nuclear Export of NP via a CRM1-Dependent Pathway

A Novel Antiviral Target Structure Involved in the RNA Binding, Dimerization, and Nuclear Export Functions of the Influenza A Virus Nucleoprotein

Inhibition of CRM1-mediated nuclear export of influenza A nucleoprotein and nuclear export protein as a novel target for antiviral drug development

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