Interaction between NS1 and Cellular MAVS Contributes to NS1 Mitochondria Targeting.

Influenza A virus nonstructural protein 1 (NS1) plays an important role in evading host innate immunity. NS1 inhibits interferon (IFN) responses via multiple mechanisms, including sequestering dsRNA and suppressing retinoic acid-inducible gene I (RIG-I) signaling by interacting with RIG-I and tripartite motif-containing protein 25 (TRIM25). In the current study, we demonstrated the mitochondrial localization of NS1 at the early stage of influenza virus infection. Since NS1 does not contain mitochondria-targeting signals, we suspected that there is an association between the NS1 and mitochondrial proteins. This hypothesis was tested by demonstrating the interaction of NS1 with mitochondrial antiviral-signaling protein (MAVS) in a RIG-I-independent manner. Importantly, the association with MAVS facilitated the mitochondrial localization of NS1 and thereby significantly impeded MAVS-mediated Type I IFN production.

Role of Enteroviral RNA-Dependent RNA Polymerase in Regulation of MDA5-Mediated Beta Interferon Activation.

Infection by enteroviruses can cause severe neurological complications in humans. The interactions between the enteroviral and host proteins may facilitate the virus replication and be involved in the pathogenicity of infected individuals. It has been shown that human enteroviruses possess various mechanisms to suppress host innate immune responses in infected cells. Previous studies showed that infection by enterovirus 71 (EV71) causes the degradation of MDA5, which is a critical cytoplasmic pathogen sensor in the recognition of picornaviruses for initiating transcription of type I interferons. In the present study, we demonstrated that the RNA-dependent RNA polymerase (RdRP; also denoted 3Dpol) encoded by EV71 interacts with the caspase activation and recruitment domains (CARDs) of MDA5 and plays a role in the inhibition of MDA5-mediated beta interferon (IFN-ß) promoter activation and mRNA expression. In addition, we found that the 3Dpol protein encoded by coxsackievirus B3 also interacted with MDA5 and downregulated the antiviral signaling initiated by MDA5. These findings indicate that enteroviral RdRP may function as an antagonist against the host antiviral innate immune response.IMPORTANCE Infection by enteroviruses causes severe neurological complications in humans. Human enteroviruses possess various mechanisms to suppress the host type I interferon (IFN) response in infected cells to establish viral replication. In the present study, we found that the enteroviral 3Dpol protein (or RdRP), which is a viral RNA-dependent RNA polymerase for replicating viral RNA, plays a role in the inhibition of MDA5-mediated beta interferon (IFN-ß) promoter activation. We further demonstrated that enteroviral 3Dpol protein interacts with the caspase activation and recruitment domains (CARDs) of MDA5. These findings indicate that enteroviral RdRP functions as an antagonist against the host antiviral response.

Altered pathogenicity for seasonal influenza virus by single reassortment of the RNP genes derived from the 2009 pandemic influenza virus.

BACKGROUND: The 2009 influenza A pandemic virus (H1N1(pdm)) may reassort with old seasonal influenza A virus (H1N1141) in humans and potentially change their pathogenicity. METHODS AND RESULTS: This study focuses on the reassortment of ribonucleoproteins (RNPs) among H1N1(pdm) and seasonal influenza A viruses. A single RNP gene reassortment altered reporter gene expression levels driven by polymerase complex in transfection system. The growth rates of recombinant viruses with different RNP recombinations were changed in A549 cells. Mice were infected with recombinant viruses containing single RNP gene reassortment, and pathogenicity was examined. The results demonstrated that the median lethal dose (LD50) of the PB2141/PB1141/PA(pdm)/NP141 recombinant virus was lower than that of the seasonal H1N1 virus. Viral titers of this reassorted virus in the lung and spleen were significantly higher than that in seasonal H1N1 virus-challenged mice. CONCLUSIONS: Although the changes of RNP activity did not exactly reflect to mice virulence, we consistently observed that the PA gene of H1N1(pdm) results in increased polymerase activity, better replication in mice, and lower LD50. Our findings suggest that monitoring of gene reassortment for the 2009 pandemic influenza and seasonal human viruses is also important, which would help to constrain the potential emergence of a more virulent influenza A variant.

Mechanism of action of the suppression of influenza virus replication by Ko-Ken Tang through inhibition of the phosphatidylinositol 3-kinase/Akt signaling pathway and viral RNP nuclear export.

AIMS OF THE STUDY: Ko-Ken Tang (KKT, aka kakkon-to), a conventional Chinese herbal medicine, has been used for the treatment of the common cold, fever and influenza virus infection. However, the underlying mechanism of its activity against influenza virus infection remains elusive. In this study, the antiviral effect and its underlying mechanism was evaluated, including the investigation of anti-influenza virus activity of KKT on MDCK cells and corresponding mechanism related to phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway and its consecutive viral RNP nuclear export. MATERIALS AND METHODS: The antiviral activity of non-toxic concentration of KKT was examined against various strains of influenza virus and enterovirus 71 by neutralization assay. PI3K/Akt signaling activated by influenza virus was inspected in A549 cells by western blot. Inhibition of influenza polymerase activity by KKT was measured with plasmid-based reverse genetics using primer extension assay and luciferase reporter assay. Inhibition of viral vRNP nuclear export was demonstrated by laser confocal microscopy and interspecies heterokaryon assay. RESULTS: KKT inhibits influenza virus replication but not entry, and it exhibits a broad spectrum inhibitory activity against human influenza A viruses and enterovirus 71. KKT does not inhibit viral polymerase activity but directly blocks the virus-induced phosphatidylinositol 3-kinase/Akt signaling pathway, which in turns causes retention of viral nucleoprotein in the nucleus, thereby interfering with virus propagation. The inhibition by KKT of the nuclear export of viral protein was further confirmed by heterokaryon assay. CONCLUSIONS: The results obtained in this study give scientific support to KKT for the treatment of influenza virus infection. KKT could be of potential use in the management of seasonal pandemic influenza virus infection in addition to other clinically available drugs.