DLG1 promotes the antiviral innate immune response by inhibiting p62-mediated autophagic degradation of IKKε.

IMPORTANCE: The type-I interferon (IFN-I) signaling pathway is the first line of antiviral innate immunity. It must be precisely regulated against virus-induced damage. The tightly regulated mechanisms of action of host genes in the antiviral innate immune signaling pathway are still worth studying. Here, we report a novel role of DLG1 in positively regulating the IκB kinase epsilon (IKKε)-mediated IFN-I signaling response against negative-stranded RNA virus replication, whereas the RNA virus inhibits the expression of DLG1 for immune escape. Importantly, the E3 ligase March2 interacts with and promotes K27-linked polyubiquitination of IKKε, and p62 is a cargo receptor that recognizes ubiquitinated IKKε for eventual autophagic degradation. Together, the current findings elucidate the role of DLG1 in the antiviral IFN-I signaling pathway and viral infection repression.

Type I interferon receptor-independent interferon-α induction upon infection with a variety of negative-strand RNA viruses.

Type I interferons (IFNs) are a first line of defence against viral infections. Upon infection, a first small wave of early type I IFN, mainly IFN-ß and particularly IFN-α4, are induced and bind to the type I IFN receptor (IFNAR) to amplify the IFN response. It was shown for several viruses that robust type I IFN responses require this positive feedback loop via the IFNAR. Recently, we showed that infection of IFNAR knockout mice with the orthomyxovirus Thogoto virus lacking the ML open reading frame (THOV(ML-)) results in the expression of unexpected high amounts of type I IFN. To investigate if IFNAR-independent IFN responses are unique for THOV(ML-), we performed infection experiments with several negative-strand RNA viruses using different routes and dosages for infection. A variety of these viruses induced type I IFN responses IFNAR-independently when using the intraperitoneal (i.p.) route for infection. In vitro studies demonstrated that myeloid dendritic cells (mDC) are capable of producing IFNAR-independent IFN-α responses that are dependent on the expression of the adaptor protein mitochondrial antiviral-signalling protein (MAVS) whereas pDC where entirely depending on the IFNAR feedback loop in vitro. Thus, depending on dose and route of infection, the IFNAR feedback loop is not strictly necessary for robust type I IFN expression and an IFNAR-independent type I IFN production might be the rule rather than the exception for infections with numerous negative-strand RNA viruses.

Nonsegmented Negative-Sense RNA Viruses Utilize N6-Methyladenosine (m6A) as a Common Strategy To Evade Host Innate Immunity.

N6-Methyladenosine (m6A) is the most abundant internal RNA modification catalyzed by host RNA methyltransferases. As obligate intracellular parasites, many viruses acquire m6A methylation in their RNAs. However, the biological functions of viral m6A methylation are poorly understood. Here, we found that viral m6A methylation serves as a molecular marker for host innate immunity to discriminate self from nonself RNA and that this novel biological function of viral m6A methylation is universally conserved in several families in nonsegmented negative-sense (NNS) RNA viruses. Using m6A methyltransferase (METTL3) knockout cells, we produced m6A-deficient virion RNAs from the representative members of the families Pneumoviridae, Paramyxoviridae, and Rhabdoviridae and found that these m6A-deficient viral RNAs triggered significantly higher levels of type I interferon compared to the m6A-sufficient viral RNAs, in a RIG-I-dependent manner. Reconstitution of the RIG-I pathway revealed that m6A-deficient virion RNA induced higher expression of RIG-I, bound to RIG-I more efficiently, enhanced RIG-I ubiquitination, and facilitated RIG-I conformational rearrangement and oligomerization. Furthermore, the m6A binding protein YTHDF2 is essential for suppression of the type I interferon signaling pathway, including by virion RNA. Collectively, our results suggest that several families in NNS RNA viruses acquire m6A in viral RNA as a common strategy to evade host innate immunity.IMPORTANCE The nonsegmented negative-sense (NNS) RNA viruses share many common replication and gene expression strategies. There are no vaccines or antiviral drugs for many of these viruses. We found that representative members of the families Pneumoviridae, Paramyxoviridae, and Rhabdoviridae among the NNS RNA viruses acquire m6A methylation in their genome and antigenome as a means to escape recognition by host innate immunity via a RIG-I-dependent signaling pathway. Viral RNA lacking m6A methylation induces a significantly higher type I interferon response than m6A-sufficient viral RNA. In addition to uncovering m6A methylation as a common mechanism for many NNS RNA viruses to evade host innate immunity, this study discovered a novel strategy to enhance type I interferon responses, which may have important applications in vaccine development, as robust innate immunity will likely promote the subsequent adaptive immunity.

Tilapia lake virus downplays innate immune responses during early stage of infection in Nile tilapia (Oreochromis niloticus).

Tilapia lake virus (TiLV) causes high mortality and high economic losses in tilapines. We describe an experimental challenge study focusing on early post challenge innate immune responses. Nile tilapia (Oreochromis niloticus) were infected with 105 TCID50/mL TiLV intraperitoneally, followed by virus quantification, histopathology and gene expression analysis in target (brain/liver) and lymphoid (spleen/headkidney) organs at 3, 7, 12, 17, and 34 days post challenge (dpc). Onset of mortality was from 21 dpc, and cumulative mortality was 38.5% by 34 dpc. Liver and kidney histopathology developed over the period 3-17 dpc, characterized by anisocytosis, anisokaryocytosis, and formation of multinucleated hepatocytes. Viral loads were highest at early time (3 dpc) in liver, spleen and kidney, declining towards 34 dpc. In brain, viral titer peaked 17 dpc. Innate sensors, TLRs 3/7 were inversely correlated with virus titer in brain and headkidney, and IFN-ß and Mx showed a similar pattern. All organs showed increased mRNA IgM expression over the course of infection. Overall, high virus titers downplay innate responses, and an increase is seen when viral titers decline. In silico modeling found that TiLV segments 4, 5 and 10 carry nucleolar localization signals. Anti-viral effects of TiLV facilitate production of virus at early stage of infection.