RNA virus infection results in expression of type 1 IFNs, especially IFN-α/β, which play a crucial role in host antivirus responses. Type 1 IFNs are induced in a cell type-specific manner through TLR and RIG-I–like receptor pathways, both of which activate IFN regulatory factors (IRFs) and NF-κB transcription factors. Although NF-κB activation and association with the IFN-β promoter after RNA virus infection is well documented, our previous work showed that, surprisingly, NF-κB is not essential for IFN-β gene expression. Thus, the actual function of NF-κB in IFN-β expression and virus replication is not clear. In this study, we found Newcastle disease virus and vesicular stomatitis virus replication is enhanced in mouse embryonic fibroblasts (MEFs) lacking the NF-κB RelA subunit. Increased virus replication was traced to a specific requirement for RelA in early virus-induced IFN-β expression. At these time points, when IFN-β expression is ~100-fold less than peak levels, impaired IFN-β production delayed IFN-induced gene expression, resulting in increased virus replication in RelA^−/− MEFs. Importantly, our results show that RelA requirement is crucial only when IRF3 activation is low. Thus, high levels of activated IRF3 expression are sufficient for induction of IFN-β in RelA^−/− MEFs, transcriptional synergism with the coactivator CREB-binding protein, and rescue of susceptibility to virus. Together, these findings indicate that NF-κB RelA is not crucial for regulating overall IFN-β production, as previously believed; instead, RelA is specifically required only during a key early phase after virus infection, which substantially impacts the host response to virus infection.

Interferons (IFNs) are cytokines with well-described immunomodulatory and antiviral properties, but less is known about the mechanisms by which they promote cell survival or cell death. Here, we show that IFN-γ induces RIP1 kinase-dependent necroptosis in mammalian cells deficient in NF-κB signaling. Induction of necroptosis by IFN-γ was found to depend on Jak1 and partially on STAT1. We also demonstrate that IFN-γ activates IκB kinase β (IKKβ)-dependent NF-κB to regulate a transcriptional program that protects cells from necroptosis. IFN-γ induced progressive accumulation of reactive oxygen species (ROS) and eventual loss of mitochondrial membrane potential in cells lacking the NF-κB subunit RelA. Whole-genome microarray analyses identified sod2, encoding the antioxidant enzyme manganese superoxide dismutase (MnSOD), as a RelA target and potential antinecroptotic gene. Overexpression of MnSOD inhibited IFN-γ-mediated ROS accumulation and partially rescued RelA-deficient cells from necroptosis, while RNA interference (RNAi)-mediated silencing of sod2 expression increased susceptibility to IFN-γ-induced cell death. Together, these studies demonstrate that NF-κB protects cells from IFN-γ-mediated necroptosis by transcriptionally activating a survival response that quenches ROS to preserve mitochondrial integrity.

Receptor-interacting protein kinase 1 (RIPK1) regulates cell fate and proinflammatory signaling downstream of multiple innate immune pathways, including those initiated by TNF-α, TLR ligands, and IFNs. Genetic ablation of Ripk1 results in perinatal lethality arising from both RIPK3-mediated necroptosis and FADD/caspase-8–driven apoptosis. IFNs are thought to contribute to the lethality of Ripk1-deficient mice by activating inopportune cell death during parturition, but how IFNs activate cell death in the absence of RIPK1 is not understood. In this study, we show that Z-form nucleic acid binding protein 1 (ZBP1; also known as DAI) drives IFN-stimulated cell death in settings of RIPK1 deficiency. IFN-activated Jak/STAT signaling induces robust expression of ZBP1, which complexes with RIPK3 in the absence of RIPK1 to trigger RIPK3-driven pathways of caspase-8–mediated apoptosis and MLKL-driven necroptosis. In vivo, deletion of either Zbp1 or core IFN signaling components prolong viability of Ripk1−/− mice for up to 3 mo beyond parturition. Together, these studies implicate ZBP1 as the dominant activator of IFN-driven RIPK3 activation and perinatal lethality in the absence of RIPK1.

Reverse transcription, an essential step in the life cycle of all retroelements, is a complex, multistep process whose regulation is not yet clearly understood. We have recently shown that reverse transcription in the pararetrovirus duck hepatitis B virus is associated with complete dephosphorylation of the viral core protein, which forms the nucleocapsid wherein reverse transcription takes place. Here we present a genetic study of the role of this dynamic nucleocapsid phosphorylation in regulating viral reverse transcription. Detailed analyses of the reverse transcription products synthesized within nucleocapsids composed of core phosphorylation site mutants revealed that alanine substitutions, mimicking the nonphosphorylated state, completely blocked reverse transcription at a very early stage. In contrast, aspartate substitutions, mimicking the phosphorylated state, allowed complete first-strand DNA synthesis but were severely defective in accumulating mature double-stranded DNA. The latter defect was due to a combination of mutant nucleocapsid instability during maturation and a block in mature second-strand DNA synthesis. Thus, the reversible phosphorylation of the nucleocapsids regulates the ordered progression of reverse transcription.

A mass vaccination campaign in India seeks to control and eventually eradicate foot-and-mouth disease (FMD). Biosanitary measures along with FMD monitoring are being conducted along with vaccination. The implementation of the FMD control program has drastically reduced the incidence of FMD. However, cases are still reported, even in regions where vaccination is carried out regularly. Control of FMD outbreaks is difficult when the virus remains in circulation in the vaccinated population. Various FMD risk factors have been identified that are responsible for FMD in vaccinated areas. The factors are discussed along with strategies to address these challenges. The current chemically inactivated trivalent vaccine formulation containing strains of serotype O, A, and Asia 1 has limitations including thermolability and induction of only short-term immunity. Advantages and disadvantages of several new-generation alternate vaccine formulations are discussed. It is unfeasible to study every incidence of FMD in vaccinated animals/areas in such a big country as India with its huge livestock population. However, at the same time, it is absolutely necessary to identify the precise reason for vaccination failure. Failure to vaccinate is one reason for the occurrence of FMD in vaccinated areas. FMD epidemiology, emerging and re-emerging virus strains, and serological status over the past 10 years are discussed to understand the impact of vaccination and incidences of vaccination failure in India. Other factors that are important in vaccination failure that we discuss include disrupted herd immunity, health status of animals, FMD carrier status, and FMD prevalence in other species. Recommendations to boost the search of alternate vaccine formulation, strengthen the veterinary infrastructure, bolster the real-time monitoring of FMD, as well as a detailed investigation and documentation of every case of vaccination failure are provided with the goal of refining the control program.

Production of type I interferons (IFNs; prominently, IFN-α/β) following virus infection is a pivotal antiviral innate immune response in higher vertebrates. The synthesis of IFN-β proceeds via the virus-induced assembly of the transcription factors IRF-3/7, ATF-2/c-Jun, and NF-κB on the ifn β promoter. Surprisingly, recent data indicate that the NF-κB subunit RelA is not essential for virus-stimulated ifn β expression. Here, we show that RelA instead sustains autocrine IFN-β signaling prior to infection. In the absence of RelA, virus infection results in significantly delayed ifn β induction and consequently defective secondary antiviral gene expression. While RelA is not required for ifn β expression after infection, it is nonetheless essential for fully one-fourth of double-stranded RNA (dsRNA)-activated genes, including several mediators of inflammation and immune cell recruitment. Further, RelA directly regulates a small subset of interferon-stimulated genes (ISGs). Finally, RelA also protects cells from dsRNA-triggered RIP1-dependent programmed necrosis. Taken together, our findings suggest distinct roles for RelA in antiviral innate immunity: RelA maintains autocrine IFN-β signaling in uninfected cells, facilitates inflammatory and adaptive immune responses following infection, and promotes infected-cell survival during this process.