Innate immune response is a universal mechanism against invading pathogens. Toll-like receptors (TLRs), being part of a first line of defense, are responsible for detecting a variety of microorganisms. Among them TLR9, which is localized in endosomes, acts as a sensor for unmethylated CpG motifs present in bacteria, DNA viruses (e.g., HSV-1), or fungi. TLRs differ from one another by the use of accessory proteins. MyD88 adapter-like (Mal) adapter molecule is considered a positive regulator of TLR2-and TLR4-dependent pathways. It has been reported that this adapter may also negatively control signal transduction induced by TLR3 anchored in the endosome membrane. So far, the role of Mal adapter protein in the TLR9 signaling pathways has not been clarified. We show for the first time that Mal is engaged in TLR9-dependent expression of genes encoding IFNβ and TNFα in HSV-1-infected or CpG-C-treated macrophages and requires a noncanonical NF-κB pathway. Moreover, using inhibitor of ERK1/2 we confirmed involvement of these kinases in TLR9-dependent induction of IFNβ and TNFα. Our study points to a new role of Mal in TLR9 signaling through a hitherto unknown mechanism whereby lack of Mal specifically impairs ERK1/2-mediated induction of noncanonical NF-κB pathway and concomitant IFNβ and TNFα production.

Sensing of viral particles and elements that initiate mechanisms of immune response is an intrinsic ability of mammalian cells. Regulatory cytokines and antiviral mediators are released after triggering of complex signaling cascades in response to interaction of pathogen particles with pattern recognition receptors (PRRs) leading to the production of interferons (IFN) and proinflammatory cytokines. Viral RNA in the cytoplasm constitute a potent danger molecule that recognition is performed by RIG-I-like receptors, the most common group of receptors in mammalian cells, capable to recognize a foreign RNA. It is known that the E3 ubiquitin ligase Pellino3 plays an important role in antibacterial and antiviral response, but its involvement in the RLR pathways remains poorly understood. In this study, we investigate the molecular mechanisms of the innate immune response in BMDMs (immortalized macrophages from mouse bone marrow) during VSV infection. Here, we present evidence that the activation of the RIG-I/Pellino3/ERK1/2 pathway in BMDMs is crucial for the protection against VSV. We demonstrate that during infection, viral particles replicate in Pellino3 knockout BMDMs more effectively than in wild-type cells. Increased viral replication resulting in cell lysis and death is aid by impaired synthesis of IFN-I and inflammatory cytokines as a consequence of disturbances in the ERK1/2 pathway regulation.

One of the key determinants of survival for organisms is proper recognition of exogenous and endogenous nucleic acids. Therefore, high eukaryotes developed a number of receptors that allow for discrimination between friend or foe DNA and RNA. Appearance of exogenous RNA in cytoplasm provides a signal of danger and triggers cellular responses that facilitate eradication of a pathogen. Recognition of exogenous RNA is additionally complicated by fact that large amount of endogenous RNA is present in cytoplasm Thus, number of different receptors, found in eukaryotic cells, is able to recognize that nucleic acid. First group of those receptors consist endosomal Toll like receptors, namely TLR3, TLR7, TLR8 and TLR13. Those receptors recognize RNA released from pathogens that enter the cell by endocytosis. The second group includes cytoplasmic sensors like PKR and the family of RLRs comprised of RIG-I, MDA5 and LGP2. Cytoplasmic receptors recognize RNA from pathogens invading the cell by non-endocytic pathway. In both cases binding of RNA by its receptors results in activation of the signalling cascades that lead to the production of interferon and other cytokines.