Viral RNA recognition by LGP2 and MDA5, and activation of signaling through step-by-step conformational changes.

Cytoplasmic RIG-I-like receptor (RLR) proteins in mammalian cells recognize viral RNA and initiate an antiviral response that results in IFN-ß induction. Melanoma differentiation-associated protein 5 (MDA5) forms fibers along viral dsRNA and propagates an antiviral response via a signaling domain, the tandem CARD. The most enigmatic RLR, laboratory of genetics and physiology (LGP2), lacks the signaling domain but functions in viral sensing through cooperation with MDA5. However, it remains unclear how LGP2 coordinates fiber formation and subsequent MDA5 activation. We utilized biochemical and biophysical approaches to observe fiber formation and the conformation of MDA5. LGP2 facilitated MDA5 fiber assembly. LGP2 was incorporated into the fibers with an average inter-molecular distance of 32 nm, suggesting the formation of hetero-oligomers with MDA5. Furthermore, limited protease digestion revealed that LGP2 induces significant conformational changes on MDA5, promoting exposure of its CARDs. Although the fibers were efficiently dissociated by ATP hydrolysis, MDA5 maintained its active conformation to participate in downstream signaling. Our study demonstrated the coordinated actions of LGP2 and MDA5, where LGP2 acts as an MDA5 nucleator and requisite partner in the conversion of MDA5 to an active conformation. We revealed a mechanistic basis for LGP2-mediated regulation of MDA5 antiviral innate immune responses.

KD-409, a Respiratory Syncytial Virus FG Chimeric Protein without the CX3C Chemokine Motif, Is an Efficient Respiratory Syncytial Virus Vaccine Preparation for Passive and Active Immunization in Mice.

Although respiratory syncytial virus (RSV) vaccine development initiatives have existed for half a century, no candidate has been approved for application at all ages from neonates to children. Developing an effective and safe RSV vaccine for pediatric use is challenging owing to RSV-associated disease and vaccine-enhanced disease (VED). We aimed to design an RSV vaccine, KD-409, by structurally incorporating the F ectodomain and G protein central conserved domain without the CX3C chemokine motif and test its efficacy and safety. KD-409 formed rosette particles or trimmers. KD-409 immunization of mice mainly induced anti-RSV F protein IgG. The induced anti-F antibodies had a higher IgG2a/IgG1 ratio than pre-fusion F, suggesting that they induced Th1-dominant immunity. Active and passive immunities were assessed by analyzing the viral titers in BALB/c mice intranasally challenged with RSV after intramuscular KD-409 immunization and pups derived from mothers who were intramuscularly vaccinated with KD-409 twice, respectively. KD-409 was more effective than post-fusion F and had a lower minimum effective dose than pre-fusion F. Thus, KD-409 demonstrated great potential as a novel RSV vaccine candidate, outperforming existing RSV F-based candidates. Our findings provide a promising strategy to overcome RSV-associated acute lower respiratory infections without the risk of VED associated with traditional approaches.