Spatiotemporal dynamics of innate immune signaling via RIG-I-like receptors.

RIG-I, MDA5, and LGP2 comprise the RIG-I-like receptors (RLRs). RIG-I and MDA5 are essential pathogen recognition receptors sensing viral infections while LGP2 has been described as both RLR cofactor and negative regulator. After sensing and binding to viral RNA, including double-stranded RNA (dsRNA), RIG-I and MDA5 undergo cytosol-to-membrane relocalization to bind and signal through the MAVS adaptor protein on intracellular membranes, thus directing downstream activation of IRF3 and innate immunity. Here, we report examination of the dynamic subcellular localization of all three RLRs within the intracellular response to dsRNA and RNA virus infection. Observations from high resolution biochemical fractionation and electron microscopy, coupled with analysis of protein interactions and IRF3 activation, show that, in resting cells, microsome but not mitochondrial fractions harbor the central components to initiate innate immune signaling. LGP2 interacts with MAVS in microsomes, blocking the RIG-I/MAVS interaction. Remarkably, in response to dsRNA treatment or RNA virus infection, LGP2 is rapidly released from MAVS and redistributed to mitochondria, temporally correlating with IRF3 activation. We reveal that IRF3 activation does not take place on mitochondria but instead occurs at endoplasmic reticulum (ER)-derived membranes. Our observations suggest ER-derived membranes as key RLR signaling platforms controlled through inhibitory actions of LGP2 binding to MAVS wherein LGP2 translocation to mitochondria releases MAVS inhibition to facilitate RLR-mediated signaling of innate immunity.

Flavivirus Nonstructural Protein NS5 Dysregulates HSP90 to Broadly Inhibit JAK/STAT Signaling.

Pathogenic flaviviruses antagonize host cell Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling downstream of interferons α/ß. Here, we show that flaviviruses inhibit JAK/STAT signaling induced by a wide range of cytokines beyond interferon, including interleukins. This broad inhibition was mapped to viral nonstructural protein 5 (NS5) binding to cellular heat shock protein 90 (HSP90), resulting in reduced Janus kinase-HSP90 interaction and thus destabilization of unchaperoned JAKs (and other kinase clients) of HSP90 during infection by Zika virus, West Nile virus, and Japanese encephalitis virus. Our studies implicate viral dysregulation of HSP90 and the JAK/STAT pathway as a critical determinant of cytokine signaling control during flavivirus infection.

RNA-binding protein isoforms ZAP-S and ZAP-L have distinct antiviral and immune resolution functions.

The initial response to viral infection is anticipatory, with host antiviral restriction factors and pathogen sensors constantly surveying the cell to rapidly mount an antiviral response through the synthesis and downstream activity of interferons. After pathogen clearance, the host's ability to resolve this antiviral response and return to homeostasis is critical. Here, we found that isoforms of the RNA-binding protein ZAP functioned as both a direct antiviral restriction factor and an interferon-resolution factor. The short isoform of ZAP bound to and mediated the degradation of several host interferon messenger RNAs, and thus acted as a negative feedback regulator of the interferon response. In contrast, the long isoform of ZAP had antiviral functions and did not regulate interferon. The two isoforms contained identical RNA-targeting domains, but differences in their intracellular localization modulated specificity for host versus viral RNA, which resulted in disparate effects on viral replication during the innate immune response.