The essential, nonredundant roles of RIG-I and MDA5 in detecting and controlling West Nile virus infection

Virus recognition and response by the innate immune system are critical components of host defense against infection. Activation of cell-intrinsic immunity and optimal priming of adaptive immunity against West Nile virus (WNV), an emerging vector-borne virus, depends on recognition by RIG-I and MDA5, cytosolic pattern recognition receptors (PRRs) of the RIG-I-like receptor (RLR) protein family that recognize viral RNA and activate defense programs that suppress infection. We evaluated the individual functions of RIG-I and MDA5 both in vitro and in vivo in pathogen recognition and control of WNV. Lack of RIG-I or MDA5 alone results in decreased innate immune signaling and virus control in primary cells in vitro and increased mortality in mice. We also generated RIG-I -/- x MDA5 -/- double knockout mice and found that lack of both RLRs results in a complete absence of innate immune gene induction in target cells of WNV infection and a severe pathogenesis during infection in vivo, similar to animals lacking MAVS, the central adaptor molecule for RLR signaling. We also found that RNA products from WNV infected cells but not incoming virion RNA display at least two distinct pathogen associated molecular patterns (PAMPs) containing 5' triphosphate and double-stranded RNA that are temporally distributed and sensed by RIG-I and MDA5 during infection. In addition to biochemical characterization of stimulatory PAMPs contained within WNV infected cells, we have also defined sequence identity of RNAs associated with RIG-I and MDA5 immunoprecipitated from infected cells. Thus, RIG-I and MDA5 are essential PRRs that recognize distinct PAMPs that accumulate during WNV replication. This study highlights the necessity and function of multiple related, cytoplasmic host sensors in orchestrating an effective immune response against an acute viral infection.