In Vivo RNAi Screening Identifies MDA5 as a Significant Contributor to the Cellular Defense against Influenza A Virus

Responding to an influenza A virus (IAV) infection demands an effective intrinsic cellular defense strategy to slow replication. To identify contributing host factors to this defense, we exploited the host microRNA pathway to perform an in vivo RNAi screen. To this end, IAV, lacking a functional NS1 antagonist, was engineered to encode individual siRNAs against antiviral host genes in an effort to rescue attenuation. This screening platform resulted in the enrichment of strains targeting virus-activated transcription factors, specific antiviral effectors, and intracellular pattern recognition receptors (PRRs). Interestingly, in addition to RIG-I, the PRR for IAV, a virus with the capacity to silence MDA5 also emerged as a dominant strain in wild-type, but not in MDA5-deficient mice. Transcriptional profiling of infected knockout cells confirmed RIG-I to be the primary PRR for IAV but implicated MDA5 as a significant contributor to the cellular defense against influenza A virus. [Display omitted] •Self-inactivating viruses can be engineered using artificial miRNAs•Virus-derived miRNAs can partially restore replication of attenuated mutants•Demonstration of a fitness-based screen using miRNA-enabled influenza viruses•miRNA-based screen implicates host factors involved in restricting influenza virus Benitez et al. describe an influenza-A-virus-driven siRNA screen performed in the context of an in vivo infection. This fitness-based screening platform identified virus strains encoding siRNAs targeting interferon inducing transcription factors, antiviral effectors, and intracellular pattern recognition receptors, including MDA5, involved in repressing influenza A virus replication.