Abstract
DEAD box RNA helicases regulate diverse facets of RNA biology. Proteins of this family carry out essential cellular functions, and emerging literature is revealing additional roles in immune defense. Using RNA interference screening, we identified an evolutionarily conserved antiviral role for the helicase DDX56 against the alphavirus Sindbis virus (SINV), a mosquito-transmitted pathogen that infects humans. Depletion of DDX56 enhanced infection in Drosophila and human cells. Furthermore, we found that DDX56 also controls the emerging alphavirus chikungunya virus (CHIKV) through an interferon-independent mechanism. Using cross-linking immunoprecipitation (CLIP-Seq), we identified a predicted stem-loop on the viral genomic RNA bound by DDX56. Mechanistically, we found that DDX56 levels increase in the cytoplasm during CHIKV infection. In the cytoplasm, DDX56 impacts the earliest step in the viral replication cycle by binding and destabilizing the incoming viral genomic RNA, thereby attenuating infection. Thus, DDX56 is a conserved antiviral RNA binding protein that controls alphavirus infection.IMPORTANCE Arthropod-borne viruses are diverse pathogens and include the emerging virus chikungunya virus, which is associated with human disease. Through genetic screening, we found that the conserved RNA binding protein DDX56 is antiviral against chikungunya virus in insects and humans. DDX56 relocalizes from the nucleus to the cytoplasm, where it binds to a stem-loop in the viral genome and destabilizes incoming genomes. Thus, DDX56 is an evolutionarily conserved antiviral factor that controls alphavirus infection.
Highlights
DEAD box RNA helicases regulate diverse facets of RNA biology
We identified antiviral helicases using a cutoff of Ͼ2-fold increase in infection; of the 22 DEAD box helicases in our panel, six exceeded this threshold, and five of those had no impact on cell survival
We tested a panel of human orthologs of the antiviral Drosophila helicases, using the human osteosarcoma cell line U2OS, as it is readily infected by many arthropod-borne viruses and has an intact interferon system
Summary
DEAD box RNA helicases regulate diverse facets of RNA biology Proteins of this family carry out essential cellular functions, and emerging literature is revealing additional roles in immune defense. Key sensors of viral RNA, including the canonical cytosolic RNA sensors, retinoic acid-inducible gene I (RIG-I/DDX58) and melanoma differentiation-associated protein 5 (MDA-5/IFIH1), are members of the DExD/H family of RNA binding proteins [2] These proteins are closely related to the larger DEAD box helicase family, which is the largest RNA helicase family and is deeply conserved. In addition to the canonical cytoplasmic RNA sensors RIG-I and MDA5, emerging data suggest that many DEAD box helicases have roles in viral RNA recognition and cellular antiviral responses. It is likely that many other helicases have uncharacterized antiviral activity
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