The DEAD box RNA helicase DDX42 is an intrinsic inhibitor of positive‐strand RNA viruses

Genome‐wide screens are powerful approaches to unravel regulators of viral infections. Here, a CRISPR screen identifies the RNA helicase DDX42 as an intrinsic antiviral inhibitor of HIV‐1. Depletion of endogenous DDX42 increases HIV‐1 DNA accumulation and infection in cell lines and primary cells. DDX42 overexpression inhibits HIV‐1 infection, whereas expression of a dominant‐negative mutant increases infection. Importantly, DDX42 also restricts LINE‐1 retrotransposition and infection with other retroviruses and positive‐strand RNA viruses, including CHIKV and SARS‐CoV‐2. However, DDX42 does not impact the replication of several negative‐strand RNA viruses, arguing against an unspecific effect on target cells, which is confirmed by RNA‐seq analysis. Proximity ligation assays show DDX42 in the vicinity of viral elements, and cross‐linking RNA immunoprecipitation confirms a specific interaction of DDX42 with RNAs from sensitive viruses. Moreover, recombinant DDX42 inhibits HIV‐1 reverse transcription in vitro. Together, our data strongly suggest a direct mode of action of DDX42 on viral ribonucleoprotein complexes. Our results identify DDX42 as an intrinsic viral inhibitor, opening new perspectives to target the life cycle of numerous RNA viruses.