Structure and Function of RNA Silencing Suppressors

RNA silencing in plants likely exists as a defense mechanism against molecular parasites such as RNA viruses, retrotransposons and transgenes. As a result, many plant viruses have adapted mechanisms to suppress gene silencing. Tombusviruses express a 19 kDa protein (p19) that suppresses RNA silencing in vivo and binds small interfering RNAs (siRNAs) in vitro. p19 likely sequesters siRNAs, thus depleting this RNA silencing effector molecule. We determined the crystal structure of p19 from the Carnation Italian ringspot virus (CIRV) bound to a 21‐nt siRNA and demonstrate that CIRV p19 protein acts as a molecular caliper to specifically select siRNAs based on the length of the duplex region of the siRNA. CIRV p19 binds tightly to siRNAs of 20–22‐nt, but progressively weaker to siRNAs of 23–26‐nt and poorly to a 19‐nt siRNA. Thus, since plants produce two size classes of siRNA, CIRV p19 likely inhibits the mRNA degradation directed by shorter siRNAs and has lesser effects on the processes associated with longer siRNAs. CIRV p19 also inhibits microRNA (miRNA)‐directed processes when expressed in plants. In order to understand the effects of mismatched or bulged nucleotides in miRNA duplexes on CIRV p19 binding, we have examined the binding of imperfect duplex RNA to CIRV p19. Furthermore, we examine the binding of other viral RNA silencing suppressors to siRNAs and different mechanisms of RNA silencing suppression.