FIG 8.

Identification of rsd-6 as key component of antiviral RNAi in C. elegans. (A) Schematic gene structures for wild-type rsd-6 and rsd-6 alleles isolated in the genetic screen. Exons are shown as solid boxes, whereas the lines indicate introns. The putative protein sizes from conceptual translation are indicated for each allele. (B) Schematic structure of the plasmid construct that was used for constitutive expression of wild-type rsd-2 gene in transgenic worms. RSD-6, the open reading frame of wild-type rsd-6. (C) Ectopic expression of wild-type rsd-6 restored antiviral RNAi in worm mutants containing the 1026a alleles. Shown here is the visualization of green fluorescence in the 1026a mutants that have been injected with plasmid construct Psur-5::RSD-6. The transgenic worm is marked by red fluorescence in the head region. (D) Constitutive overexpression of wild-type rsd-6 restored antiviral RNAi in worm mutants containing the 1026a alleles. Shown here is the detection of FR1gfp genomic and subgenomic RNA accumulation after heat induction. An asterisk indicates 1026a mutants that contain integrated transgene derived from the plasmid construct Psur-5::RSD-6. (E) Same as panel D, except that the replication of Orsay virus RNA1 was detected through Northern blotting. The replication of Orsay virus RNA1 in drh-1 loss-of-function mutants was detected as the reference.