Fig. 4. GLH/VASA mutants exhibit defects in producing secondary WAGO-22G-RNAs.
a A scatter plot showing the abundance of all 22G-RNAs (left) or HRDE-1 bound 22G-RNAs (right) mapped to each WAGO target in wild-type worms compared to indicated strains. (Left) The percentage of WAGO targets with 2-fold increased or decreased 22G-RNAs in mutants are shown. (Right) The percentage of significantly changed (adjusted P < 0.05 [see Methods for details] and 2-fold) WAGO targets are shown. The three diagonal lines indicate a two-fold increase (top), no change (middle), or a two-fold depletion (bottom) in the indicated mutant strains. b A scatter plot showing the mRNA vs 22G-RNA log2 expression changes for WAGO targets in glh-1 glh-4 mutants vs wild type worms. The upper left quadrant corresponds to WAGO targets that have become activated in the mutant (increased mRNAs and decreased 22G-RNAs). Percentages are shown in each quadrant to indicate the proportion of WAGO targets with reduced HRDE-1-associated 22G-RNAs in the glh-1 mutant that fall in that quadrant. c 22G-RNA changes for WAGO targets with reduced HRDE-1-associated 22G-RNAs in the glh-1 mutant versus all other WAGO targets in the indicated strains versus wild type worms. Statistical analysis was performed using a two-tailed Mann–Whitney Wilcoxon test. For all boxplots, lines display median values, boxes display first and third quartiles, and whiskers display 5th and 95th percentiles. d Density of 22G-RNAs within a 100-nt window around predicted piRNA target sites in the indicated strains. Computed by summing 22G-RNA density per piRNA targeting site in WAGO targets with reduced HRDE-1 associated 22G-RNAs in the glh-1 mutant. The plots are centered on the 10th nucleotide of piRNAs.