Fig 1: Isolation of fve-8 as a new mutant with attenuated RNA silencing through a genetic screening.(A) Schematic construct of E5-4 used for the EMS screening in this study. E4 and E5, the fourth and fifth exons of PHB, respectively; PAGO10, the native promoter of AGO10 locus; Nos Ter, nopaline synthase terminator. (B) The established mutations in miRNA and siRNA pathways increased LUC luminescence. Five-day seedlings of Ec5-4/se-2, Ec5-4/hyl1-2, and Ec5-4/rdr6-11 were photographed in bright field (top), under charge-coupled device (CCD) camera for LUC signal (middle), and under regular camera (bottom). The crossing lines of M17-1 with sgs3-1 displayed similar patterns (see also fig. S1A). Scale bars, 0.5 cm. (C) Mutation in FVE caused increased LUC luminescence. Five-day seedlings of ars1-1 (fve-8), E5-4, and fve-8; gFVE were photographed in bright field (top) and under CCD camera for LUC signal (middle). The signals were displayed by LightField software. Scale bars, 1 cm. Bottom: Six-week-old ars1-1/fve-8 and complementation lines displayed later and normal flowering phenotypes, respectively. Scale bars, 1 cm. (D) RNA blot analysis showed LUC transcript accumulated in ars1-1(fve-8) and E5-4. Ribosomal RNA is control. (E) Next-generation mapping (NGM) analysis of F2 mapping population delineated eight candidates in 1.3 Mb of chromosome 2. bp, base pairs. (F) Top: The gene structure of FVE includes untranslated regions (gray boxes), exons (black boxes), and introns (lines). The fve-8 mutation (triangle) is labeled. Bottom: The protein schematic shows a low complexity region (light gray boxes), 6 × WD40 domains (dark gray boxes), and putative nuclear localization signal (NLS; black box).