ANI-2 and ANI-1 have opposing activities in germline organization. (A) Mid-section confocal images of the germline of wild-type L4 (top) and adult (bottom) hermaphrodites, treated with control(RNAi) or ani-1(RNAi), and expressing GFP::ANI-2 (green) and a membrane marker (red). (B) Proportion of germ cells showing rachis bridges with a diameter >0.8 µm (turquoise) or <0.8 µm (red) in animals of the specified condition, as measured by fluorescent marker distribution. (C) Maximal rachis bridge diameter in germ cells of animals of the specified condition, as measured with membrane (red) or GFP::ANI-2 (green) fluorescent distribution. (D) Mid-section confocal images of the germline of wild-type and ani-2(−) young adult hermaphrodites (20 h post-L4), treated with ani-1(RNAi), and expressing a membrane marker (green) and mCherry::Histone H2B (red). Arrowheads indicate germ cells with open rachis bridges. (E) Proportion of germ cells showing rachis bridges with a diameter >0.8 µm (turquoise) or <0.8 µm (red) in animals of the specified condition, as measured by fluorescent marker distribution. (F) Maximal rachis bridge diameter in germ cells of animals of the specified condition, as measured with membrane fluorescent distribution. (G) Average number of multinucleated germ cells in various germline regions of ani-2(−) mutant hermaphrodites (20 h post-L4) treated with control(RNAi) or ani-1(RNAi). Error bars represent SD over 10–17 animals at each stage. (H) Proportion of multinucleated germ cells with 2, 3, 4, 5–10, and >10 nuclei in various germline regions of ani-2(−) hermaphrodites (20 h post-L4) treated with control(RNAi) or ani-1(RNAi). In B, C, E, F, and H, the numbers in brackets represent the total number of germ cells analyzed. Bars, 10 µm. *, P < 0.01; **, P < 0.001.