Fig. 4
From: Methyl group donors abrogate adaptive responses to dietary restriction in C. elegans
Choline and methionine abrogate the enlargement of the lipid droplets of in utero embryos (F1) produced by dietarily restricted mothers (Po). a DIC images of in utero embryos (F1 generation) derived from ad libitum-fed (AL, a) and dietarily restricted (DR, b) adult worms (P0 generation) supplemented with choline (c) or methionine (d). Images were captured using a ×63 objective. Scale bar, 20 μm. Enlargements of the framed areas are shown in a´–d´ (×5, scale bar, 10 μm). b The lipid droplet marker ATGL::GFP (a, a´) and the vitellogenin marker Vit-2::GFP (b, b´) are shown. Images were captured using a ×100 objective. Scale bar, 20 μm; ×5 magnification a´, b´, scale bar, 10 μm. c Lipid droplets from isolated embryos obtained from ad libitum-fed (AL, a) and dietarily restricted (DR, b) adult worms were visualized using short-term vital BODIPY™ 493/503 staining. Merged DIC and BODIPY™ 493/503 fluorescence images (a, b) and fluorescence images (a´, b´) are shown. Image stacks (a´, b´) with a depth of 10 μm, consisting of 21 focal planes at 0.5 μm intervals, were captured and rendered as maximum projections of the BODIPY™ 493/503 signals. d Quantification of intestinal lipid droplets in ad libitum-fed (AL) L1 larvae (F1 generation; 24 h after hatching), L1/L2 larvae (F1 generation; 30 h after hatching), and L4 (F1 generation; 48 h after hatching) obtained from ad libitum-fed (AL) and dietarily restricted (DR) adult worms (P0 generation). Analyses were performed using the 3D spot segmentation plug-in of ImageJ software. The distribution of the relative lipid-droplet volume/fraction ratio per lipid droplet size category (<1, 1–3, 3–5, >5 μm3) is shown. The data were derived from six to nine individual worms per feeding condition in three independent experiments. ALAL24/30/48, ad libitum feeding of the P0 generation and ad libitum feeding of F1 larvae 24, 30, or 48 h after hatching; DRAL24/30/48, dietary restriction of the P0 generation and ad libitum feeding of F1 larvae 24, 30, or 48 h after hatching