Fig. 5: The mitochondria-AMPK-DBL-1/TGFβ axis is indispensable for MAE-mediated offspring adult size shaping.

a Schematic illustration of C. elegans DBL-1/TGFβ pathway. b Adult body length comparisons between OD2 and OD5 animals from the indicated DBL-1/TGFβ signaling mutant mothers. c Adult body length comparisons between OD2 and OD5 animals born to sma-3(e491) mutant mothers with the indicated global (sma-3) or tissue-specific SMA-3 rescue in the pharynx (myo-2), intestine (vha-6) and hypodermis (vha-7). d Adult body length comparisons between OD2 and OD5 offspring of dbl-1(ok3749) mutant mothers with a neuron-specific rescue driven by the unc-17 promoter. e, f The expression of dbl-1 gene (e) and the secretion of DBL-1 protein (f) in OD2 and OD5 animals in the indicated stages. The scale bars represent 20 μm (e) and 5 μm (f), respectively. g, h Adult body length measurements in unc-3(tm4776) mutant animals challenged by MAE (g) or treated with AICAR (h) in early-life stages (L1–L4). i The transcriptional changes of dbl-1 indicated by the dbl-1-RFP reporter upon treatment with indicated RNAi and/or compound from P0 mothers to F1 L2 stage. The scale bars represent 20 μm. j Relative DBL-1 secretion in L2 offspring from mothers with the indicated treatments. Rotenone (Rot, 5 μM) or AICAR (1 mM) was applied. The examined head coelomocytes are highlighted with white dotted lines. The scale bars represent 5 μm. k Adult offspring body length measurements in unc-3(tm4776) mutant animals treated with 5 μM Rot at D2 maternal stage for 12 h. l Adult body length measurements in offspring of WT and sma-3(e491) mutant mother worms treated with 1 mM AICAR or vehicle (Veh). m, n DBL-1/TGFβ signaling activity test using the RAD-SMAD reporter in OD2 and OD5 animals (m) or AICAR-treated offspring (n) born to WT, dbl-1(wk70) or aak-2(ok524) mutant animals. The scale bars represent 5 μm. o Adult body length measurements in OD2 animals of WT, dbl-1(wk70) or aak-2(ok524) mutant mothers. These OD2 animals were treated with 1 mM AICAR or Veh in early-life stages. p Annulus width measurements in tail regions of OD2 and OD5 worms. q Levamisole sensitivity assay for OD2 and OD5 animals born to WT or sma-3(e491) mutant mothers. r Fat deposit assay for OD2 and OD5 animals born to sma-3(e491) mutants. s Chemotaxis assay for OD2 and OD5 animals born to sma-3(e491) mutants. t Heat response assay for OD2 and OD5 animals born to sma-2(e502) mutants. u Model of MAE-mediated offspring adult size shaping. Dots in box plots represent worm numbers, and dots in bar plots indicate biological replicates. The data are presented as the means ± SEM with bar plots (q, s, t), box plots (b–d, g, h, k, l, o) or scatter dot plots (e, f, i, j, m, n). In the box plots, the centerline is the median, the box range shows the 25th–75th percentiles, and the whiskers indicate the minimum–maximum values. The bar plots (q, s, t), box plots (b–d, g, h, k, l, o) and scatter dot plots (e, f, i, j, m, n) were analyzed by unpaired t-test. Biological replicates: 4 (b), 3 (c–r), 5 (s, t).