Fig. 7: Loss of intestinal DAF-2 triggered gene expression changes in other tissues through cross-tissue DAF-2 to DAF-16 signaling.
a Isolation of neurons, hypodermis, body wall muscle (BWM), and intestine cells from intestinal DAF-2 AID worms for RNA-seq. Left panel, tissue-specific transgenic reporters in the intestinal DAF-2 AID strain used to isolate tissue-specific cells. Neurons, labeled by rgef-1p::NuGFP; hypodermis, labeled by dpy-7p::NLSSV40::GFP; BWM, labeled by myo-3p::NuGFP; intestine, labeled by ges-1p:: NuGFP. Right panel, representative images of tissue-specific cells isolated by FACS for RNA-seq. A similar pattern of expression was observed in two independent experiments. b GSEA analysis of up-regulated and down-regulated GO terms enriched in each isolated tissue. GO terms with q-value < 0.01 in at least two tissues are shown. For multiple comparisons, adjustments were made with Benjamini–Hochberg (BH) method. c Degrading DAF-2 from the intestine induces DAF-16 nuclear accumulation in the hypodermis at the L2 larval stage. A similar pattern of expression was observed in three independent experiments. d Five DEGs selected to verify the cross-tissue effect of intestinal IIS on non-intestinal tissues by mCherry transgenic reporters in the intestinal DAF-2 AID strain. Representative images of each reporter are shown in the bottom panel. A similar pattern of expression was observed in three independent experiments. e Schematic of the combination of tissue-specific GFP nanobody-mediated ZIF-1 system and tissue-specific AID system to simultaneously degrade DAF-2 and DAF-16 in two different tissues, the intestine and non-intestinal tissues (neurons or hypodermis). f Lifespan phenotypes following simultaneous degradation of intestinal DAF-2 and non-intestinal DAF-16. Degrading intestinal DAF-2 by GFP nanobody-mediated ZIF-1 system extended lifespan by 51.8% (top panel, p < 0.0001). Degrading DAF-16 in the hypodermis (bottom panel, p = 0.003), but not in the neurons (middle panel, p = 0.337), moderately but significantly decreased the lifespan of the worms in which the intestinal DAF-2 level was reduced. p-values are calculated by log-rank tests. See survival statistics in Supplementary Data 1. g Motif enrichment analysis of transcriptional binding sites among 1-kb promoter sequence of the intestinal, hypodermal, and neuronal DEGs. See statistics in Supplementary Data 5. Source data are provided as a Source Data file.
