Fig. 7: AMPK triggers DAF-16-dependent innate immune signaling pathway to defense against Bt infection.
a Venn diagram showing the overlap in genes activated by nematicidal Bt and the target genes of DAF-16 in C. elegans. b qRT-PCR analysis of the expression of DAF-16 target genes in wild-type N2, aak-2(ok524), and daf-16(mu861) mutant worms after Bt infection. Error bars denote the Standard deviation. Data represent three biological replicates of representative results from three independent experiments with the same trend. c DAF-16 translocation observed during the no-RNAi and aak-2 RNAi transgenic worms TJ356(Isdaf-16::gfp) fed with BMB171/Cry5Ba or BMB171/pHT304 for 2 h, respectively. The representative images for each treatment are shown. Scale bar represents 20 µm. N = 3 independent experiments. d The quantification of DAF-16 distribution after Bt infection (CK: BMB171/pHT304; NBt: BMB171/Cry5Ba). Worms with cytosolic (Cyt), intermediate (Int), or nuclear (Nuc) DAF-16 location were counted, and the percentages of each pattern of DAF-16 nuclear translocation were calculated. N = 3 independent experiments containing at least 30 worms. e The survival assay of the N2 worms after relevant gene knockdowns by RNAi under BMB171/Cry5Ba treatment for 4 days. N = 3 independent experiments contain three replication of at least 30 worms. Data points represent the mean values of three independent replicates, error bars denote the SD. f The expression level of Psod::GFP was observed using a reporter strain CF1553 (Psod::GFP) during the aak-2 RNAi, daf-16 RNAi, or no-RNAi after Bt infection. Scale bar represents 20 µm. Representative images of worms in each condition are shown. N = 3 independent experiments. g Quantification of Psod::GFP expression levels calculated by the fluorescence intensity of individual worms. Worms were treated for 4 h and then photographed. N = 3 independent experiments containing at least 20 worms. Each point represents a worm. The p-value was determined by a Student’s t-test (Welch’s correction for unequal variances) in (b), (e), and (g); and by a two-sided Fisher exact test in (a) and (d) (*: vs CK; #: vs NBt). ***p < 0.001, **p < 0.01, *p < 0.05 and ns indicates no significant difference.
