Fig. 3: LPWJL volatiles Promote tracheal branching formation, gut oxygenation, and gut growth.
Germ-free embryos were exposed to LPWJL volatiles throughout larval stages (+) or left untreated (−). Third-instar GF larvae were used. a OR42b expression in the enterocytes (open arrowhead) and gut-associated trachea (filled arrowhead) was observed. b LPWJL volatiles enhanced tracheal Btl signaling activation (left: representative image; right: tracheal coverage normalized to untreated larvae, set to 1). n = 3 biologically independent experiments. c Btl signaling activation is dependent on exposure to LPWJL volatiles. Larvae carrying Btl-Gal4 > UAS-GFP were used. To exclude the effect of larval size on tracheal Btl signaling activation, volatile-exposed larvae and non-exposed larvae of similar size were used (left). Relative tracheal coverage of these animals is shown (right). The tracheal coverage of untreated animals was taken arbitrarily as 1. n = 3 biologically independent experiments. LPWJL volatiles induced enterocyte Bnl expression (d) and tracheal Btl expression (e). f Gut-specific Bnl knockdown reduced tracheal branching formation. Relative tracheal coverage was shown and the tracheal coverage of untreated control animals was taken arbitrarily as 1. n = 4 biologically independent experiments. g LPWJL volatiles increased gut oxygenation, measured using Timer, a fluorescence-based oxygen reporter (left: merged oxidized red/unoxidized green Timer image; right: oxidized red/unoxidized green ratio). n = 4 biologically independent experiments. h The gut lengths, from proventriculus to hindgut, of untreated larvae at 144 h AEL were taken arbitrarily as 1. n = 3 biologically independent experiments. i Gut-specific Bnl knockdown abolishes VSF-induced GF larval growth at 144 h AEL. n = 3 biologically independent experiments. Data are mean ± s.e.m., with P values (*p < 0.05, ***p < 0.001, ns not significant) from unpaired two-tailed t-test (b, c, g, h) or two-way ANOVA with Tukey’s multiple comparison test (f, i). The gut is arranged from anterior to posterior, moving from top to bottom in the photo (a, b, d, e, g). Nuclei were stained with DAPI (blue). Scale bar, 50 μm. Genotypes, sample sizes, and statistical analyses are shown in Supplementary Data 1. Source data are provided in the Source Data file.
