Fig. 4: Hi-1 acts as a salivary (labial) gland-specific gene and is highly expressed at early and late larval stages of M. sexta.

a Relative transcript abundance of Hi-1 throughout different developmental stages. Expression levels of each stage are relative to the expression level in the embryo. Different letters indicate significant differences (p < 0.05) between developmental stages (One-way ANOVA analysis with log2 transformed data followed by Tukey HSD post-hoc analysis, F_8,35 = 152.3, p < 0.0001, n = 5 biologically independent samples). b (3Z):(2E)-hexenal isomerase activity of oral secretions from larvae at different developmental stages. Equal amounts of total protein (0.5 µg) per sample (n = 3 biologically independent samples) or buffer control (n = 8 biologically independent samples) were incubated with Z-3-hexenal (1 mM), and a SPME-guided in vitro assay was performed. The proportion of E-2-hexenal (in percentages) emitted from the total aldehyde headspace (Z-3-hexenal + E-2-hexenal) was calculated. Different letters indicate significant differences (p < 0.05) between tissues (One-way ANOVA followed by Tukey HSD post-hoc analysis, F_5,17 = 38.21, p < 0.0001). c Representative extracted ion chromatograms (ion 69) as output of the SPME-guided assay. Green highlight of the E-2-hexenal peak indicates a clear conversion of Z-3-hexenal to E-2-hexenal. d Relative transcript abundance of Hi-1 in different organ tissues including brain plus ganglia (BR), foregut (FG), midgut (MG), hindgut (HG), Malpighian tubules (MT), fat body (FB) and salivary glands (SG) of fourth instar larvae (expression relative to BR). Different letters indicate significant differences (p < 0.05) between tissues (One-way ANOVA analysis with log2 transformed data followed by Tukey HSD post-hoc analysis, F_6,14 = 10.05, p = 0.0002, n = 3 biologically independent samples). Error bars are presented as mean values ± SEM.