Fig. 1
NK2R-HA induces ectopic activation of the N signalling pathway. A–B”’ N-HA and NK2R-HA were expressed under control of hh-GAL4, tub-GAL80ts for 14.5 h in the posterior compartment of late third instar larvae wing imaginal discs. Activity of the N signalling pathway was indicated by the N target gene Wg and the activity reporter construct NRE-GFP. The expression of hh-Gal4 is restricted to the right side of the stippled yellow line. The left side is the wild-type control. In contrast to N-HA, NK2R-HA induced a strong activation of NRE-GFP and a slight ectopic expression of Wg (A, A’, B, B’). C N-HA and NK2R-HA western blot analysis of whole larvae lysate. Transgenic N constructs were identified by HA antibody staining. In contrast to N-HA, NK2R-HA showed a strong accumulation of NICD. D Quantification of N activation in wing imaginal discs by fluorescence intensity measurement, shown in Fig. 1A, B. NRE-GFP intensity of the posterior compartment was normalised to the anterior compartment (wt: n = 9 / N-HA: n = 7 / NK2R-HA: n = 11, significance test: one-way ANOVA p ≥ 0.05 (n.s.) / p ≤ 0.05 (*) / p ≤ 0.01 (**) / p ≤ 0.001 (***) / p ≤ 0.0001 (****)). NK2R-HA induced a significantly stronger activation of the N-pathway than N-HA. E Quantification of N activation in an S2 cell luciferase-assay. Activation of the N signalling pathway was indicated by NRE-Fluc. NRE-Fluc fluorescence intensity was normalised to N-HA activity (n = 3 luciferase-assays, significance test: one-way ANOVA p ≥ 0.05 (n.s.) / p ≤ 0.05 (*) / p ≤ 0.01 (**) / p ≤ 0.001 (***) / p ≤ 0.0001 (****)). Basal N activation induced by NK2R-HA was significantly increased