Fig. 10.

Expression of endodermal marker genes. a Fluorescent in situ hybridization of two endodermal marker genes, Pdu-Nk2.1 and Pdu-Otx and b quantification of the effect of activation/inhibition of Wnt/β-catenin pathway on their expression by assignment of all individuals to phenotypic classes. The expression of these genes is typical for neuroectoderm in early larvae but similar to Pdu-Tcf, they obtain new expression domains in the gut later in development while still retaining their neurospecific expression. Pdu-Nk2.1 (top row, green) is expressed in both midgut and hindgut under normal circumstances, and the same pattern is observed in the presence of any of the tested inhibitors or the activator. This is consistent with the theory of midgut to hindgut conversion upon Wnt/β-catenin signalling inhibition since the gene is normally expressed in both tissues. Pdu-Otx is typical for mandibular cells, the entire midgut, whereas it is absent from most of the hindgut except for the very posterior cells which form the sides of the anus. Its midgut expression disappears in the presence of the stronger inhibitor (IWR-1-endo), while a normal pattern is observed with the milder JW55 inhibition (which might just point to unsuccessful inhibition on this batch). Neural and mandibular expression is preserved under all conditions, although it is somewhat lower after Wnt/β-catenin inhibition given the overall smaller size of mandibles. Interestingly, the cells surrounding the anus lose Pdu-Otx expression also in the presence of either CHIR99021 activator of Wnt/β-catenin signalling