Sequential Signalling at the Fng Boundary Figure 7. Importance of Notch activation in dorsal or ventral boundary cells for wing development. (A–D) Expression of UAS psn in psn mutant wing imaginal discs exclusively in dorsal cells with apGal4. Arrowhead points to the inner ring-like expression domain of Wg in the proximal wing anlage. (A, B) Expression of Ap (green) and Wg (red) in PsnC1 mutant discs. Note that the boundary of Ap expressing and non-expressing cells is irregular because the compartment boundary fails to form. In addition the diameter of the inner ring-like domain of Wg is strongly reduced and the expression domain along the D/V boundary is lost (C, D) A similar Phenotype is observed if UAS psn is expressed exclusively in dorsal cells with apGal4. Thus restoring a functional Notch pathway in only dorsal cells is not sufficient to rescue expression of Wg along the D/V boundary. (E) In contrast supplying psn activity simultaneously in dorsal and ventral cells by expressing UAS Psn with dppGal4 restores Wg expression (arrow in E). (F– H) Expression of UAS mib1 in mib12/mib13 mutant wing discs with apGal4. (F) Expression of Wg in a mib mutant control disc. The diameter of the residual ring-like domain of Wg expression is dramatically reduced and the expression domain along the D/V boundary is lost. (G, H) Expression of UAS mib1 exclusively in dorsal cells with apGal4 is sufficient to restore expression of Wg along the D/V boundary (arrow) and to allow wing development to occur. (I–N) Expression of UAS Su(H)-RNAi during wing development with apGal4 (I–M) and sdGal4 (N). Expression with apGal4 results in the formation of a nearly normal looking wing anlage. However, the comparison with the expression of Ap expression revealed that the Wg domain is restricted to VBCs in early (I, J) and late (K–M) third instar discs. (N) Expression on both half of the wing results in abolishment of expression of Wg along the D/V boundary. doi:10.1371/journal.pone.0049007.g007 expression of H causes a restriction of expression of Wg to VBCs, but not its abolishment. Thus, the Dl/Ser loop is probably mainly required for the later occurring patterning of the future wing margin, but not for the establishment of the wing primordium. A further addition is that the basic expression of Dl is independent of Ser signalling. This is indicated by the observation Figure 8. Clonal analysis of the allele fng13. (A–E) A clone crossing the D/V boundary interrupts expression of Wg along the boundary in early (A–C) and late (D, E) third instar discs. The arrowheads point to the clone area, which is labelled by loss of the GFP marker. doi:10.1371/journal.pone.0049007.g008 PLOS ONE | www.plosone.org 12 November 2012 | Volume 7 | Issue 11 | e49007