Fig. 3: Oncogene expression induces epithelial tumourigenesis in the accessory gland.

a–l GFP-Ras^V12 expressing clones display hallmarks of cancer. a Visualisation of the tumour cell membranes with a myristoylated form of Tomato (Myr-Tomato) indicates that the cells conserve their typical binucleation. b–g Staining reveals that tumour-like GFP-Ras^V12 expressing clones lost expression of the epithelial markers. Epithelial markers visualised by staining are: (b) Fasciclin III (FasIII), (d) Coracle, (f) E-Cadherin (E-Cad). h Phosphorylated Src (P-Src) is detected at in GFP-Ras^V12 expressing clones, a feature of mesenchymal cells and/or transformed epithelial cells. i–l Tracheal system is revealed by Matrix Metalloprotease 1 (MMP1, i, l) or Gasp (j, k) staining. i Presence of tumour-like GFP-Ras^V12 expressing clones is correlated to strong hypertrophy of tracheal system. j–k Gasp staining reveals trachea network associated with tumour-like GFP- Ras^V12 expressing clones. k Zoom of the boxed section in (j); white arrowhead points at a strong gasp expression in a low diameter trachea, a feature of developing trachea. l MMP1 staining reveals an arborescence of trachea into the tumour and from accessory gland tissue (white arrowhead). DAPI (blue) reveals nuclei in (a–k). Representative images in (b–i, l) from three or more experiments, in (j)–(k) from two experiments and in (a) from one. Scale bars: 50 μm.