Fig. 3. Ectopic expression of Dystroglycan interferes with epithelial cell polarity. (A) A schematic drawing of the UAS-DG construct and a Western blot to detect overexpression of UAS-DG driven by a Gal4 driver. (B) UAS-DGcyto contains the cytoplasmic and the transmembrane domain of DG linked to a FLAG-tag. Western blot analysis of UAS-DGcyto (20kDa) driven by daGal4 using anti-Flag antibody. (C) Drawing of an embryo at stage 13 to show the location of the salivary gland. The apical surface of the salivary epithelium is indicated by green line (in the center of the salivary gland). (D) A Nomarski image of a wild-type embryo at stage 12-13 stained with the Crb antibody, which shows apical staining (arrow). (E) Stage 13 embryo derived from da-GAL4 x UAS-DG cross. No Crb staining in the salivary gland can be detected (arrow). Notice that Crb staining in the pharynx is normal (arrowhead). (F) An enlarged drawing of an embryonic salivary gland at stage 13. The apical surface of the epithelial cells is shown in green. (G) A confocal image of the wild-type salivary gland around stage 13 stained with Crb (green) and DG (red) antibodies. Wild-type DG expression is below detection level. (H,H') Salivary gland of stage 13 embryo expressing UAS-DG driven by elav-GAL4. Notice the strong staining of DG (red, H) and the strong reduction of Crb (green, H; white, H') at the apical membrane. (I) Stage 10 follicle cells that overexpress UAS-DG (marked with GFP, green) accumulate high levels of DG protein (red) both in the apical (arrow) and basal surfaces. By contrast, stage 10 wild-type follicle cells express very low levels of DG (arrowhead). (J,J') Apical localization of ßH-Spec (red in J, white in J') is reduced in DG-overexpressing follicle cells (green; arrow). (K,K') Overexpression of DG in follicle cells also causes reduction (arrow) of the apical localization (arrowhead) of Baz (red in K, white in K'). (L) Summary of the localization of apical markers (red line) in the wild-type (i), Dg mutant epithelial cell clones (ii) and cells that overexpress DG (iii). Apical markers are expanded to the basolateral surface of the epithelium in Dg mutant clones, and their apical localization is substantially reduced because of DG overexpression.