Fig. 2. Localization of Dgo during PCP establishment in the eye. Anterior is leftwards and dorsal upwards in all panels except F and I, where proximal is leftwards. (A) Single apical confocal microscopy section showing ommatidial rows 1-7 stained with anti-Dgo (green) and phalloidin (highlighting F-actin, red) to mark the center of each cluster. Morphogenetic furrow (anterior) is on the left of the panel and the equator is at the bottom. (Right panels) Monochrome higher magnifications of single clusters from row 3, row 5 and row 6 (as indicated). Dgo is first detected apical at the membrane in all cells ahead of and posterior to MF (up to row 2). More posteriorly, the initial uniform apical localization changes (row 3) within five cell preclusters, and Dgo is present at higher levels in R3/4 cells within the equatorial-polar (DV) axis. (Row 5) Dgo is maintained at the border between the R3/R4 cells, but it is now detected less in parts of R3 and at higher levels in R4. (Row 6) The R4 specific enrichment is now complete and persists through row 10. Whereas there is no Dgo detected at membranes where R3/R4 abut the R2/R5 pair, Dgo is enriched at the opposing R8, R2/R5 membranes. (B-D) Mosaic analysis of GFP-Dgo to determine the precise localization of Dgo at the R3/R4 cell boundary. GFP-Dgo is in green, anti-DE Cadherin is in red. Right side panels show a schematic of actual clusters on the left. Cells are numbered according to their position; cells shown in white express GFP-Dgo in the given cluster; GFP-Dgo at localization membranes is highlighted with green lines. (B,C) Mosaic clusters in row 3 with either R4 (B) or R3 (C) expressing GFP-Dgo. Dgo localization reflects a horseshoe-like pattern in both R3 and R4 cells. (D,E) Mosaic clusters in rows 5-6, with (again) either R4 (D) or R3 (E) expressing GFP-Dgo. Dgo is now enriched only at the R3 side of the R3/R4 cell boundary and the polar side of R4 (very similar to the localization of Fz). (F) GFP-Dgo (green) clones in pupal wings at 60-80 hours at 18°C. lacZ (blue) marks the cells that express GFP-Dgo; F-actin (phalloidin, red) labels the growing actin hairs; proximal is leftwards. GFP-Dgo localizes to the distal membrane of each cell that expresses it. (G-I) Schematic summary presentation of the Dgo localization patterns in developing eye (G, row 3; H, row 6) and pupal wing (I) cells; again, Dgo shows the same localization pattern as Fz and Dsh.