Wb and LanA play distinct roles in midgut MET. (a–f) Midgut cells in stage 15 wb (a and b) and LanA (d and e) mutant embryos; plots of the average fluorescence intensity (c and f, represented as mean gray value) of Baz, E-Cad, and βPS in stage 15 midgut cells mutant for wb (c) and LanA (f) measured along the apical (A) to basal (B) axis of a cell (represented in b and e by red line). Each n represents the average of 10 cells measured in one embryo (black dotted lines). n = 6 per condition, the mean for each condition is plotted in red. (a–c) In wb mutants, midgut cells fail to form an epithelial monolayer, and Baz (b1), E-Cad (b2), and βPS (b3) are found delocalized throughout the cells. (c) Accordingly, the apical peaks of Baz and E-cad are lost in wb mutants. There is just a small peak of βPS apically and a low broad peak basally, which correlates with diffuse βPS throughout the mesoderm layer. (d–f) In LanA mutants, midgut cells form a disorganized-looking monolayer of columnar or wedge-shaped cells, with βPS integrins localized to the apical and basal sides of the cells (e3), although quantification of levels reveals the peaks of βPS to be half that in wild type (f). In contrast, Baz (e1) and E-Cad (e2) are no longer restricted to the apical side of the cells (d and e) and are found in lower levels throughout the lateral membranes (d and e). Plots of the average fluorescence intensity (f) reveal the apical peak of Baz to be half that of wild-type cells and much broader, and a lower apical peak of E-Cad. White dashed lines in a and b indicate the apical side of the midgut, and yellow lines, the basal. Red boxes in a and d depict the area of the midgut epithelium shown in b and e. Confocal images are oriented with the anterior to the left and posterior to the right. Scale bars, 10 µm.