Fig. 2

Dynamics and Compositions of the PAAs and OAAs

(A) Diagrams illustrate the principle of GFP-assisted sagittal serial microscopy for visualizing proteins at the opposing apical surfaces: the opposing apical surfaces at the midline region (S^M), which contain the OAAs, must be flanked by two spatial references—PAA regions of the left and right halves of the tissue (S^L and S^R). Green represents transiently expressed GFP in some cells. The bottom panels are images collected with this technique: N-Cad and β-catenin were detected at the PAA regions as circumferential belts (S^L and S^R, arrows) as well as in the OAA regions as punctate foci (S^M; arrowheads). By contrast, ZO-1 and actin were detected in the PAA regions (S^L and S^R, arrows) but not in the OAA regions (S^M, arrowheads).

(B) Diagrams illustrate the strategy to visualize transiently expressed N-Cad^wt-GFP or N-Cad^m117-GFP at 14-ss. Note that N-Cad^wt-GFP enriched on the opposing apical surfaces (illustrated by the drawing on the right).

(C) The PAAs (arrows) and the opposing apical surfaces (arrowheads indicating the OAAs) survived the Triton X-100 extraction of the anti-GFP immunogold electron microscopic procedure, whereas the lateral membranes were dissolved (basal of asterisks).

(D) The magnification of the boxed region in C revealed the labeling of N-Cad-GFP with 5-nm gold particles at the PAAs (arrows, 72% of the PAAs were labeled with gold particles) and OAAs (arrowhead). Insets are magnifications of boxed regions.

(E) Crb2a juxtaposed with ZO-1 sites at the apical ends (26-ss).

(F) The table summarizes the developmental and compositional changes of the OAAs and PAAs. Crb*, Crb-based adhesion; NaKATPase*, Na⁺/K⁺-ATPase-based adhesion; +, presence; -, absence. Also see Figure S2.