Subcellular localization of Smash in embryos. (A) Smash colocalizes with Baz at the ZA of the epidermis. Epidermis of an embryo at stage 13 stained for Smash (A, green in Merge), Baz (red in Merge), Dlg (blue in Merge), and DAPI (cyan in Merge). Arrowheads point to colocalization of Smash and Baz at the ZA. (B) In zygotic bazEH747 mutant embryos, maternal Baz is lost from individual cells at stage 11 (arrows). In these cells, Smash is also lost at the ZA (Smash and Merge panels). (C) Smash (C, green in Merge) is enriched at myotendinous junctions of an embryo at stage 16 and colocalizes there with α-actinin (red in Merge) and βPS integrin (blue in Merge). (D and E) Smash is planar polarized and enriched at A/P junctions. Embryos at stage 8 were stained for Smash (D, green in Merge) and Baz (D, middle and red in Merge) or for Smash (E, red in Merge) and Sqh-GFP (E, middle and green in Merge). In D (Merge panel), some A/P junctions are marked by green arrowheads and some D/V junctions are marked by red arrowheads. (F) Quantification of PCP of Smash, Sqh, and Baz in stage 8 embryos. ***, P < 0.001. n = 200 cell contacts analyzed for each protein. (G) Smash is enriched at tricellular junctions (G, arrows) to a higher extent than Baz (middle) and E-cadherin (right). (H) Quantification of the enrichment at tricellular junctions. ****, P < 0.0001; ***, P < 0.001; **, P < 0.01; n = 6 embryos. At least 10 cells were analyzed per embryo. In D–G, anterior is to the left and dorsal is up. Images in A–C and G are single optical sections taken with an Airyscan detector. Images in D and E are maximum-intensity projections of three optical sections 1 µm apart taken from the apical region of the epidermis with an Airyscan detector. Bars, 20 µm. Error bars in F and H show mean ± SEM.
