Mispolarized UNC-40 colocalizes with F-actin in the absence of UNC-6. Anterior is left; ventral is down. (A–C) All animals were examined at the P6.p two-cell stage. Shown are UNC-40::GFP and UNC-40(ΔFN4/5)::GFP (left), F-actin (middle), and an overlay (right). Magnification (below) and colocalization quantification graphs (far right) show extensive UNC-40 and F-actin overlapping localization in all cases (arrowheads; colocalization was measured along the yellow line in the insets from the outside to the inside of the cell, fluorescent intensity was plotted in arbitrary units, and distances are given in pixels). (D) A schematic diagram illustrates the structural domains of the wild-type UNC-40 protein and the UNC-40(ΔFN4/5) protein, an engineered form of UNC-40 lacking the putative UNC-6 binding site in the fourth and fifth FNIII domains. (E) In unc-40 mutants, UNC-40(ΔFN4/5)::GFP colocalized with F-actin in the apical and lateral membranes of the AC (arrowheads). Bars, 5 µm.