The RB, SB, and PH domains regulate distinct aspects of Rho-kinase localization. (A) Drosophila Rho-kinase (Rok) contains an N-terminal kinase domain (aa 81–349), central coiled-coil domain (CC; aa 448–1,046), Shroom-binding domain (SB; aa 834–938), Rho-binding domain (RB; aa 966–1,046), and a C-terminal pleckstrin homology domain (PH; aa 1,134–1,391). (B) Domains required for Rho-kinase cortical, junctional, and planar polarized localization. (C and D) Planar polarized enrichment of Venus:Rok^K116A proteins at AP cell boundaries (oriented at 75–90° with respect to the AP axis) relative to DV cell boundaries (oriented at 0–15°). Δ denotes deletion of a domain. Dotted line at 1.0 indicates no polarity, and top dotted line indicates wild-type polarity. (E–P) Images of Venus:Rok^K116A proteins in stage 7 wild-type embryos. Full-length Rok^K116A (FL Rok; E and M), Rok ΔPH (F), Rok ΔRB (G and N), Rok ΔCC,SB (H and P), Rok ΔSB (I), Rok PH (J), Rok RB:PH (K and O), and Rok SB:RB:PH (L) are shown. (E–L) Anterior is left, and ventral is down. (M–P) Cross sections, with apical up. Rok ΔRB, Rok ΔCC,SB, Rok ΔSB, Rok PH, and RB:PH were less planar polarized than full-length Rok^K116A (P < 0.003). Rok ΔPH was slightly less planar polarized (P = 0.013). SB:RB:PH planar polarity was similar to full-length Rok^K116A (P = 0.87). A single value was obtained for each image by averaging 100–200 edges/image; 4–15 images in 4–8 embryos were analyzed/genotype. Means ± SEM between images are shown. Bars, 10 µm.