CK1-γ/gish regulates dynamic apical Rab11 trafficking. (A and A′) Endogenous Rab11 in apical confocal projections of 30–32-h APF pupal wings (Fig. S4 B, diagram). actin-Gal4 FLP-out clones expressing UAS-gish^IR (blue) show an absence of Rab11 foci (green and monochrome in A and A′) at the trichome base (rhodamine-phalloidin, red; yellow arrowheads indicate Rab11 foci at the base of initiating prehairs). (A′′) Rab11 in subapical projections of the same gish^IR clones from A and A′. (B–B′′) In 32–34-h APF wings, Rab11 puncta (see insets) were absent within trichomes in UAS-gish^IR clones (blue; B and B′), whereas subapical regions appeared normal (B’’). (C and C′) actin-Gal4 FLP-out clones of UAS-mwh^IR (blue) exhibit no effect on Rab11 localization (green and monochrome in C and C′). (D–D′′) Z sections of wings from B–B′′ (yellow lines indicate the gishIR clone border, which is blue in D). (E, left) Quantification of the percentage of cells with Rab11 puncta in multiple and single gish^IR mutant trichomes and multiple mwh^IR trichomes. (right) Rab11 detection in wild-type (nonclone [nc]) tissue is included to assess antibody/assay sensitivity. Error bars indicate SDs; unpaired t tests were performed on three independent animals (***, P < 0.001). (F–M) Time-lapse images of live notum epithelia highlighted with pnr-Gal4 expressed UAS-mCD8-RFP to mark membranes and UAS-YFP-Rab11^WT labeling recycling endosomal structures. All images show nota oriented with anterior to the left. (F) 36-h APF notum displays YFP-Rab11^WT localization to the base and within small prehairs (red). (G) The YFP-Rab11^WT pattern is lost upon UAS-gish^IR coexpression. (H) 38-h APF notum shows YFP-Rab11^WT (green) localization along the length of the prehair (red). (H′–J) Time-lapse images of YFP-Rab11^WT dynamics (yellow arrowheads mark initial position shown in H′ and serve as a reference in I and J). (K) 38-h APF notum coexpressing UAS-gish^IR displays diffuse YFP-Rab11^WT localization and rare trafficking in prehair (red). (K′–M) Time-lapse images tracking remaining YFP-Rab11^WT dynamics in prehair (magenta arrowheads mark trafficking shown in K′ and serve as a reference in L and M). Also see Videos 1, 2, 3, and 4. Yellow lines mark clone borders. Bars: (A–D) 10 µm; (F–H, and K) 5 µm.