Daple regulates the planar polarized distribution of MTs in a cell-autonomous manner. (A) Representative immunofluorescence images of keratin8 (green) and Odf2 (blue) in WT and Daple-KO MTECs grown at the ALI for 12–14 d. The Odf2 signal indicates MCCs. (B) Representative immunofluorescence images of actin (red) and Odf2 (blue) in WT and Daple-KO MTECs grown on the ALI for 12–14 d. The Odf2 signal indicates MCCs. (C and D) Statistical analyses of keratin8 (C) and actin (D) expression in MCCs of WT and Daple-KO tracheas (n = 8–18 cells). Significant differences were evaluated using two-tailed Mann-Whitney U tests; n.s., P ≥ 0.05. (E) Representative images of α-tubulin (green) and Daple (magenta) in WT and Daple-KO MTECs grown at the ALI for 12–14 d. The Odf2 signal indicates MCCs. The intensity of the α-tubulin signal is represented by the color map, in which red indicates high intensity and blue indicates low intensity. (F) Schematic illustration of the co-culture method of MTECs prepared from the tracheas of WT and Daple-KO mice. (G) Representative immunofluorescence images of Daple (magenta), ZO-1 (blue), and tyrosinated tubulin (green) in co-cultured MTECs of WT (orange) and Daple-KO (blue) cells grown at the ALI for 12 d. The intensity of the tyrosinated tubulin signal is illustrated by the color map (red indicates high intensity; blue indicates low intensity). Tyr-tubulin, tyrosinated tubulin. (H and I) Immunofluorescence image (H) and intensity profiles for Tyr-tubulin (I) in MCCs of WT and Daple-KO co-cultured MTECs. Scale bars represent 5 µm. A.U., arbitrary units.