Figure 3.
Establishment of planar polarized apical MTs requires Daple in tracheal MCCs. (A) Representative super-resolution images of α-tubulin as a marker of cilia and apical MTs in different z-slices (A', A'', and A''') in tracheal MCCs of WT MTECs grown on the ALI for 12–14 d, with illustrations showing z-slices in tracheal MCCs. Arrows display MCCs. (B) Comparison between a super-resolution fluorescence (IF) image for α-tubulin (upper panels) and a UHVEMT image in which MTs are reconstituted in 3D (green; lower panels) in tracheal MCCs of isolated WT MTECs grown at the ALI for 12–14 d. (C) Immunofluorescence images of α-tubulin and Fzd6 in WT and Daple-KO MTECs grown at the ALI for 12–14 d and color map of intensities of fluorescent signals for α-tubulin in which red indicates high intensity and blue indicates low intensity. (D and E) Linear ROI setting (D) and intensity profiles along the ROI of immunofluorescence for α-tubulin (E) in MCCs of WT (orange) and Daple-KO MTECs (blue). (F) Statistical analysis of the slope coefficient (Coef.) acquired from the intensity profiles (E) along the linear ROI setting in MCCs of WT (orange; n = 8 cells) and Daple-KO MTECs (blue; n = 8 cells). Two-tailed Mann-Whitney U test; *, P < 0.05. (G) Representative UHVEMT images in which the MTs are reconstituted in 3D (green) in WT and Daple-KO adult tracheas. The dashed line shows the cell membrane. The double arrows indicate the direction of the mounted trachea in B and G. Scale bars represent 5 µm in A, C, and D, and 500 nm in B and G.

Establishment of planar polarized apical MTs requires Daple in tracheal MCCs. (A) Representative super-resolution images of α-tubulin as a marker of cilia and apical MTs in different z-slices (A', A'', and A''') in tracheal MCCs of WT MTECs grown on the ALI for 12–14 d, with illustrations showing z-slices in tracheal MCCs. Arrows display MCCs. (B) Comparison between a super-resolution fluorescence (IF) image for α-tubulin (upper panels) and a UHVEMT image in which MTs are reconstituted in 3D (green; lower panels) in tracheal MCCs of isolated WT MTECs grown at the ALI for 12–14 d. (C) Immunofluorescence images of α-tubulin and Fzd6 in WT and Daple-KO MTECs grown at the ALI for 12–14 d and color map of intensities of fluorescent signals for α-tubulin in which red indicates high intensity and blue indicates low intensity. (D and E) Linear ROI setting (D) and intensity profiles along the ROI of immunofluorescence for α-tubulin (E) in MCCs of WT (orange) and Daple-KO MTECs (blue). (F) Statistical analysis of the slope coefficient (Coef.) acquired from the intensity profiles (E) along the linear ROI setting in MCCs of WT (orange; n = 8 cells) and Daple-KO MTECs (blue; n = 8 cells). Two-tailed Mann-Whitney U test; *, P < 0.05. (G) Representative UHVEMT images in which the MTs are reconstituted in 3D (green) in WT and Daple-KO adult tracheas. The dashed line shows the cell membrane. The double arrows indicate the direction of the mounted trachea in B and G. Scale bars represent 5 µm in A, C, and D, and 500 nm in B and G.

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