Figure 2.

Daple Localizes to the Fzd/Dvl Side of the Cell Cortex and Regulates Microtubule Dynamics

(A) Whole-mount staining of the LV wall with antibodies against Dvl1 (green) and Daple (red). The enlarged image of the indicated box area is shown below.

(B) Whole-mount staining of the LV wall with antibodies against Fzd6 (green), Dvl1 (red), Daple (white/blue), and tyrosinated tubulin (white/magenta). Each panel shows the apical surface of the ependymal cells. Arrowheads indicate sites of co-localization of Fzd6, Dvl1, and Daple with tyrosinated tubulin at the cell cortex.

(C) Confocal microscopy images of the apical surface of Daple^+/+ or Daple^−/− ependymal cells on the LV wall stained with antibodies against Fzd6 (green), tyrosinated tubulin (red), and γ-tubulin or β-catenin (white). Arrowheads indicate the accumulation of Fzd6 or tyrosinated tubulin at the cell cortex. Dashed lines indicate BB clusters.

(D) Accumulation of tyrosinated tubulin at the Fzd6 side of the cell cortex was quantified for Daple^+/+ and Daple^−/− ependymal cells. The mean intensity of tyrosinated tubulin at the Fzd6 area of the cell cortex was normalized against the mean intensity of tyrosinated tubulin for the entire cell surface area. Normalized intensities are plotted as mean values ± SEM for 30 ependymal cells, sampled from three mice in each group.

(E) Kymographs of single microtubules from Daple^+/+ or Daple^−/− ependymal cells. The tips of EMTB-EGFP at the cell edge were monitored by fluorescence time-lapse microscopy imaging, see also Videos S1 and S2.

(F) Quantification of the state of microtubule dynamics in Daple^+/+ or Daple^−/− ependymal cells. Shrinking, growing, and pausing (pause defined as > 0.1 μm/2 s) states of 30 microtubules were measured over 2.55 min for each group. Values are represented in Table S1.