Rab11 vesicle transport requires microtubules and dynein. (A) An example of Rab11 vesicle movement (yellow circle) along the microtubules (Jupiter-GFP). Scale bar, 1 μm. (B) Acute disruption of microtubules via colchicine injection results in immediate inhibition of Rab11 vesicle transport. Magenta arrowheads: residual microtubules. Yellow circles: region where microtubules were completely disrupted. N = 3 embryos. Scale bars, 2 μm. (C) En face view of the constricting domain shortly (<2 min) after on-stage injection of water or colchicine. Injection of colchicine shortly before but not after the onset of gastrulation (N = 5 embryos for each condition) results in reduced Rab11 vesicle accumulation compared to water injected controls (N = 4 embryos). Scale bar, 10 μm. Bottom panels: zoom-in views. Scale bar, 2 μm. (D–F) Quantification of Rab11 vesicle density after water or colchicine injection within a 3.5 μm-stack from apical surface (D) or in a single z plane near apical surface (E) generates consistent results (F). The central mark indicates the median, and the bottom and top edges of the box indicate the 25th and 75th percentiles, respectively. The whiskers extend to the most extreme data points not considered outliers. Two-sample, two-tailed, unpaired t test. **, P < 0.01; n.s., P > 0.05. (G) Injection of dynein antibody rapidly abolishes both apical (red) and basal (green) transport of Rab11 vesicles. Trajectories show vesicle movement within a 20-s window at the indicated time after injection. N = 5 embryos for each condition. Scale bars, 2 μm. (H) En face view showing the inhibition of apical accumulation of Rab11 vesicles upon on-stage injection of dynein antibody immediately before the onset of gastrulation. N = 5 and 6 embryos for GST antibody and dynein antibody injection, respectively. Scale bar, 10 μm. Bottom panels: zoom-in views. Scale bar, 2 μm.