Pk2 KD in the Xenopus NE attenuates AJ remodeling. (A) Experimental scheme. Pk2 MO was injected with GFP reporter RNA into one dorsal-animal blastomere (green) at the four- to eight-cell stage. RFP reporter RNA was injected into the other dorsal-animal blastomere (red) as a control. The superficial layer of the posterior NE near the brain‒spinal cord border was imaged. (B) Representative images of the control (Scarlet-UtrCH) and Pk2 MO (Lifeact-GFP) NEs from Video 1. Imaging was initiated at stage 15 with 5-min intervals and a duration of 3 h 40 min. Arrowheads indicate the midline. A: anterior. P: posterior. (C) Frequency distribution histogram of the apical domain size in the posterior NE in control and Pk2 MO embryos. Stage 15–16 embryos were stained with phalloidin. Five embryos per group were used. The total number of cells was as follows: n = 1,098 (control) and n = 1,168 (Pk2 MO). (D–G) Representative control (D and E) and Pk2 KD (F and G) images of stage 15–16 NEs from Video 2. The rectangular areas in D and F are enlarged in E and G. The colored dots represent cell tracking. The paired inward white arrows show shrinking AJs before T1 transition. The white circles represent T1 junctions. The white double arrows show the elongation of newly formed AJs after the T1 transition. The yellow circle represents four-cell junctions after cytokinesis. The yellow double arrow represents newly formed AJs after cytokinesis. (H) Frequency of T1 transition was compared between the control (blue) and Pk2 MO (red) sides of NEs in the same embryos. The dots represent the average frequency of T1 transition in individual embryos paired by a line. Five embryos per group were used. (I and J) Rates of AJ elongation (I) and AJ shrinkage on the control and Pk2-KD sides of NEs. A total of 312 AJs (control) and 285 AJs (Pk2 KD) were randomly selected from three embryos. The bold and thin lines represent the medians and quartiles, respectively. (K) Representative still images of Pk1-KD NEs from Video 5. Imaging was initiated at stages 15–16 with 5-min intervals and a duration of 1 h 45 min. A: anterior. P: posterior. Statistical significance was determined as follows: the Mann‒Whitney test was used to compare the mean ranks of the apical domain size (C). Paired Student’s t test (two-sided) was used to compare the means of the cell intercalation rates. Data distribution was assumed to be normal, but this was not formally tested (H). The Mann‒Whitney test was used to compare the mean ranks of AJ elongation (I) and shrinkage (J). *P < 0.05, **P < 0.01. n.s., not significant. Scale bars: 20 μm.