Pk2 promotes the elongation of newly formed AJs after cytokinesis. (A and B) Representative snapshot images of new AJ formation after cytokinesis. Xenopus gastrula ectoderm expressing GFP-ZO1 only (A) or with Flag-Pk2 (B). Time-lapse imaging was initiated at stage 11, and t = 0 was set at the time when the contractile ring was closed. (C) Length changes of newly formed AJs in cells expressing only GFP-ZO1 (blue) or with HA-RFP-Pk2 (red). Three embryos per group were used. The total number of newly formed AJs was as follows: n = 38 (control); n = 20 (HA-RFP-Pk2). The maximum value of AJ length after cytokinesis was used for statistical analysis. Unpaired Student’s t test (two-sided). The data distribution was tested for normality using the Kolmogorov–Smirnov normality test. P < 0.0001. (D) Cell proliferation rate in stage 10–11.5 embryos expressing GFP-ZO1 only or GFP-ZO1 with Flag-Pk2. The dots represent the average rate of cell proliferation in individual embryos. The lines represent the means of seven (control) or six (+Pk2) embryos. Unpaired Student’s t test (two-sided). The data distribution was tested for normality using the Kolmogorov–Smirnov normality test. The total number of cells was as follows: n = 875 (control); n = 853 (Flag-Pk2). (E) Quantification of the correlation between AJ length changes and AJ orientation in the control (left) and Pk2 MO (right) gastrula ectoderms. 30 AJs were randomly selected from time-lapse images of three embryos per group. Changes in AJ length were categorized as follows: elongation (blue, ≥0.8 μm/3 min), shrinkage (red, less than or equal to −0.8 μm/3 min), and no change (green, −0.8< and <0.8 μm/3 min). On the basis of orientation, AJs were categorized into AP (≤30° from the AP axis), ML (≤30° from the ML axis), and mid (30–45° from either the AP or ML axes). The two-sample Kolmogorov‒Smirnov test was used on original numerical data of AJ length changes. Distributions were compared between AP and ML junctions. Three embryos per group were used. 10 AJs per embryo were used. The total number of AJ length changes was as follows: n = 1,258 (control) and n = 910 (Pk2 KD). n.s., not significant. Scale bars: 10 μm.