aPKC organizes cytoskeleton to promote AJ remodeling, but apical localization of aPKC does not require Rab11. (A-C) Temporospatial expression pattern of aPKC in zebrafish lens. (A) aPKC displays non-polarized distribution in lens epidermal cells when Crb2a was not expressed at 24 hpf. (B) Coupling with Crb2a expression in epithelia, aPKC was accumulated to the apical domains at 28 hpf. (C) aPKC was restricted at the apical domains and colocalized with Crb2a in epithelial cells at 36 hpf. In contrast to Crb2a, aPKC was also expressed in lens mesenchymal cells (broken arrows) and primary fiber cells (arrowheads). Arrows show the apical aggregation of aPKC, Crb2a and actin in lens epithelial cells. Broken arrows show the lens mesenchymal cells. Arrowheads show the primary fiber cells. a (anterior) and p (posterior) in panel A show the lens orientation. (D) Apical zonula adherens disappeared in lens epithelial cells in aPKCλ mutants at 36 hpf. a (apical) and b (basal) show the orientation of lens epithelial cells. The black boxed region in D shows the apical zone between two epithelial cells. (E) Tricellular adherens (white arrowhead) and lateral punctum adherens (black arrow) between lens epidermal cells in aPKCλ mutants at 22 hpf. (F) In aPKCλ mutants, aPKC staining had disappeared (F), actin lost its apical enrichment. Arrows show the apical regions of lens epithelial cells. (G) In aPKCλ mutants, Crb2a and Rab11 lost their apical distribution in most epithelial cells at 36 hpf. Arrows show the apical colocalization of Crb2a and Rab11. (H) In rab11a/rab11ba dKO mutants, Crb2a lost its apical aggregation at 36 hpf. In contrast, aPKC and actin were still enriched at the apical domains in lens epithelial cells. Arrows show the apical regions of lens epithelial cells. (I) Quantification of apical/basal relative fluorescence intensity of aPKC, actin, Crb2a and Rab11 in WT and rab11a/rab11ba dKO mutants shown in H. aPKC displayed normal apical enrichment in rab11a/rab11ba dKO mutant lens epithelial cells; however, apical enrichment of actin, Crb2a and Rab11 was significantly decreased although actin reduction was mild (n=20 lenses from 10 embryos). (J) The apical localization of aPKC and actin displayed a heterogeneity in different lens epithelial cells in nok mutants at 36 hpf. Broken arrows show that aPKC and actin lost their apical enrichment in the anterior epithelial cells. Arrows show that aPKC and actin were still enriched in the apical regions in lateral epithelial cells. (K) Quantification of apical/basal relative fluorescence intensity of aPKC, actin, Crb2a and Rab11 in nok mutants shown in J and Fig. 3C. The apical enrichment of actin was associated with aPKC, whereas the apical enrichment of Rab11 was associated with Crb2a. Crb2a and aPKC showed concurrent apical localization in anterior epithelia, but not in lateral epithelial cells (n=20 lenses from 10 embryos). Scale bars: 10 μm (A-C,F-I), 400 nm (D,E). **P<0.01, ***P<0.001.
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