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. 2017 Nov 6;216(11):3729–3744. doi: 10.1083/jcb.201611061

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© 2017 Zilberman et al.

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Figure 2. — cdc-42 is not required for epidermal cell polarity or junction maturation. (A and B) PAR-6 staining of control embryos (cdc-42 mutants rescued with ha-zf1-cdc-42) and cdc-42(MZ) embryos. Red arrows indicate PAR-6 enrichment at the apical domain in epidermis; white arrows indicate enrichment at the apical domain in intestine. Red lines show t where the intensity profile shown in E was taken. (C and D) PAR-6-GFP and PAR-6ΔCRIB-GFP imaged live in par-6(MZ) embryos. (E) Intensity profile of PAR-6 in control (representative of n = 24) and in cdc-42(MZ) (representative of n = 20) epidermal cells (taken from micrographs shown in A and B). (F) Quantification of apical membrane to cytoplasm intensity ratio of PAR-6 immunostaining in the epidermis of control (n = 24) and cdc-42(MZ) (n = 20) embryos. Individual data points from two independent experiments were pooled. (G) Quantification of apical membrane to cytoplasm intensity ratio in the epidermis of PAR-6-GFP (n = 13) and PAR-6ΔCRIB-GFP (n = 14) in live par-6(MZ) embryos. Individual data points from two independent experiments were pooled. (H–I) Frames from time-lapse movies of control and cdc-42(MZ) embryos expressing HMR-1-GFP, showing polarization and junction maturation of dorsal epidermal cells. Insets show a side view of lateral epidermal cells, in which apical enrichment of HMR-1-GFP to junctions is observed by 50 min. t = 0 is defined as the initiation of polarization, when HMR-1-GFP puncta are first detected in the intestine. See Videos 2 and 3. (J) Quantification of junctional enrichment of HMR-1-GFP in epidermal cells at t = 50 min (control, n = 9 embryos; cdc-42(MZ), n = 12 embryos). (K and L) Immunostaining of junctional HMP-1 and basolateral LET-413 in control and cdc-42(MZ) embryos. Insets below are of boxed region. Line and error bars in graphs (F, G, and J) indicate mean ± SD. P-values were calculated using a Mann-Whitney U test. *, P ≤ 0.05; ***, P ≤ 0.001. NS, P > 0.05. Bars, 5 µm.