Figure 3.
Crb and the RhoGEF Cysts are required for normal NB ingression dynamics. (A) Mean apical area loss during NB ingression in crb dsRNA injected embryos. T = 0 min, onset of ingression. Means ± SEM. Control: 20 NBs, three embryos, 260 time points spaced by 6 s, crb-RNAi: 38 NBs, five embryos, 176 time points spaced by 6 s. (B) NB ingression speed in crb-RNAi (n = 91 NBs, 12 embryos), cyst-RNAi (21 NBs, 3 embryos), and control embryos (68 NBs, 10 embryos; 19 NBs, 4 embryos, respectively). Median values: −1.5 (H2O injected), −1.6 (crb-RNAi), −1.6 (cyst-RNAi control), −2.6 (cyst-RNAi). KS tests: ns, P = 0.09, **, P = 0.0014; F-tests (to test for differences in variance of ingression speed): ****, P = 1.7 × 10−5; ns, P = 0.23. Bars are IQRs. (C) Mean apical area loss, the 10 fastest, and the 10 slowest ingressing NBs in crb-RNAi (five embryos; control, H2O injected, 20 NBs, three embryos). T = 0 min, time point when the average apical surface of each ingressing cell population was 22 μm2. Means ± SEM. (D and E) Junctional and medial myosin levels (D; ****, P = 2.5 × 10−85; 3.4 × 10−19) and Ecad (E; ****, P = 1.1 × 10−40) in ingressing NBs from crb dsRNA injected and H2O-injected embryos. N values as in B (two-tailed T test). Means ± SD. (F) Cyst is one of several Crb complex (Crb/Sdt) effectors. Cyst stimulates Rho1 to enhance junctional myosin II. Other Crb/Sdt effectors are the Par6/aPKC complex, which undergoes negative feedback regulation with basolateral polarity proteins (Scrib/Dlg/Lgl and Yurt/Cora), and Moesin and βH-Spectrin, which support apical membrane stability (Tepass, 2012; Silver et al., 2019). (G) Mean apical area loss during NB ingression in cyst-RNAi embryos compared to best-match controls. T = 0 min indicates the onset of ingression. Data presented are means ± SEM. N values: control (19 NBs, four embryos, 97 time points spaced by 15 s) versus cyst-RNAi (21 NBs, three embryos, 59 time points spaced by 15 s).

Crb and the RhoGEF Cysts are required for normal NB ingression dynamics. (A) Mean apical area loss during NB ingression in crb dsRNA injected embryos. T = 0 min, onset of ingression. Means ± SEM. Control: 20 NBs, three embryos, 260 time points spaced by 6 s, crb-RNAi: 38 NBs, five embryos, 176 time points spaced by 6 s. (B) NB ingression speed in crb-RNAi (n = 91 NBs, 12 embryos), cyst-RNAi (21 NBs, 3 embryos), and control embryos (68 NBs, 10 embryos; 19 NBs, 4 embryos, respectively). Median values: −1.5 (H2O injected), −1.6 (crb-RNAi), −1.6 (cyst-RNAi control), −2.6 (cyst-RNAi). KS tests: ns, P = 0.09, **, P = 0.0014; F-tests (to test for differences in variance of ingression speed): ****, P = 1.7 × 10−5; ns, P = 0.23. Bars are IQRs. (C) Mean apical area loss, the 10 fastest, and the 10 slowest ingressing NBs in crb-RNAi (five embryos; control, H2O injected, 20 NBs, three embryos). T = 0 min, time point when the average apical surface of each ingressing cell population was 22 μm2. Means ± SEM. (D and E) Junctional and medial myosin levels (D; ****, P = 2.5 × 10−85; 3.4 × 10−19) and Ecad (E; ****, P = 1.1 × 10−40) in ingressing NBs from crb dsRNA injected and H2O-injected embryos. N values as in B (two-tailed T test). Means ± SD. (F) Cyst is one of several Crb complex (Crb/Sdt) effectors. Cyst stimulates Rho1 to enhance junctional myosin II. Other Crb/Sdt effectors are the Par6/aPKC complex, which undergoes negative feedback regulation with basolateral polarity proteins (Scrib/Dlg/Lgl and Yurt/Cora), and Moesin and βH-Spectrin, which support apical membrane stability (Tepass, 2012; Silver et al., 2019). (G) Mean apical area loss during NB ingression in cyst-RNAi embryos compared to best-match controls. T = 0 min indicates the onset of ingression. Data presented are means ± SEM. N values: control (19 NBs, four embryos, 97 time points spaced by 15 s) versus cyst-RNAi (21 NBs, three embryos, 59 time points spaced by 15 s).

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