![Crb dynamics during apical contractions, and NB apical area, myosin, and Ecad dynamics in crb-RNAi embryos. (A) Left dot plot showing the fraction of apical perimeter increase (expansions [exp.]) or decrease (contractions [cont.]) during early and late ingression. ns, P = 0.14; ****, P (from left to right) = 3 × 10−9, 3.17 × 10−6, and 4.6 × 10−12. Right dot plot showing the fraction of apical Crb gain or loss during apical oscillations in early and late ingression. ns, P = 0.78 and 0.19; ****, P = 8.57 × 10−10; ***, P = 3 × 10−4. Note a significant increase in relative contractions during late ingression, matching with higher reductions in apical Crb. Column height, median; error bars, IQR; 385 contractions and 302 expansions analyzed in 58 NBs from 12 embryos expressing endo-Crb::GFP. (B) Ventral views of late stage 8 embryos injected with H2O (top, four embryos) or crb-dsRNA (bottom, seven embryos) stained for Crb and Armadillo (Arm). Notice the strong uniform reduction in Crb levels in crb-RNAi embryos. Scale bars, 50 μm. (C) Stills from time-lapse movies of ingressing NBs (arrowheads) expressing Ecad (endo-Ecad::GFP; green) and myosin::mCherry (sqh-Sqh::mCherry; red) in crb-RNAi and embryos (H2O injected control is shown in Fig. 2 A). T = 0 min, onset of ingression. Scale bars, 5 μm. (D) Amplitude and duration of individual contractions/expansions during ingression in control versus crb-RNAi slowest and fastest NBs. Median amplitudes for control/slow/fast crb-RNAi of contractions, 3.04/2.35/6.95 μm2/min; **, P = 7.6 × 10−3; ****, P = 1.5 × 10−16 and 2.0 × 10−17; and expansions, 1.42/1.32/2.4 μm2/min; ns, P = 0.23; ****, P = 3.3 × 10−5 and 5.4 × 10−6. Median duration for control/slow/fast crb-RNAi of contractions, 46.6/38.3/53.2 s, ns, P = 0.10; *, P = 0.046; **, P = 5.4 × 10−3; and expansions, 20.6/21.2/19.6 s, ns, P = 0.09, 0.16, 0.06 (KS test); n = 176–551 events per condition. (E) Representative plots showing the rate of medial myosin change (a.u./min) throughout ingression in a control NB (blue), and crb-RNAi slow and fast NB (red). Positive rates indicate myosin assembly; negative rates indicate disassembly. T = 0 min, onset of ingression. (F) Medial myosin assembly and disassembly rates during ingression in control NBs, crb-RNAi slow and fast NBs. N values as in Fig. 3 C. Median assembly rates for control/slow/fast crb-RNAi: 0.06/0.09/0.09 a.u./min; ***, P = 5.0 × 10−4; *, P = 0.01; median disassembly rates for control/slow/fast crb-RNAi: 0.05/0.09/0.06 a.u./min; ***, P = 1 × 10−4; ns, P = 0.07; n = 151–429 events per condition. (G) Mean levels of Ecad (endo-Ecad::GFP) in control NBs and slow/fast crb-RNAi NBs. N values as in Fig. 3 C. ****, P = 1.8 × 10−10, 8.3 × 10−79, 1.7 × 10−49 (two-tailed T test). Means + SD.](https://cdn.rupress.org/rup/content_public/journal/jcb/221/7/10.1083_jcb.202108076/5/jcb_202108076_figs1.png?Expires=1771551873&Signature=B0HVylkXHdCDQ0cThkl49B3zpSwABQWeOkagTDlR5VkikCxGhFiq3P03WUHFoiDjRaWpnil2IfSF1WfqCEOJBSEXN7ImUuS8cmU4RO5O-VURzfkcFr6hlt6HdpRgTcznbGDAUHNgh8s9cBqOR02nkZv9EMx2RfOwFLD4Iv2IxXAEOKNesGTE5Kgn7PhnCMoogsNZ7a4Fe3Cz9R7e3Geod68gx1I0IfJNDPe~xXJqZ7Q3Y9u6sLEVu0ZruGU2VnLOjxSCnTDeA4BLEwYjyC-0w90cXrX7ebNetzRTm5bw984dW2yad1eWN2id6Y2L4vfxojuduARz9IyFg5~Vt0DUiA__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Crb dynamics during apical contractions, and NB apical area, myosin, and Ecad dynamics in crb-RNAi embryos. (A) Left dot plot showing the fraction of apical perimeter increase (expansions [exp.]) or decrease (contractions [cont.]) during early and late ingression. ns, P = 0.14; ****, P (from left to right) = 3 × 10−9, 3.17 × 10−6, and 4.6 × 10−12. Right dot plot showing the fraction of apical Crb gain or loss during apical oscillations in early and late ingression. ns, P = 0.78 and 0.19; ****, P = 8.57 × 10−10; ***, P = 3 × 10−4. Note a significant increase in relative contractions during late ingression, matching with higher reductions in apical Crb. Column height, median; error bars, IQR; 385 contractions and 302 expansions analyzed in 58 NBs from 12 embryos expressing endo-Crb::GFP. (B) Ventral views of late stage 8 embryos injected with H2O (top, four embryos) or crb-dsRNA (bottom, seven embryos) stained for Crb and Armadillo (Arm). Notice the strong uniform reduction in Crb levels in crb-RNAi embryos. Scale bars, 50 μm. (C) Stills from time-lapse movies of ingressing NBs (arrowheads) expressing Ecad (endo-Ecad::GFP; green) and myosin::mCherry (sqh-Sqh::mCherry; red) in crb-RNAi and embryos (H2O injected control is shown in Fig. 2 A). T = 0 min, onset of ingression. Scale bars, 5 μm. (D) Amplitude and duration of individual contractions/expansions during ingression in control versus crb-RNAi slowest and fastest NBs. Median amplitudes for control/slow/fast crb-RNAi of contractions, 3.04/2.35/6.95 μm2/min; **, P = 7.6 × 10−3; ****, P = 1.5 × 10−16 and 2.0 × 10−17; and expansions, 1.42/1.32/2.4 μm2/min; ns, P = 0.23; ****, P = 3.3 × 10−5 and 5.4 × 10−6. Median duration for control/slow/fast crb-RNAi of contractions, 46.6/38.3/53.2 s, ns, P = 0.10; *, P = 0.046; **, P = 5.4 × 10−3; and expansions, 20.6/21.2/19.6 s, ns, P = 0.09, 0.16, 0.06 (KS test); n = 176–551 events per condition. (E) Representative plots showing the rate of medial myosin change (a.u./min) throughout ingression in a control NB (blue), and crb-RNAi slow and fast NB (red). Positive rates indicate myosin assembly; negative rates indicate disassembly. T = 0 min, onset of ingression. (F) Medial myosin assembly and disassembly rates during ingression in control NBs, crb-RNAi slow and fast NBs. N values as in Fig. 3 C. Median assembly rates for control/slow/fast crb-RNAi: 0.06/0.09/0.09 a.u./min; ***, P = 5.0 × 10−4; *, P = 0.01; median disassembly rates for control/slow/fast crb-RNAi: 0.05/0.09/0.06 a.u./min; ***, P = 1 × 10−4; ns, P = 0.07; n = 151–429 events per condition. (G) Mean levels of Ecad (endo-Ecad::GFP) in control NBs and slow/fast crb-RNAi NBs. N values as in Fig. 3 C. ****, P = 1.8 × 10−10, 8.3 × 10−79, 1.7 × 10−49 (two-tailed T test). Means + SD.
