Sbf and Rab35 work as a ratchet to ensure efficient apical constriction. (A) Time-lapse images of individual cells in control, Sbf shRNA, and Rab35 shRNA embryos during apical constriction. Expansion and constriction rates are color-coded (see color bar on the right). Red indicates constriction and green expansion. Central, black-dotted cells show expansion periods in Sbf- and Rab35-disrupted embryos that are not present in control. (B–D) Automated step detection of periods of active cell area change (shaded area) in control (B), Sbf shRNA (C), and Rab35 shRNA embryos (D). (E and F) Duration (E) and frequency (F) of active steps in control, Sbf shRNA, and Rab35 shRNA background. (G) Quantification of the fraction of negative steps followed by positive steps in control, Sbf shRNA, and Rab35 shRNA embryos. (H) Quantification of active steps that cause area change. Bracket marks active but expending steps. (E–H) n = 835 steps from 167 cells (control), 1,090 steps from 237 cells (Sbf shRNA), and 1,109 steps from 223 cells (Rab35 shRNA). Scale bar in A represents 5 μm. Error bars indicate standard errors. Statistical significance was calculated using a Student’s t test. ***, P < 0.0005. In A, embryos are oriented with anterior up and posterior down.