ANI-1 tethers IT-RHO-1 to linear structures after Formin ^CYK-1 depletion but does not affect IT-RHO-1 levels and dynamics. (A) Single z-plane images of the cell cortex of one-cell C. elegans embryos expressing IT-RHO-1 treated with the indicated RNAi conditions. A zoom-in of the equatorial region for the indicated time points is shown on the right. Selected images are reproduced in Fig. S3 A to illustrate the quantification of IT-RHO-1 structures. Scale bars are 5 µm. (B) Normalized cortical fluorescence intensity of IT-RHO-1 from the anterior (0%) to the posterior (100%) pole for indicated RNAi conditions. (C) Mean normalized fluorescence intensity of IT-RHO-1 at the cell equator 180 s after NEBD for indicated RNAi conditions and dots represent data points of individual embryos. (D) Mean number of linear and nonlinear structures at the cell equator for the indicated RNAi conditions 180 s after NEBD. (E and F) IT-RHO-1 was bleached in a small region immediately after furrow ingression in control and ani-1(RNAi) treated embryos (E, left). Plotted are the mean fluorescence recovery in the bleached region (F) and recovery half time and the percentage of recovery for each embryo (E). For all, time in seconds (s) after NEBD is indicated, error bars are SEM, and n = number of embryos analyzed. P values were calculated using the two-tailed Student’s t test or Mann–Whitney U test and are n.s. P > 0.05 and *P < 0.05.