RTKN-1-CT is a determinant for actin integrity during basolateral recycling. (A and A′) The actin-binding potential of the GST-RBD-PH was measured using the cosedimentation assay. Multiple concentrations were deployed to make a curve to estimate interaction (n = 3 independent experiments). (B and B′) Confocal images showing localization of RTKN-1-GFP and RBD-PH-GFP in intestinal cells. Box-and-whisker plots (n = 18 cells): 10–90th percentile; dots, outliers; red midline, median of WT; boundaries, quartiles. ***, P < 0.001 by Mann-Whitney test. (C) GST-RBD or GST-RBD-PH did not affect GST-UNC-60A–mediated actin depolymerization; t_1/2 has been indicated. (D and D′) Confocal images showing the colocalization between endogenous UNC-60A and Utrophin-CH–labeled actin structures. The white arrowheads indicate structures labeled by both GFP and mCherry. OE, overexpression. Mander's coefficients were calculated; error bars are 95% CIs (n = 12 animals). (E–G′) Confocal images showing the distribution of GFP-Utrophin-CH, hTAC-GFP, and hTfR-GFP in intestinal cells. Box-and-whisker plots (n = 18 cells): 10–90th percentile; dots, outliers; red midline, median of WT; boundaries, quartiles. ***, P < 0.001 by one-way ANOVA followed by Dunn’s post hoc multiple comparison test. The black asterisks in the image panels indicate intestinal lumen. Scale bars, 10 µm. MC, mCherry; P, pellet; S, supernatant.