Expression of a mutant GMFβ cannot rescue GMFβ depletion. (A) Sequence alignment of human GMFβ (Hs) with S. cerevisiae (Sc), Drosophila (Dm), and C. elegans (Ce) GMF homologues. The mutated site is indicated by the red bracket. (B) Localization of mutant GMFβ-GFP in unsynchronized cells. IF of Arp2/3 and F-actin is shown. Bar, 10 µm. (C) Ratio of mutant GMFβ-GFP to soluble tRFP. The boxed region is enlarged on the right. Bars, 25 µm. (D) Cell area quantified from micrographs for mutant GMFβ OE. (E) Cell area quantified from micrographs for GMFβ KD cells rescued with mutant GMFβ (KDR). (F) Retraction rate in micrometers per minute for mutant GMFβ OE. (G) Retraction rate in micrometers per minute for mutant GMFβ KDR. (H) The percentage of cell edge positive for high Arp2/3 for mutant GMFβ OE. (I) The percentage of cell edge positive for high Arp2/3 for mutant GMFβ KDR. For all graphs, error bars represent 10th–90th percentile. Kruskal-Wallis multiple comparison testing was performed, and significance was measured with a Dunn’s post-test. ***, P < 0.001; **, P < 0.01; *, P < 0.05.