FIGURE 4:

Inhibition of NM2 elongates microvilli and limits actin turnover. SIM MaxIP images of representative phalloidin-stained (F-actin) Ls174T-W4 cells fixed after (A) 60 min exposure to 20 μM DMSO vehicle control, (B) 10 min exposure to 20 μM blebbistatin, or (C) 60 min exposure to 20 μM blebbistatin; signals are inverted to facilitate visualization of dim structures. (D) Quantification of microvillar length from Ls174T-W4 cells fixed after 10, 30, and 60 min in 20 µM blebbistatin; each data point is a single microvillus, minimum of 10 cells per condition, at least 10 microvilli per cell. Ordinary one-way ANOVA with Dunnett’s multiple comparisons test; ****, p < 0.0001 vs. DMSO control. (E) Image montage of the brush border of a Ls174T-W4 cell expressing EGFP-UtrCH (magenta) and Halo-NM2C labeled with JF585 (green), imaged for 22.5 min after the addition of blebbistatin using spinning-disk confocal microscopy; 2.5 min interval between frames. (F) Image montage of FRAP analysis of Ls174T-W4 cells expressing mNeon-Green β-actin in the absence (top row) or presence (bottom row) of 20 µM blebbistatin for 15 min before imaging.