Panel A shows fluorescence microscopy images of Green Fluorescent Protein-membrane in Dimethyl sulfoxide and Golgicide A treated cells at 1 hour and 4 hours, with cell diameter measurements indicated. Panel B shows a superplot of Cell adhesion area (square micrometers), with the x-axis labeled Dimethyl sulfoxide and Golgicide A at 1 hour and 4 hours, and the y-axis labeled Cell adhesion area (square micrometers). Panel C shows fluorescence microscopy images of Active Beta 1 (9EG7) in Dimethyl sulfoxide and CRT0066101 treated cells cultured on Fibronectin for 1 hour and 4 hours. Panel D shows a superplot of Active Beta 1 signal per cell (arbitrary units), with the x-axis labeled 1 hour, Dimethyl sulfoxide, 1 hour, CRT0066101, 4 hours, Dimethyl sulfoxide, and 4 hours, CRT0066101, and the y-axis labeled Active Beta 1 signal per cell (arbitrary units). Panel E shows a superplot of Focal adhesion area per cell (square micrometers), with the x-axis labeled 1 hour, Dimethyl sulfoxide, 1 hour, CRT0066101, 4 hours, Dimethyl sulfoxide, and 4 hours, CRT0066101, and the y-axis labeled Focal adhesion area per cell (square micrometers). Panel F shows fluorescence microscopy images of Acetylated Tubulin, Total tubulin, and merged images in Dimethyl sulfoxide and CRT0066101 treated cells. Panel G shows a superplot of Acetylated/Total Tubulin fluorescence intensity (arbitrary units), with the x-axis labeled 4 hour, Dimethyl sulfoxide and 4 hour, CRT0066101, and the y-axis labeled Acetylated/Total Tubulin fluorescence intensity (arbitrary units).
Golgi-derived export is necessary for cell spreading and mechanoadaptation. (A) Representative confocal microscopy images of fixed HeLa cells transfected with the GFP-mem plasmid, subjected or not to a treatment with 10 µM Golgicide A. Scale bars, 10 µm; representative cell length scales shown. (B) SuperPlots showing individual cell measurements (small, light-colored symbols; N∼10 per biological replicate) and the mean value for each independent biological replicate (larger, black-outlined circles; n = 3). Each color represents a different experimental replicate. A repeated-measures two-way ANOVA test using Fisher’s LSD test was performed. (C) Confocal microscopy images of HeLa cells pre-seeded on FN-coated glass coverslips for 30 min and then treated with CRT0066101 (5 µM) or DMSO for 30 min (total time 1 h) or 210 min (total time 4 h), fixed, and processed for immunofluorescence microscopy against endogenous active β1 integrin (9EG7 antibody) and paxillin. Scale bar, 10 µm. (D and E) SuperPlots showing quantification of the fluorescence intensity signal of active β1 integrin per cell (D) and of the FA area per cell (E) from cells in C. A repeated-measures two-way ANOVA test was performed using Tukey’s post hoc multiple comparison test (N = 3 replicates, n∼10 cells per replicate). (F) Confocal microscopy images of HeLa cells pre-seeded on FN-coated glass coverslips for 30 min and then treated with CRT0066101 (5 µM) or DMSO for 210 min (total time 4 h), fixed and processed for immunofluorescence microscopy against acetylated and total α-tubulin. Scale bar, 10 µm. (G) SuperPlot showing quantification of the acetylated vs. total α-tubulin intensity signal per cell, from cells in F. A repeated-measures two-way ANOVA test was performed using Tukey’s post hoc multiple comparison test (N = 3 replicates, n∼10 cells per replicate). A two-sided parametric ratio paired t test was used. The P value is indicated in the plot.