Figure 5.
Fbp17 is required to maintain cortical integrity. (A) WT, fbp17−, and GFP-Fbp17/fbp17− (rescue) cells grown for 60 h in shaken suspension were fixed and stained with DAPI to visualize the nuclei. (B) Quantification of nuclei in cells. n, number of cells analyzed. (C) Top: Trajectories of randomly migrating cells (n = 43 for WT and 46 for fbp17−). Bottom: Summary of motility parameters. (D and E) Velocity and Euclidean distance of cells are shown in C. At least 13 cells were quantified per experiment (each experiment shown in a different color); mean ± SEM. (F) Top: Trajectories of cells migrating under 2% agarose along a folic acid gradient (n = 61 for WT and 62 for fbp17−). Bottom: Summary of chemotaxis parameters; FMI, forward migration index. (G and H) Directness and FMI of cells shown in F. At least 17 cells were quantified per experiment (each experiment shown in a different color); mean ± SEM. (I) Localization of GFP-ABD in WT and fbp17− cells. (J) Quantification of the cortex-to-cytoplasm fluorescent intensity ratios of GFP-ABD. (K) WT and fbp17− cells were fixed and stained with Alexa Fluor 555-labeled phalloidin. (L) Quantification of the fluorescence intensity of cortical phalloidin. (M) Projection length (Lp) of WT and fbp17− cells determined by micropipette aspiration using a constant pressure of 500 Pa for 5 min. (N) Quantitative analysis of the projection lengths of probed cells. For C and F, data were from three independent experiments (the average of each biological replicate was used to calculate the mean and SEM). For J, L, and N, data were from three independent experiments; the scatter plots show data points with mean ± SEM; n, number of cells analyzed. Scale bars, 5 μm.

Fbp17 is required to maintain cortical integrity. (A) WT, fbp17−, and GFP-Fbp17/fbp17− (rescue) cells grown for 60 h in shaken suspension were fixed and stained with DAPI to visualize the nuclei. (B) Quantification of nuclei in cells. n, number of cells analyzed. (C) Top: Trajectories of randomly migrating cells (n = 43 for WT and 46 for fbp17−). Bottom: Summary of motility parameters. (D and E) Velocity and Euclidean distance of cells are shown in C. At least 13 cells were quantified per experiment (each experiment shown in a different color); mean ± SEM. (F) Top: Trajectories of cells migrating under 2% agarose along a folic acid gradient (n = 61 for WT and 62 for fbp17−). Bottom: Summary of chemotaxis parameters; FMI, forward migration index. (G and H) Directness and FMI of cells shown in F. At least 17 cells were quantified per experiment (each experiment shown in a different color); mean ± SEM. (I) Localization of GFP-ABD in WT and fbp17− cells. (J) Quantification of the cortex-to-cytoplasm fluorescent intensity ratios of GFP-ABD. (K) WT and fbp17− cells were fixed and stained with Alexa Fluor 555-labeled phalloidin. (L) Quantification of the fluorescence intensity of cortical phalloidin. (M) Projection length (Lp) of WT and fbp17− cells determined by micropipette aspiration using a constant pressure of 500 Pa for 5 min. (N) Quantitative analysis of the projection lengths of probed cells. For C and F, data were from three independent experiments (the average of each biological replicate was used to calculate the mean and SEM). For J, L, and N, data were from three independent experiments; the scatter plots show data points with mean ± SEM; n, number of cells analyzed. Scale bars, 5 μm.

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