Fig. 2: WWP GOF led to detachment-induced apoptosis.
From: WWP1 gain-of-function drives developmental anoikis through TGFβ pathway during neurodevelopment
A Representative images of HEK293T and hNPCs transfected with control or WWP1 variant constructs co-expressing GFP. Scale bar, 50 μm. B Scheme of live imaging to monitor apoptosis events in hNPCs. The image is created with Biorender.com. Fluorogenic dye is activated upon cleavage by active caspase 3. C Quantification of the percentage of detached cells among GFP+ cells transfected with control or WWP1 variants. n = 3. One-way ANOVA with Tukey’s post hoc test. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. D Quantification of time taken from GFP expression to detachment in hNPCs transfected with control or WWP1 variants. Control, n = 33; WWP1OE, n = 36; WWP1E798V, n = 44; WWP1C890A, n = 36; WWP1E798V C890A, n = 39. Kruskal-Wallis test with Dunn’s post hoc test. **p < 0.01; ****p < 0.0001. E Quantification of the percentage of GFP+CC3+ and GFP+CC3- hNPCs transfected with control or WWP1 variants. n = 3. Two-way ANOVA with Tukey’s post hoc test. The significance of the GFP+CC3+ group is labeled. *p < 0.05; **p < 0.01; ****p < 0.0001. F Quantification of time taken from GFP expression to CC3 signal in hNPCs transfected with control or WWP1 variants. Control, n = 27; WWP1OE, n = 29; WWP1E798V, n = 30; WWP1C890A, n = 28; WWP1E798V C890A, n = 30. Kruskal-Wallis test with Dunn’s post hoc test. *p < 0.05; **p < 0.01; ***p < 0.001. G Quantification of the percentage of hNPCs transfected with control or WWP1 variants categorized by detachment and apoptosis sequence. n = 3. Two-way ANOVA with Tukey’s post hoc test. Significance of the detachment to CC3+ group is labeled. *p < 0.05; ****p < 0.0001. H Representative images of apical junction belt in E15.5 brain electroporated with control or WWP1 variants. Green, GFP; magenta, Phalloidin. White arrows indicate disruption in the apical belt. Scale bar, 50 μm. I Pie chart showing the ratio of disrupted apical belt in E15.5 brain electroporated with control or WWP1 variants. Control, n = 10; WWP1E798V, n = 9; WWP1E798V C890A, n = 7. Bar graphs indicate mean ± SEM.
