Figure 1

Polycystin-1 levels correlate with cytoskeleton protrusions formation in wound-healing assay and microtubules stabilization. (A) MDCK^Zeo (clone F6) and MDCK^PKD1Zeo (clone 68) cells were subjected to wound healing, allowed to migrate for one hour and stained with α-tubulin antibodies (white/red), FITC-phalloidin (white/green) and DAPI staining (blue). Full images and zoom-in of the boxed areas show that MDCK^PKD1Zeo cells have both actin filaments and microtubules elongated towards the wound. Bar: 20 μm. (B) Western blot analysis of PC-1 levels using LRR antibody in fibroblasts Pkd1 ^+/+ and Pkd1 ^−/−, and in control MDCK^Zeo (clone F2, F6) and PKD1-overexpressing MDCK^PKD1Zeo (clone 36, 68, G3) epithelial cells. (C) Western blot analysis using PC-1 LRR antibody of mIMCD cells: two clones isolated from shScrambled-infected (M3 and M4) and two shPkd1-infected (C12 and C16). (D) Pkd1 ^+/+ and Pkd1 ^−/− fibroblasts, as well as mIMCD^shCtrl and mIMCD^shPkd1, were subjected to wound, allowed to migrate for one hour and stained with DAPI (blue) and FITC-phalloidin (white) or α-tubulin antibodies (white). Images show defective protrusions in the two cytoskeletons in Pkd1 ^−/− and mIMCD^shPkd1 cells, compared to their respective controls. Bar: 40 μm. (E) Quantification of nocodazole resistance assay in MDCK cells, revealing that MDCK^PKD1Zeo cells have more persistent microtubules compared to MDCK^Zeo cells. The graph is representative of three independent experiments; averages and SD are shown. Statistical analysis: ANOVA; *p < 0.05, **p < 0.01, relative to both control clones (F2 and F6). (F) Quantification of nocodazole resistance assay in fibroblasts, revealing that Pkd1 ^−/− cells have less persistent microtubules compared to Pkd1 ^+/+ cells. The graph is representative of three independent experiments; averages and SD are shown. Statistical analysis: T-test; ****p < 0.0001.