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. 2023 Feb 6;24(4):e55069. doi: 10.15252/embr.202255069

Figure 5. TRPV2 activity controls the calpain‐dependent mechanical maturation of focal adhesions.

Figure 5

  1. Quantification of vinculin‐stained focal adhesion sites per cell. Confocal images taken on low confluency melanoma cells seeded on fibronectin‐coated coverslips were analyzed using the imageJ software to count the number of vinculin clusters per cell. The total number of cells counted for each cell line is indicated in the boxes. Data are presented as box and whiskers plots (Boxes extend from the 25th to 75th percentiles, whiskers from min to max, the horizontal line in each box is plotted at the median). **P = 0.0066, the Mann–Whitney test (501mel); ****P < 0.0001 the Kruskal–Wallis test followed by Dunn's multiple comparisons tests (WM266.4 and 451Lu).
  2. Comparison of calpain activity in control (GFP) or overexpressing TRPV2 (GFP‐TRPV2) 501mel cells, and in control (shRNA Ctrl) or TRPV2‐silenced (shRNA V2‐1, ‐2) WM266.4 cells or 451Lu cells. Bar graphs show mean normalized calpain activity ± SEM (n = 3 biological replicates). For 501mel the Mann–Whitney test was used for statistical analysis (*P = 0.05); For WM266.4&451Lu one‐way ANOVA followed by Dunnett's multiple comparisons tests was used (**P < 0.01; See Appendix Table S2 for exact P‐values).
  3. Representative immunoblots showing the full‐length (230 kDa; Black Arrows) and the calpain‐mediated degradation product (190 kDa; Gray Arrows) of talin after overexpression or repression of TRPV2, in the corresponding cell lines. Tubulin was used as a loading control.
  4. Densitometric analysis of the cleaved‐talin ratios from three independent experiments (as described in C) normalized to control. Data are presented as mean ± SEM. For 501mel the Mann–Whitney test was used for statistical analysis (ns P = 0.2); For WM266.4&451Lu one‐way ANOVA followed by Dunnett's multiple comparisons tests was used (*P < 0.05; ***P < 0.001; See Appendix Table S2 for exact P‐values).

Source data are available online for this figure.