Figure 2. Y155 is required for the correct localization of RLC to actomyosin filaments and adhesion elongation.

(A) Images of fibronectin-bound, CHO.K1 cells transfected with the indicated GFP-tagged versions of RLC. Bars=10 μm. Pictures are representative of >500 cells examined in three independent experiments.

(B) Quantification of the filamentous localization of the RLC mutants shown in (A). Boxes represent the 25^th and 75^th percentile of three independent experiments in which >500 cells were analyzed. p<0.01 between wild type/Y142F and the other conditions; also between Y155E and the other conditions.

(C-D) Quantification of the number of adhesions per μm² (C) and total percentage of the area of the cell that corresponds to focal adhesions, that is, adhesive area (D). Boxes represent the 25^th and 75^th percentile and whiskers correspond to the 10^th and 90^th percentile of three independent experiments in which n ≥ 20 cells (>2000 adhesions) were analyzed. Throughout the study, thick line represents the mean and thin line the median. There are no statistically significant differences among conditions.

(E) Images of fibronectin-bound, CHO.K1 cells transfected with the indicated GFP-tagged versions of RLC. After 2h of adhesion, cells were starved overnight and then stimulated with IGF for 3h. Bars=10 μm. Pictures are representative of >200 cells from three independent experiments.

(F-G) Quantification of the number of adhesions per μm² (F) and adhesive area (G) in the conditions represented in (E). n ≥ 20 cells from three separate experiments (>2000 adhesions per condition). The significances of the relevant comparisons between conditions are shown on the top right corner of each panel. WT stands for wild type throughout the study.