Fig. 4.

A, Inhibitors of MLC phosphorylation modulate recruitment of vinculin and talin to adhesive structures on micropatterned substrates. Fibroblasts were cultured on micropatterned substrates for 16 hours and treated with Y-27632 (50μM), H-7 (500μM), and HA-1077 (50μM) for 30 minutes. Controls received fresh media without inhibitors. The cells were then fixed, extracted, and stained for vinculin, talin, or actin. B, Inhibition of MLC phosphorylation reduces focal adhesion formation. Fibroblasts were cultured overnight on glass substrates coated with 10 μg/ml FN and treated with indicated condition 30 minutes before IRM imaging. C, Inhibitors of contractility modulate recruitment of vinculin and talin to focal adhesions. Western blotting for vinculin and talin recruited to focal adhesions as analyzed by wet-cleaving method. D, relative amounts of localized vinculin and talin were analyzed by Western blotting (mean ± standard deviation, n = 3 from two independent experiments). *p<0.05 relative to no treatment controls which received fresh media without inhibitors.