FIGURE 6.

FHL2 modulates focal adhesions and Rho activation. A, endogenous FHL2 localizes to focal adhesions, along some stress fibers and the cytoplasm. Control or FHL2 shRNA MTLn3 cells were cultured on fibronectin-coated coverslips and stained with rhodamine phalloidin (actin) and immunolabeled for FHL2 and vinculin. Scale bar = 20 μm. B, average focal adhesion number in MTLn3 control or FHL2 shRNA cells. n > 50 cells per experiment. C, average focal adhesion area in control or FHL2 shRNA cells; n > 40 focal adhesions per experiment; *, p = 0.0190. D, FHL2 mediates RhoA activation. Control and FHL2 shRNA cells were serum-starved 24 h and treated with 5 nm EGF for 5 min. Lysates were incubated with Rhotekin beads to pull down active Rho. RhoA levels were detected by immunoblotting. E, quantification of active RhoA pulled down by densitometry with normalization to control. Active RhoA was reduced in FHL2 deficient cells. n = 8. *, p = 0.0314. All experiments were performed in triplicate. Error bars represent S.E.