Focal Adhesion Size Controls Tension-Dependent Recruitment of$\alpha-Smooth$Muscle Actin to Stress Fibers

Expression of$\alpha-smooth$muscle actin ($\alpha-SMA$) renders fibroblasts highly contractile and hallmarks myofibroblast differentiation. We identify$\alpha-SMA$as a mechanosensitive protein that is recruited to stress fibers under high tension. Generation of this threshold tension requires the anchoring of stress fibers at sites of$8-30-\mu m-long$"supermature" focal adhesions (suFAs), which exert a stress approximately fourfold higher ($\sim 12 nN/\mu m^2$) on micropatterned deformable substrates than$2-6-\mu m-long$classical FAs. Inhibition of suFA formation by growing myofibro blasts on substrates with a compliance of$\leq 11 kPa$and on rigid micropattems of$6-\mu m-long$classical FA islets confines$\alpha-SMA$to the cytosol. Reincorporation of$\alpha-SAAA$into stress fibers is established by stretching$6-\mu m-long$classical FAs to$8.1-\mu m-long suFA islets$on extendable membranes; the same stretch producing$5.4-\mu m-long$classical FAs from initially$4-\mu m-long$islets is without effect. We propose that the different molecular composition and higher phosphorylation of FAs on supermature islets, compared with FAs on classical islets, accounts for higher stress resistance.