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. 2012 May 15;23(10):1846–1859. doi: 10.1091/mbc.E11-12-1057

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© 2012 Hoffman et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

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FIGURE 3: — Zyxin responds independent of signaling molecule p130Cas. (A) Western immunoblot for p130Cas in WT and Cas-null fibroblasts. (B, C) Fibroblasts from p130Cas^−/− mice were stained for F-actin (phalloidin) on unstretched (ø) membranes or after uniaxial cyclic stretch (1 h, 15%, 0.5 Hz). Stretch direction is in the horizontal plane (double-headed arrow; scale, 40 μm). (D) SFTI analysis of the stretch-induced actin thickening in Cas^−/− cells, ****p < 0.0001. (E, F) Immunofluorescence microscopy of zyxin in fibroblasts from p130Cas^−/− mice on unstretched (ø) and stretched membranes. (G) Immunoblot analysis of zyxin in cell lysates from unstretched (ø) and stretched (+) membranes (WT, lanes 1 and 2; Cas^−/−, lanes 3 and 4) showed the stretch-induced zyxin shift seen in WT cells persists in p130Cas^−/− cells. (H) Fibroblasts on unstretched (ø) membranes or after uniaxial cyclic stretch (5 min, 15%, 0.5 Hz) with dimethyl sulfoxide control or with ERK activation inhibitor PD98059 (100 μM) were immunoblotted for total ERK1/2 (p44/p42), P-ERK, P-zyxin, and β-actin. (I) WT fibroblasts unstretched (ø) and stretched for 5, 30, and 60 min, electrophoresed, and immunoblotted for P-ERK, total zyxin, P-zyxin, and β-actin indicate maximal ERK phosphorylation at 5 min, followed by zyxin phosphorylation.