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Logo of nihpaAbout Author manuscriptsSubmit a manuscriptHHS Public Access; Author Manuscript; Accepted for publication in peer reviewed journal;
 
From:
Nat Mater. Author manuscript; available in PMC 2010 July 1.
Published in final edited form as:
Nat Mater. 2010 January; 9(1): 82–88.
Published online 2009 October 18. doi: 10.1038/nmat2563

Figure 3

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Stress-induced ES cell spreading depends on myosin II activity, Src, Cdc42, but not on Rac activity

a, Summarized data after drug treatments were compared with those of untreated cells (n=5 cells). Control = cell areas before stress application. Inhibiting myosin II ATPase with Blebbistatin (50 μM for 30min; n=7 cells), inhibiting myosin light chain kinase with ML7 (25 μM for 20min; n=5 cells), inhibiting ROCK with Y27632 (50 μM for 20min; n=5 cells), or inhibiting Src activity with PP1 (10 μM for 1hr; n=5 cells), all prevented stress-induced cell spreading, i.e., no significant changes in cell areas between 0 and 10 min and between 0 and 20 min (p>0.05). For inhibiting Rac with NSC23766 (100 μM for 1hr; n=5 cells), there were significant changes in cell areas (p<0.006 and p<0.0009) between 0 and 10 min and between 0 and 20 min. Latrunculin A (0.1 μg/ml for 30 min) (n=10 cells) to disrupt F-actin also prevented stress-induced spreading. Mean±s.e. b, Cdc42 is necessary for stress-induced spreading in mES cells. Western blots of Cdc42 in mES cells under different conditions. Lane 1, non-target shRNA control; Lane 2–4, different constructs to knockdown Cdc42. An independent experiment showed similar results. c, Corresponding changes in cell areas after stress application after Cdc42 knockdown (17.5 Pa at 0.3 Hz). n=9, 8, 9, 8 cells for Lane 1–4 respectively; mean±s.e. (for Lane 1, p<8.68×10−7 and p<2.66×10−6 comparing between 0 and 5 min, 0 and 10 min; there were no significant changes (p>0.05) for Lane 2 through Lane 4). Note that cdc42 knockdown correlated strongly with abolishment of stress-induced spreading response, suggesting that Cdc42 is critical in stress-induced protrusion and spreading.

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