Rapid Actions of Plasma Membrane Estrogen Receptors Regulate Motility of Mouse Embryonic Stem Cells through a Profilin-1/Cofilin-1-Directed Kinase Signaling Pathway

Long-term estrogen actions are vital for driving cell growth, but more recent evidence suggests that estrogen mediates more rapid cellular effects. However, the function of estradiol-17β (E2)-BSA in mouse embryonic stem cells has not been reported. Therefore, we examined the role of E2-BSA in mouse embryonic stem cell motility and its related signal pathways. E2-BSA (10−8m) significantly increased motility after 24 h incubation and increased filamentous (F)-actin expression; these effects were inhibited by the estrogen receptor antagonist ICI 182,780, indicating that E2-BSA bound membrane estrogen receptors and initiated a signal. E2-BSA increased c-Src and focal adhesion kinase (FAK) phosphorylation, which was attenuated by ICI 182,780. The E2-BSA-induced increase in epidermal growth factor receptor (EGFR) phosphorylation was inhibited by Src inhibitor PP2. As a downstream signal molecule, E2-BSA activated cdc42 and increased formation of a complex with the neural Wiskott-Aldrich syndrome protein (N-WASP)/cdc42/transducer of cdc42-dependent actin assembly-1 (TOCA-1), which was inhibited by FAK small interfering RNA (siRNA) and EGFR inhibitor AG 1478. In addition, E2-BSA increased profilin-1 expression and cofilin-1 phosphorylation, which was blocked by cdc42 siRNA. Subsequently, E2-BSA induced an increase in F-actin expression, and cell motility was inhibited by each signal pathway-related siRNA molecule or inhibitors but not by cofilin-1 siRNA. A combined treatment of cofilin-1 siRNA and E2-BSA increased F-actin expression and cell motility more than that of E2-BSA alone. These data demonstrate that E2-BSA stimulated motility by interacting with profilin-1/cofilin-1 and F-actin through FAK- and c-Src/EGFR transactivation-dependent N-WASP/cdc42/TOCA-1 complex.