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[摘要]：The ability of transforming growth factor beta (TGF beta) to induce epithelial-mesenchymal transition (EMT) is mediated by SMAD-dependent and SMAD-independent pathways such as the activation of Rho GTPase signalling. Upon activation, GTP-bound Rho stimulates the ROCK kinases, which in turn phosphorylate numerous substrates including the LIM kinases (LIMK). The net result of ROCK activation is increased actin-myosin contractile force generation, with a contribution from LIMK-induced actin filament stabilisation. In this study, we made use of siRNA-mediated knockdown and selective inhibitors to determine the contributions of ROCK and LIMK to TGF beta-induced responses. We find that both ROCK and LIMK are required for TGF beta stimulation of serum-response factor (SRF) transcriptional activity and actin stress fibre formation during EMT. In contrast, although LIMK inhibition had little effect on cell motility in scratch wound and Transwell migration assays, ROCK inhibition actually promoted TGF beta-induced cell motility by helping individual cells to break free from the epithelial sheet. Furthermore, we demonstrate that selective inhibition of LIMK, but not ROCK, effectively blocked TGF beta driven invasion through a layer of matrigel extracellular matrix protein. These results indicate that the roles of LIMK and ROCK in the Rho signalling pathway downstream of TGF beta are not identical and suggest that LIMK represents an attractive therapeutic target in TGF beta driven organ fibrosis and metastatic cancer spread. (C) 2010 Elsevier GmbH. All rights reserved. |
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