Activation of ROCK by RhoA is regulated by cell adhesion, shape, and cytoskeletal tension
Activation of ROCK by RhoA is regulated by cell adhesion, shape, and cytoskeletal tension
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DOI：
10.1016/j.yexcr.2007.07.002
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发表时间：
2007-10-01
影响因子：
3.7
通讯作者：
Chen, Christopher S.
中科院分区：
文献类型：
--
作者：
Bhadriraju, Kiran;Yang, Michael;Chen, Christopher S.
与细胞外基质的黏附通过调节Rho家族的小GTP酶的活性来调节肌动蛋白细胞骨架的许多变化。在这里，我们报告了GTP结合的RhoA激活其下游效应器ROCK所必需的黏附以及相关的细胞形状和细胞骨架张力的变化。使用内源性ROCK的体外激酶分析，我们发现悬浮中的细胞，附着在低密度纤维连接蛋白包裹的底物上，或在铺展限制性微图案岛上，都表现出低的ROCK活性和相应的低肌球蛋白轻链磷酸化，面对高水平的GTP结合的RhoA。相反，允许细胞在底物上扩散可以挽救岩石和肌球蛋白的活性。有趣的是，用细胞松弛素D或Bablbistatin抑制张力也在20分钟内抑制了岩石的活性。通过细胞分离或张力抑制而导致的岩石活性的丧失不能被具有结构性活性的RhoA-V14挽救。这些结果表明，细胞骨架张力、黏附成熟和ROCK信号之间存在反馈环路，这可能有助于许多机械力化学过程。(C)2007 Elsevier Inc.保留所有权利。
Adhesion to the extracellular matrix regulates numerous changes in the actin cytoskeleton by regulating the activity of the Rho family of small GTPases. Here, we report that adhesion and the associated changes in cell shape and cytoskeletal tension are all required for GTP-bound RhoA to activate its downstream effector, ROCK. Using an in vitro kinase assay for endogenous ROCK, we found that cells in suspension, attached on substrates coated with low density fibronectin, or on spreading-restrictive micropatterned islands all exhibited low ROCK activity and correspondingly low myosin light chain phosphorylation, in the face of high levels of GTP-bound RhoA. In contrast, allowing cells to spread against substrates rescued ROCK and myosin activity. Interestingly, inhibition of tension with cytochalasin D or blebbistatin also inhibited ROCK activity within 20 min. The abrogation of ROCK activity by cell detachment or inhibition of tension could not be rescued by constitutively active RhoA-V14. These results suggest the existence of a feedback loop between cytoskeletal tension, adhesion maturation, and ROCK signaling that likely contributes to numerous mechanochemical processes. (c) 2007 Elsevier Inc. All rights reserved.