Focal contacts as mechanosensors: externally applied local mechanical force induces growth of focal contacts by an mDia1-dependent and ROCK-independent mechanism.
Focal contacts as mechanosensors: externally applied local mechanical force induces growth of focal contacts by an mDia1-dependent and ROCK-independent mechanism.
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DOI：
10.1083/jcb.153.6.1175
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发表时间：
2001-06-11
期刊：
影响因子：
--
通讯作者：
Bershadsky AD
中科院分区：
文献类型：
--
作者：
Riveline D;Zamir E;Balaban NQ;Schwarz US;Ishizaki T;Narumiya S;Kam Z;Geiger B;Bershadsky AD
细胞 - 基质黏附从最初的点状黏着斑复合物转变为成熟的细长形式（称为黏着斑接触）需要GTP酶Rho的活性。特别是，一种被称为Rho相关激酶（ROCK）的Rho靶标激活肌球蛋白II驱动的收缩性已被证明对黏着斑接触的形成至关重要。为了剖析Rho依赖性诱导黏着斑接触的机制并阐明细胞收缩性的作用，我们使用微量移液器对细胞边缘含纽蛋白的点状黏附施加机械力。局部向心牵拉导致这些结构的局部组装和伸长，并使其发展成条纹状黏着斑接触，这通过绿色荧光蛋白标记的纽蛋白或桩蛋白的动力学以及干涉反射显微镜得以揭示。C3转移酶对Rho活性的抑制可抑制这种力诱导的黏着斑接触形成。然而，另一种Rho靶标——成蛋白同源蛋白mDia1（渡边，N.，加藤，T.，藤田，A.，石崎，T.，鸣海，S. 1999.《自然细胞生物学》1:136 - 143）的组成型活性突变体足以在经C3转移酶处理的细胞中恢复力诱导的黏着斑接触形成。在受到肌球蛋白II和ROCK抑制的细胞中，力诱导的黏着斑接触形成仍然会发生。因此，只要mDia1有活性，外部张力在诱导黏着斑接触时就可绕过ROCK介导的肌球蛋白II收缩性的需求。我们的实验表明，含整合素的黏着斑复合物作为单个机械传感器发挥作用，对局部力表现出定向组装。
The transition of cell–matrix adhesions from the initial punctate focal complexes into the mature elongated form, known as focal contacts, requires GTPase Rho activity. In particular, activation of myosin II–driven contractility by a Rho target known as Rho-associated kinase (ROCK) was shown to be essential for focal contact formation. To dissect the mechanism of Rho-dependent induction of focal contacts and to elucidate the role of cell contractility, we applied mechanical force to vinculin-containing dot-like adhesions at the cell edge using a micropipette. Local centripetal pulling led to local assembly and elongation of these structures and to their development into streak-like focal contacts, as revealed by the dynamics of green fluorescent protein–tagged vinculin or paxillin and interference reflection microscopy. Inhibition of Rho activity by C3 transferase suppressed this force-induced focal contact formation. However, constitutively active mutants of another Rho target, the formin homology protein mDia1 (Watanabe, N., T. Kato, A. Fujita, T. Ishizaki, and S. Narumiya. 1999. Nat. Cell Biol. 1:136–143), were sufficient to restore force-induced focal contact formation in C3 transferase-treated cells. Force-induced formation of the focal contacts still occurred in cells subjected to myosin II and ROCK inhibition. Thus, as long as mDia1 is active, external tension force bypasses the requirement for ROCK-mediated myosin II contractility in the induction of focal contacts. Our experiments show that integrin-containing focal complexes behave as individual mechanosensors exhibiting directional assembly in response to local force.
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