Modulation of acto-myosin contractility in skeletal muscle myoblasts uncouples growth arrest from differentiation
Modulation of acto-myosin contractility in skeletal muscle myoblasts uncouples growth arrest from differentiation
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DOI：
10.1242/jcs.01197
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发表时间：
2004-08-01
影响因子：
4
通讯作者：
Helfman, DM
中科院分区：
文献类型：
--
作者：
Dhawan, J;Helfman, DM
细胞-基质相互作用触发调节生长控制和组织特异性基因表达的关键信号通路。我们以前已经表明，废除粘附相互作用的悬浮培养的结果在去逮捕成肌细胞。我们报告说，在粘附细胞中阻断粘附依赖性信号的细胞内传递模拟了粘附接触的缺乏。我们研究了肌动球蛋白收缩性的药理学抑制剂对C2 C12肌原细胞生长和分化的影响。ML 7（5-碘萘-1-磺酰基高哌嗪）和BDM（2，3，丁二酮单肟）分别是肌球蛋白轻链激酶和肌球蛋白重链ATP酶的特异性抑制剂。ML 7和BDM通过减少粘着斑和应力纤维影响细胞形状。这两种抑制剂以剂量依赖性、可逆的方式迅速阻断DNA合成。此外，ML 7和BDM都抑制MyoD和肌细胞生成素的表达，诱导p27（kip 1），但不诱导p21（cip 1），并抑制分化。因此，与悬浮-停滞一样，抑制粘附细胞中的肌动蛋白-肌球蛋白收缩性导致停滞与分化解偶联。过表达的抑制剂的小GTAL-RhoA（显性负RhoA和C3转移酶）模仿肌球蛋白抑制剂的影响。相比之下，野生型RhoA诱导停滞，维持MyoD并激活肌细胞生成素和p21。表情Rho效应激酶ROCK似乎不介导Rho对MyoD的作用。因此，ROCK和MLCK在肌源性程序中发挥不同的作用。由MLCK调节的信号是关键的，因为MLCK的抑制抑制MyoD的表达，但ROCK的抑制没有。抑制收缩抑制MyoD，但没有减少肌动蛋白聚合物水平。然而，肌动蛋白解聚与latrunculin B抑制MyoD的表达。两者合计，我们的观察表明，肌动蛋白聚合物的状态和收缩调节MyoD的表达。我们认为，在成肌细胞中，Rho通路和调节肌动球蛋白的收缩性可能定义一个控制点的分化和细胞周期的条件解偶联。
Cell-substratum interactions trigger key signaling pathways that modulate growth control and tissue-specific gene expression. We have previously shown that abolishing adhesive interactions by suspension culture results in Go arrest of myoblasts. We report that blocking intracellular transmission of adhesion-dependent signals in adherent cells mimics the absence of adhesive contacts. We investigated the effects of pharmacological inhibitors of acto-myosin contractility on growth and differentiation of C2C12 myogenic cells. ML7 (5-iodonaphthalene-1-sulfonyl homopiperazine) and BDM (2,3, butanedione monoxime) are specific inhibitors of myosin light chain kinase, and myosin heavy chain ATPase, respectively. ML7 and BDM affected cell shape by reducing focal adhesions and stress fibers. Both inhibitors rapidly blocked DNA synthesis in a dose-dependent, reversible fashion. Furthermore, both ML7 and BDM suppressed expression of MyoD and myogenin, induced p27(kip1) but not p21(cip1), and inhibited differentiation. Thus, as with suspension-arrest, inhibition of acto-myosin contractility in adherent cells led to arrest uncoupled from differentiation. Over-expression of inhibitors of the small GTPase RhoA (dominant negative RhoA and C3 transferase) mimicked the effects of myosin inhibitors. By contrast, wild-type RhoA induced arrest, maintained MyoD and activated myogenin and p21. expression. The Rho effector kinase ROCK did not appear to mediate Rho's effects on MyoD. Thus, ROCK and MLCK play different roles in the myogenic program. Signals regulated by MLCK are critical, since inhibition of MLCK suppressed MyoD expression but inhibition of ROCK did not. Inhibition of contractility suppressed MyoD but did not reduce actin polymer levels. However, actin depolymerization with latrunculin B inhibited MyoD expression. Taken together, our observations indicate that actin polymer status and contractility regulate MyoD expression. We suggest that in myoblasts, the Rho pathway and regulation of acto-myosin contractility may define a control point for conditional uncoupling of differentiation and the cell cycle.