Myosin-mediated cytoskeleton contraction and Rho GTPases regulate laminin-5 matrix assembly
Myosin-mediated cytoskeleton contraction and Rho GTPases regulate laminin-5 matrix assembly
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DOI：
10.1002/cm.10161
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发表时间：
2004-02-01
影响因子：
--
通讯作者：
Jones, JCR
中科院分区：
文献类型：
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作者：
deHart, GW;Jones, JCR
层粘连蛋白-5是上皮组织基底膜的主要结构元件。在培养的上皮细胞基质中，层粘连蛋白-5排列成复杂的图案。在这里，我们测试了一个假设，即肌球蛋白II介导的肌动蛋白收缩是必要的层粘连蛋白-5基质培养的SCC 12上皮细胞的正确组装。为此，细胞用ML-7（一种肌球蛋白H轻链激酶抑制剂）或Y-27632（一种Rho激酶（ROCK）抑制剂）处理，这两种物质都能阻断肌动球蛋白收缩。在这些条件下，层粘连蛋白-5在细胞周边的致密斑块中显示异常定位。由于ROCK活性受小GTP酶Rho调节，这表明GTP酶Rho家族的成员对于SCC 12细胞的层粘连蛋白-5基质组装也可能是重要的。我们证实了这一假设，因为SCC 12细胞表达抑制RhoA，Rac和Cdc 42的突变蛋白组装相同的异常层粘连蛋白-5蛋白阵列作为药物处理的细胞。我们还评估了ML-7和Y-27632处理的细胞或RhoA，Rac和Cdc 42活性被抑制的细胞中层粘连蛋白-5受体α 3 β 1和α 6 β 4整联蛋白和半桥粒蛋白的组织。在所有情况下，α 3 β 1和α 6 β 4整联蛋白异源二聚体以及半桥粒蛋白与层粘连蛋白-5精确定位在细胞基质中。总之，我们的研究结果提供的证据表明，肌球蛋白II介导的肌动蛋白收缩和Rho GTPases的活性是必要的适当组织的层粘连蛋白-5矩阵和本地化的半桥粒蛋白阵列在上皮细胞。(C)2004威利-利斯公司
Laminin-5 is a major structural element of epithelial tissue basement membranes. In the matrix of cultured epithelial cells, laminin-5 is arranged into intricate patterns. Here we tested a hypothesis that myosin II-mediated actin contraction is necessary for the proper assembly of a laminin-5 matrix by cultured SCC12 epithelial cells. To do so, the cells were treated with ML-7, a myosin H light chain kinase inhibitor, or Y-27632, an inhibitor of Rho-kinase (ROCK), both of which block actomyosin contraction. Under these conditions, laminin-5 shows an aberrant localization in dense patches at the cell periphery. Since ROCK activity is regulated by the small GTPase Rho, this suggests that members of the Rho family of GTPases may also be important for laminin-5 matrix assembly by SCC12 cells. We confirmed this hypothesis since SCC12 cells expressing mutant proteins that inhibit RhoA, Rac, and Cdc42 assemble the same aberrant laminin-5 protein arrays as drug-treated cells. We have also evaluated the organization of the laminin-5 receptors alpha3beta1 and alpha6beta4 integrin and hemidesmosome proteins in ML-7- and Y-27632-treated cells or in cells in which RhoA, Rac, and Cdc42 activity were inhibited. In all instances, alpha3beta1 and alpha6beta4 integrin heterodimers, as well as hemidesmosome proteins, localize precisely with laminin-5 in the matrix of the cells. In summary, our results provide evidence that myosin II-mediated actin contraction and the activity of Rho GTPases are necessary for the proper organization of a laminin-5 matrix and localization of hemidesmosome protein arrays in epithelial cells. (C) 2004 Wiley-Liss, Inc.