Involvement of the RASSF1A tumor suppressor gene in controlling cell migration
Involvement of the RASSF1A tumor suppressor gene in controlling cell migration
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DOI：
10.1158/0008-5472.can-05-0247
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发表时间：
2005-09-01
期刊：
影响因子：
11.2
通讯作者：
Latif, F
中科院分区：
文献类型：
--
作者：
Dallol, A;Agathanggelou, A;Latif, F
我们之前已经证明RASSF1A与微管相关。这种关联改变了微管动力学，似乎对RASSF1A的肿瘤抑制功能至关重要。不完全与微管结合的RASSF1A突变变体降低了稳定微管的能力，并导致细胞周期停滞。在这里，我们发现RASSF1A的过表达降低了A549非小细胞肺癌细胞通过transwell过滤器或关闭伤口的能力。此外，我们采用基因敲除和Rassfla敲除小鼠的小鼠胚胎成纤维细胞（mef）来分析RASSF1A在控制细胞运动中的功能。与对照相比，稳定转染RASSF1A的A549细胞表现出细胞间黏附增加和折射形态减少。相反，在HeLa中，RASSF1A敲低导致细胞-细胞粘附丧失和更多的折射形态。Rassf1a耗尽的HeLa细胞以及Rassf1a(-/-) mef显示出细胞迁移增加，这可能部分依赖于磷脂酰肌醇3-激酶。延时显微镜显示，rassf1a缺失的细胞具有高度运动性，呈成纤维细胞样形态，细胞间粘连减弱。rassf1a缺失的HeLa细胞和mef的细胞骨架染色显示，微管生长和肌动蛋白应激纤维形成方面存在显著差异。这一观察结果与rassf1a敲低细胞和Rassfla(-/-) mef中Rac1活化增加有关。此外，在rassf1a缺失的HeLa细胞中，Rac1的显性阴性变体的表达降低了它们形成板足和其他突起的能力。这些发现代表了RASSF1A的一种新功能，这可能有助于解释其肿瘤抑制能力独立于其对细胞周期和凋亡的影响。
We have previously shown that RASSF1A associates with the microtubules. This association alters the microtubule dynamics and seems essential for RASSF1A tumor suppressive function. Mutant variants of RASSF1A that do not associate fully with the microtubules have reduced ability to stabilize them and cause cell cycle arrest. Here we show that overexpression of RASSF1A diminished the ability of A549 non-small cell lung cancer cells to migrate either through a transwell filter or to close a wound. In addition, we employed gene knockdown as well as mouse embryonic fibroblasts (MEFs) from Rassfla knockout mice to analyze RASSF1A function in controlling cell motility. A549 cells stably transfected with RASSF1A exhibited increased cell-cell adhesion and less refractive morphology compared with controls. Conversely, RASSF1A knockdown in HeLa caused loss of cell-cell adhesion and a more refractive morphology. RASSF1Adepleted HeLa cells as well as Rassf1a(-/-) MEFs displayed increased cell migration that could be partly phosphatidylinositol 3-kinase dependent. Time-lapse microscopy showed the RASSF1A-depleted cells are highly motile with fibroblastlike morphology and diminished cell-cell adhesion. Staining of the cytoskeleton in RASSF1A-depleted HeLa cells and MEFs show marked differences in terms of microtubules outgrowth and actin stress fibers formation. This observation was associated with increased activation of Rac1 in RASSF1A-knockdown cells and the Rassfla(-/-) MEFs. In addition, expression of a dominant-negative variant of Rac1 in the RASSF1A-depleted HeLa cells reduced their ability to form lamellipodia and other protrusions. These findings represent a novel function for RASSF1A, which may help explain its tumor suppression ability independently of its effects on cell cycle and apoptosis.