Overexpression of FAK promotes Ras activity through the formation of a FAK/p120RasGAP complex in malignant astrocytoma cells
Overexpression of FAK promotes Ras activity through the formation of a FAK/p120RasGAP complex in malignant astrocytoma cells
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DOI：
10.1038/sj.onc.1207541
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发表时间：
2004-05-13
期刊：
影响因子：
8
通讯作者：
Gladson, CL
中科院分区：
文献类型：
--
作者：
Hecker, TP;Ding, Q;Gladson, CL
粘着斑激酶(FAK)信号转导可能是通过调节RAS活性来实现的。我们以前已经证明，III级恶性星形细胞瘤活检标本显示FAK水平升高，并且FAK在U-251 MG恶性星形细胞瘤细胞中的过表达促进了Shc的磷酸化，Shc是潜在的RAS活性的上游中介。在这里，我们报告了FAK的过表达促进了U-251 MG恶性星形细胞瘤细胞的RAS活性，这些细胞在聚集性悬浮培养中或作为单层黏附在玻璃体连接蛋白上培养。在聚集体悬浮培养的U-251 MG细胞中，FAK的过表达也促进了FAK与RAS活性的负调控因子p120RasGAP的结合，这种联系在细胞与Vitronectin结合后被取消。在恶性星形细胞瘤活检标本中也观察到FAK与p120RasGAP的相关性，但在正常脑标本中未观察到。由于在聚集体悬浮培养的U-251 MG细胞中FAK的过表达减少了p120RasGAP与活性RAS的络合量，我们推测FAK与p120RasGAP的结合可能促进RAS的活性。Y397突变为F的FAK过表达并未导致FAK/p120RasGAP复合体的形成，也没有促进RAS活性，表明FAK的Y397残基在该复合体的形成和RAS的激活中起作用。此外，突变的FAK(397F)过表达抑制了锚定非依赖性生长。这些数据为一个以前未描述的机制提供了基础，在该机制中，FAK的高表达可以通过竞争性地招募p120RasGAP来促进RAS活性，从而减少p120RasGAP与活性RAS的关联。
Focal adhesion kinase (FAK) signaling may be mediated through the modulation of Ras activity. We have shown previously that grade III malignant astrocytoma biopsy samples exhibit elevated levels of FAK, and that overexpression of FAK in U-251MG malignant astrocytoma cells promotes the phosphorylation of Shc, a potential upstream mediator of Ras activity. Here, we report that overexpression of FAK promotes Ras activity in U-251MG malignant astrocytoma cells cultured in aggregate suspension or as monolayers adherent to vitronectin. The overexpression of FAK also promoted the association of FAK with p120RasGAP, which is a negative regulator of Ras activity, in the U-251MG cells cultured in aggregate suspension, with this association being abrogated upon plating of the cells onto vitronectin. An association of FAK with p120RasGAP also was observed in malignant astrocytoma biopsy samples, but not in normal brain samples. As overexpression of FAK in U-251MG cells in aggregate suspension culture reduced the amount of p120RasGAP complexed with active Ras, we hypothesize that the association of FAK with p120 RasGAP may facilitate Ras activity. The overexpression of a mutated FAK in which the Y397 had been mutated to F did not result in the formation of the FAK/p120RasGAP complex and did not promote Ras activity, indicating that the Y397 residue of FAK plays a role in the formation of this complex and in the activation of Ras. Moreover, the overexpression of mutated FAK (397F) was found to inhibit anchorage-independent growth. These data provide the basis for a previously undescribed mechanism in which the elevated expression of FAK can promote Ras activity through its competitive recruitment of p120RasGAP, thereby diminishing the association of p120RasGAP with active Ras.