Oncogenic Ras leads to Rho activation by activating the mitogen-activated protein kinase pathway and decreasing Rho-GTPase-activating protein activity
Oncogenic Ras leads to Rho activation by activating the mitogen-activated protein kinase pathway and decreasing Rho-GTPase-activating protein activity
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DOI：
10.1074/jbc.m207943200
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发表时间：
2003-01-31
影响因子：
4.8
通讯作者：
Pilz, RB
中科院分区：
文献类型：
--
作者：
Chen, JC;Zhuang, SH;Pilz, RB
通过致癌Ras的转化需要通过Rho家族蛋白包括RhoA的信号传导，但是致癌Ras调节RhoA活性的机制是未知的。我们研究了Ras对NIH 3 T3细胞中RhoA活性的影响，NIH 3 T3细胞在诱导型启动子控制下稳定转染H-Ras（V12）或瞬时表达活化的H-Ras。使用一种新的方法来定量与Rho结合的GTP，我们发现致癌Ras的表达使Rho活性增加约2倍。Rho活性增加与RhoA的质膜结合增加和Rho/Ras调节的p21（WAF 1/CIP 1）启动子的活性降低相关。致癌Ras激活RhoA可以解释为胞质p190 Rho-GAP活性的降低和p190 Rho-GAP从胞质转移到洗涤剂不溶性的细胞骨架部分。Ras/Raf/MEK/ERK通路的药理学抑制防止Ras诱导的RhoA活化和p190 Rho-GAP易位;组成性活性Raf-1激酶或MEK的表达足以诱导p190 Rho-GAP易位。我们的结论是，在NIH 3 T3细胞中，致癌Ras通过Raf/MEK/ERK途径激活RhoA，通过降低胞浆活性和改变p190 Rho-GAP的亚细胞定位。
Transformation by oncogenic Ras requires signaling through Rho family proteins including RhoA, but the mechanism(s) whereby oncogenic Ras regulates the activity of RhoA is (are) unknown. We examined the effect of Ras on RhoA activity in NIH 3T3 cells either stably transfected with H-Ras(V12) under control of an inducible promoter or transiently expressing the activated H-Ras. Using a novel method to quantitate enzymatically the GTP bound to Rho, we found that expression of the oncogenic Ras increased Rho activity similar to2-fold. Increased Rho activity was associated with increased plasma membrane binding,of RhoA and decreased activity of the Rho/Ras-regulated p21(WAF1/CIP1) promoter. RhoA activation by oncogenic Ras could be explained by a decrease in cytosolic p190 Rho-GAP activity and translocation of p190 Rho-GAP from the cytosol to a detergent-insoluble cytoskeletal fraction. Pharmacologic inhibition of the Ras/Raf/MEK/ERK pathway prevented Ras-induced activation of RhoA and translocation of p190 Rho-GAP; expression of constitutively active Raf-1 kinase or MEK was sufficient to induce p190 Rho-GAP translocation. We conclude that in NIH 3T3 cells oncogenic Ras activates RhoA through the Raf/MEK/ERK pathway by decreasing the cytosolic activity and changing the subcellular localization of p190 Rho-GAP.