Crosstalk between PI3 Kinase/PDK1/Akt/Rac1 and Ras/Raf/MEK/ERK Pathways Downstream PDGF Receptor
Crosstalk between PI3 Kinase/PDK1/Akt/Rac1 and Ras/Raf/MEK/ERK Pathways Downstream PDGF Receptor
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DOI：
10.1159/000350108
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发表时间：
2013-01-01
影响因子：
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通讯作者：
Nishizaki, Tomoyuki
中科院分区：
文献类型：
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作者：
Niba, Emma Tabe Eko;Nagaya, Hisao;Nishizaki, Tomoyuki
背景/目标：我们早期的研究表明IRS/PI 3激酶/PDK 1/Akt/Rac 1/ROCK和（Shc 2/Grb 2/SOS）/Ras/Raf/MEK/ERK通路下游的PDGF-β β受体之间的串扰负责恶性间皮瘤细胞的趋化和增殖。本研究的目的就是为了获得这方面的证据。研究方法：为了评估Akt、MEK和ERK的活化，使用针对磷酸-Thr 308-Akt、磷酸-Ser 473-Akt、Akt、磷酸-MEK、MEK、磷酸-ERK 1/2和ERK 1/2的抗体对MSTO-211 H恶性间皮瘤细胞进行蛋白质印迹。为了敲低Akt、PI 3激酶、PDK 1和Rac 1，构建沉默每个靶向基因的siRNA并转染细胞。为了监测Rac 1活性，对活细胞和固定细胞进行FRET监测。结果如下：ERK在MSTO-211 H细胞中在基础条件下被激活，并且通过PI 3激酶、PDK 1、Akt和Rac 1的抑制剂或通过敲低PI 3激酶、PDK 1、Akt和Rac 1来阻止激活。Akt在基础条件下也被激活，并且该激活被MEK抑制剂和ERK 1/2抑制剂抑制。在FRET分析中，Rac 1在基础条件下被激活，并且该激活被MEK抑制剂和ERK 1/2抑制剂抑制。结论：本研究的结果表明，ERK可以被PI 3激酶，PDK 1，Akt和Rac 1激活，或者，Akt和Rac 1可以被MEK和ERK激活。版权所有（C）2013 S. Karger AG，巴塞尔
Background/Aims: Our earlier studies suggested crosstalk between IRS/PI3 kinase/PDK1/Akt/Rac1/ROCK and (Shc2/Grb2/SOS)/Ras/Raf/MEK/ERK pathways downstream PDGF-beta beta receptor responsible for chemotaxis and proliferation of malignant mesothelioma cells. The present study was conducted to obtain evidence for this. Methods: To assess activation of Akt, MEK, and ERK, Western blotting was carried out on MSTO-211H malignant mesothelioma cells using antibodies against phospho-Thr308-Akt, phopho-Ser473-Akt, Akt, phospho-MEK, MEK, phopho-ERK1/2, and ERK1/2. To knock-down Akt, PI3 kinase, PDK1, and Rac1, siRNAs silencing each-targeted gene were constructed and transfected into cells. To monitor Rac1 activity, FRET monitoring was carried out on living and fixed cells. Results: ERK was activated under the basal conditions in MSTO-211H cells, and the activation was prevented by inhibitors for PI3 kinase, PDK1, Akt, and Rac1 or by knocking-down PI3 kinase, PDK1, Akt, and Rac1. Akt was also activated under the basal conditions, and the activation was suppressed by a MEK inhibitor and an ERK1/2 inhibitor. In the FRET analysis, Rac1 was activated under the basal conditions, and the activation was inhibited by a MEK inhibitor and an ERK1/2 inhibitor. Conclusion: The results of the present study show that ERK could be activated by PI3 kinase, PDK1, Akt, and Rac1 and that alternatively, Akt and Rac1 could be activated by MEK and ERK in MSTO-211H cells. Copyright (C) 2013 S. Karger AG, Basel