Diacylglycerol Kinase η Augments C-Raf Activity and B-Raf/C-Raf Heterodimerization
Diacylglycerol Kinase η Augments C-Raf Activity and B-Raf/C-Raf Heterodimerization
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DOI：
10.1074/jbc.m109.043604
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发表时间：
2009-10-23
影响因子：
4.8
通讯作者：
Sakane, Fumio
中科院分区：
文献类型：
--
作者：
Yasuda, Satoshi;Kai, Masahiro;Sakane, Fumio
Ras/B-Raf/C-Raf/MEK/ERK 信号级联对于控制许多基本细胞过程（包括增殖、存活和分化）至关重要。这项研究表明，二酰基甘油激酶 eta (DGK eta) 的小干扰 RNA 依赖性敲低会损害 HeLa 细胞中表皮生长因子 (EGF) 激活的 Ras/B-Raf/C-Raf/MEK/ERK 通路。相反，DGK eta 1 的过表达可以以 DGK 活性无关的方式激活 Ras/B-Raf/C-Raf/MEK/ERK 通路，表明 DGK eta 可以作为支架/适配器蛋白。通过测定 DGK eta 沉默的 HeLa 细胞中该通路所有成分的活性，本研究表明 DGK eta 激活 C-Raf，但不激活 B-Raf。此外，这项研究表明，DGK eta 增强了 EGF 诱导的 C-Raf 与 B-Raf 的异二聚化，从而将信号传递给 C-Raf。 DGK eta 与 B-Raf 和 C-Raf 发生物理相互作用，调节 EGF 诱导的 B-Raf 和 C-Raf 从细胞质到细胞膜的募集。 C-Raf 的 DGK eta 依赖性激活发生在下游或独立于已知的 C-Raf 修饰，例如 Ser-259 处的去磷酸化、Ser-338 处的磷酸化以及与 14-3-3 蛋白的相互作用。总而言之，这些结果有力地支持了 DGK eta 通过先前未识别的机制作为 Ras/B-Raf/C-Raf/MEK/ERK 信号级联的新型关键调节组件。
The Ras/B-Raf/C-Raf/MEK/ERK signaling cascade is critical for the control of many fundamental cellular processes, including proliferation, survival, and differentiation. This study demonstrated that small interfering RNA-dependent knockdown of diacylglycerol kinase eta (DGK eta) impaired the Ras/B-Raf/C-Raf/MEK/ERK pathway activated by epidermal growth factor (EGF) in HeLa cells. Conversely, the overexpression of DGK eta 1 could activate the Ras/B-Raf/C-Raf/MEK/ERK pathway in a DGK activity-independent manner, suggesting that DGK eta serves as a scaffold/adaptor protein. By determining the activity of all the components of the pathway in DGK eta-silenced HeLa cells, this study revealed that DGK eta activated C-Raf but not B-Raf. Moreover, this study demonstrated that DGK eta enhanced EGF-induced heterodimerization of C-Raf with B-Raf, which transmits the signal to C-Raf. DGK eta physically interacted with B-Raf and C-Raf, regulating EGF-induced recruitment of B-Raf and C-Raf from the cytosol to membranes. The DGK eta-dependent activation of C-Raf occurred downstream or independently of the already known C-Raf modifications, such as dephosphorylation at Ser-259, phosphorylation at Ser-338, and interaction with 14-3-3 protein. Taken together, the results obtained strongly support that DGK eta acts as a novel critical regulatory component of the Ras/B-Raf/C-Raf/MEK/ERK signaling cascade via a previously unidentified mechanism.