Kinoshita T, Shirouzu M, Kamiya A, Hashimoto K, Yokoyama S, Miyajima A
Department of Cellular Biosynthesis, Institute of Molecular and Cellular Bioscience, The University of Tokyo, Bunkyo-ku, Japan.
Oncogene. 1997 Aug 7;15(6):619-27. doi: 10.1038/sj.onc.1201234.
The Ras signal transduction pathway is activated by a number of hematopoietic cytokines and is implicated in the prevention of apoptotic death in hematopoietic cells. Recent studies have provided evidence that the downstream of Ras is highly divergent and several independent pathways appear to mediate distinct biological functions of Ras. In the present study, we investigated the downstream pathway(s) of Ras responsible for the maintenance of hematopoietic cell survival by using various mutants of signaling molecules. Activation of the Raf/MAPK pathway in interleukin (IL) 3-dependent cells by expression of an oncogenic Raf or a Ras mutant (G12V/T35S) prevented apoptosis following IL-3 deprivation. In contrast, another Ras mutant (G12V/V45E), which is apparently incapable of activating MAPK, efficiently blocked apoptosis as well. It is therefore likely that the activation of the Raf/MAPK pathway is not an absolute requirement for the prevention of apoptosis, and there appears to be a Raf/MAPK-independent pathway that contributes to hematopoietic cell survival. Since Ras(G12V/V45E) was able to cause the phosphorylation of p70/S6 kinase, we inhibited the S6 kinase pathway by rapamycin and by wortmannin, and found that the anti-apoptotic function of Ras(G12V/V45E), but not of Ras(G12V), was critically influenced by both inhibitors. These results indicate that the Raf/MAPK and a rapamycin/wortmannin-sensitive pathways mediate Ras function to prevent apoptotic death in hematopoietic cells.
Ras信号转导通路可被多种造血细胞因子激活，并与预防造血细胞的凋亡性死亡有关。最近的研究表明，Ras的下游高度分化，几条独立的通路似乎介导了Ras不同的生物学功能。在本研究中，我们使用信号分子的各种突变体，研究了负责维持造血细胞存活的Ras下游通路。通过表达致癌性Raf或Ras突变体（G12V/T35S）激活白细胞介素（IL）3依赖性细胞中的Raf/MAPK通路，可防止IL-3缺失后的细胞凋亡。相比之下，另一种明显无法激活MAPK的Ras突变体（G12V/V45E）也能有效阻断细胞凋亡。因此，Raf/MAPK通路的激活可能不是预防细胞凋亡的绝对必要条件，似乎存在一条不依赖Raf/MAPK的通路有助于造血细胞存活。由于Ras（G12V/V45E）能够导致p70/S6激酶磷酸化，我们用雷帕霉素和渥曼青霉素抑制S6激酶通路，并发现这两种抑制剂对Ras（G12V/V45E）而非Ras（G12V）的抗凋亡功能有严重影响。这些结果表明，Raf/MAPK和雷帕霉素/渥曼青霉素敏感的通路介导了Ras在预防造血细胞凋亡性死亡中的功能。
