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Raf/MAPK and rapamycin-sensitive pathways mediate the anti-apoptotic function of p21Ras in IL-3-dependent
T Kinoshita1, M Shirouzu, A Kamiya
1Department of Cellular Biosynthesis, Institute of Molecular and Cellular Bioscience, The University of Tokyo, Bunkyo-ku, Japan.
Abstract:
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.
Insights
Ras signaling prevents hematopoietic cell death through both the Raf/MAPK pathway and a separate rapamycin/wortmannin-sensitive pathway, indicating divergent downstream signaling for cell survival.
Area of Science:
- Cell Biology
- Molecular Biology
- Signal Transduction
Background:
- The Ras signaling pathway is crucial for hematopoietic cell survival, activated by cytokines.
- Downstream pathways of Ras are diverse, mediating distinct cellular functions.
- Understanding these pathways is key to preventing apoptotic death in hematopoietic cells.
Purpose of the Study:
- To investigate the specific downstream pathways of Ras involved in maintaining hematopoietic cell survival.
- To differentiate between the roles of Raf/MAPK and other pathways in Ras-mediated anti-apoptosis.
Main Methods:
- Utilized various Ras signaling molecule mutants in interleukin (IL)-3-dependent cells.
- Assessed apoptosis prevention following IL-3 deprivation.
- Investigated the effect of rapamycin and wortmannin on Ras mutant-induced anti-apoptotic functions.
Main Results:
- Activation of Raf/MAPK pathway by Ras mutants (G12V/T35S) prevented apoptosis.
- A Ras mutant (G12V/V45E), not activating MAPK, also blocked apoptosis, suggesting a MAPK-independent pathway.
- Ras(G12V/V45E)-mediated anti-apoptosis was sensitive to rapamycin and wortmannin, implicating p70/S6 kinase signaling.
Conclusions:
- Ras-mediated prevention of hematopoietic cell apoptosis involves at least two distinct pathways: Raf/MAPK and a rapamycin/wortmannin-sensitive pathway.
- The Raf/MAPK pathway is not essential for all Ras-mediated anti-apoptotic functions.
- Divergent Ras signaling pathways contribute to hematopoietic cell survival.