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p120 GAP modulates Ras activation of Jun kinases and transformation
G J Clark1, J K Westwick, C J Der
1Department of Pharmacology, University of North Carolina, and The Lineberger Comprehensive Cancer Center, Chapel Hill, North Carolina 27599, USA.
Abstract:
Although recent evidence demonstrates that Ras causes transformation by activation of multiple downstream pathways, the specific role of non-Raf effector pathways is presently unknown. Although Ras causes activation of the Jun NH2-terminal kinases (JNKs) via a Raf-independent pathway, the contribution of JNK activation to Ras transformation and the effector that mediates JNK activation have not been established. We observed that a dominant negative mutant of SEK1/JNKK, an activator of JNKs, selectively inhibited oncogenic Ras activation of JNK and Ras transformation, but not Ras activation of the p42 mitogen-activated protein kinase. In contrast, overexpression of wild type SEK1 enhanced Ras activation of JNK and transforming activity. Thus, JNK activation promotes Ras transformation. Furthermore, a dominant negative mutant of p120 GAP (designated N-GAP), a candidate Ras effector, blocked Ras, but not Raf, transformation and blocked Ras, but not Rac, activation of JNK. Since N-GAP overexpression reduced the association of p190 Rac/Rho GAP with endogenous p120 GAP, N-GAP may form nonproductive complexes with components critical for p120 GAP function. In summary, p120 GAP may function as an effector for Ras activation of JNK and Ras transformation.
Insights
Ras transformation involves Jun NH2-terminal kinases (JNKs) acting through a Raf-independent pathway. p120 GTPase-activating protein (GAP) appears to be a key effector mediating Ras-induced JNK activation and cell transformation.
Area of Science:
- Cellular Biology
- Oncology
- Molecular Biology
Background:
- Ras proteins are key regulators of cell signaling pathways.
- Ras-mediated cell transformation involves multiple downstream effectors.
- The role of non-Raf effector pathways in Ras transformation is not fully understood.
Purpose of the Study:
- To investigate the role of Jun NH2-terminal kinases (JNKs) in Ras-mediated cell transformation.
- To identify the specific Ras effector responsible for JNK activation.
- To elucidate the contribution of JNK signaling to oncogenic Ras activity.
Main Methods:
- Utilized dominant-negative mutants of SEK1/JNKK and p120 GTPase-activating protein (GAP).
- Assessed oncogenic Ras activation of JNK and p42 mitogen-activated protein kinase (MAPK).
- Examined Ras-induced cell transformation and protein-protein interactions using N-GAP.
Main Results:
- SEK1/JNKK inhibition blocked Ras-induced JNK activation and transformation.
- Overexpression of SEK1 enhanced Ras-mediated JNK activation and transforming activity.
- A dominant-negative mutant of p120 GAP inhibited Ras transformation and JNK activation.
Conclusions:
- JNK activation is a critical mediator of Ras-induced cell transformation.
- p120 GAP functions as a Ras effector for JNK activation.
- p120 GAP may regulate JNK signaling through interactions with other GAP proteins.