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Cross-cascade activation of ERKs and ternary complex factors by Rho family proteins
J A Frost1, H Steen, P Shapiro
1U.T. Southwestern Medical Center, Department of Pharmacology, Dallas 75235-9041, USA.
Abstract:
Mitogens promote cell growth through integrated signal transduction networks that alter cellular metabolism, gene expression and cytoskeletal organization. Many such signals are propagated through activation of MAP kinase cascades partly regulated by upstream small GTP-binding proteins. Interactions among cascades are suspected but not defined. Here we show that Rho family small G proteins such as Rac1 and Cdc42hs, which activate the JNK/SAPK pathway, cooperate with Raf-1 to activate the ERK pathway. This causes activation of ternary complex factors (TCFs), which regulate c-fos gene expression through the serum response element. Examination of ERK pathway kinases shows that neither MEK1 nor Ras will synergize with Rho-type proteins, and that only MEK1 is fully activated, indicating that MEKs are a focal point for cross-cascade regulation. Rho family proteins utilize PAKs for this effect, as expression of an active PAK1 mutant can substitute for Rho family small G proteins, and expression of an interfering PAK1 mutant blocks Rho-type protein stimulation of ERKs. PAK1 phosphorylates MEK1 on Ser298, a site important for binding of Raf-1 to MEK1 in vivo. Expression of interfering PAK1 also reduces stimulation of TCF function by serum growth factors, while expression of active PAK1 enhances EGF-stimulated MEK1 activity. This demonstrates interaction among MAP kinase pathway elements not previously recognized and suggests an explanation for the cooperative effect of Raf-1 and Rho family proteins on cellular transformation.
Insights
Small G proteins like Rac1 and Cdc42hs cooperate with Raf-1 to activate the ERK pathway by targeting MEK1. This cross-cascade regulation impacts gene expression and cellular transformation.
Area of Science:
- Cellular biology
- Molecular signaling
- Signal transduction pathways
Background:
- Mitogens regulate cell growth via complex signal transduction networks.
- MAP kinase cascades, regulated by small GTP-binding proteins, propagate many mitogenic signals.
- Interactions between these signaling cascades are suspected but not fully understood.
Purpose of the Study:
- To investigate the interaction between Rho family small G proteins and MAP kinase pathways.
- To elucidate the mechanism by which Rho proteins influence ERK pathway activation.
- To identify key regulatory points in cross-cascade signaling.
Main Methods:
- Utilized expression of active and interfering mutants of PAK1 (a downstream effector of Rho proteins).
- Examined the phosphorylation of MEK1 at Ser298.
- Assessed the activation of ternary complex factors (TCFs) and c-fos gene expression.
- Investigated the synergistic effects of Rho proteins, Raf-1, MEK1, and Ras on ERK pathway activation.
Main Results:
- Rho family proteins (Rac1, Cdc42hs) cooperate with Raf-1 to activate the ERK pathway.
- MEK1, but not Ras, is a focal point for cross-cascade regulation, being fully activated by Rho proteins.
- Rho proteins utilize PAKs, which phosphorylate MEK1 at Ser298, facilitating Raf-1 binding.
- PAK1 activity correlates with TCF function and c-fos expression, demonstrating cross-talk between pathways.
Conclusions:
- Rho family proteins interact with MAP kinase pathway elements, specifically MEK1, through PAK1.
- This interaction provides a mechanism for the cooperative effects of Raf-1 and Rho proteins on cellular transformation.
- Identified MEK1 as a critical node for cross-cascade regulation within MAP kinase signaling networks.