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Lysophosphatidic acid stimulates mitogen-activated protein kinase activation via a G-protein-coupled pathway
1Institute of Cancer Research, Chester Beatty Laboratories, London, United Kingdom.
Abstract:
Activation of tyrosine kinase receptors causes mitogen-activated protein (MAP) kinase stimulation via a pathway involving p21ras, p74raf-1 (acting as a MAP kinase kinase kinase), and MAP kinase kinases; however, the pathway by which heterotrimeric G-protein-coupled receptors activate MAP kinases is undefined. Since there are several MAP kinase kinase kinases it has been suggested that p74raf-1 may only couple tyrosine kinase receptors to MAP kinase activation. We therefore investigated the requirement for p21ras and p74raf-1 in G-protein receptor-mediated MAP kinase activation. Lysophosphatidic acid stimulates MAP kinase via a pertussis toxin-sensitive pathway, which is blocked by dominant negative Ras. Lysophosphatidic acid-stimulated MAP kinase activation is potentiated by overexpression of p74raf-1 and blocked by expression of a dominant negative Raf protein comprising the N-terminal 259 amino acids. We conclude that lysophosphatidic acid activates MAP kinases by a G-protein-coupled pathway that requires both p21ras and p74raf-1.
Insights
G-protein-coupled receptors activate mitogen-activated protein (MAP) kinases through a pathway involving p21ras and p74raf-1. This study clarifies the signaling cascade for G-protein-coupled receptor-mediated MAP kinase activation.
Area of Science:
- Cellular signaling pathways
- Molecular biology
- Signal transduction
Background:
- Mitogen-activated protein (MAP) kinase activation is well-defined downstream of tyrosine kinase receptors.
- The specific pathway for MAP kinase activation by heterotrimeric G-protein-coupled receptors remains unclear.
- p74raf-1 is a potential MAP kinase kinase kinase, but its role in G-protein-coupled receptor signaling is debated.
Purpose of the Study:
- To investigate the involvement of p21ras and p74raf-1 in G-protein-coupled receptor-mediated MAP kinase activation.
- To elucidate the signaling pathway linking G-protein-coupled receptors to MAP kinase.
- To determine if p74raf-1 is essential for this signaling cascade.
Main Methods:
- Utilized lysophosphatidic acid as a stimulus for G-protein-coupled receptor activation.
- Employed dominant-negative Ras and dominant-negative Raf constructs to inhibit specific pathway components.
- Assessed MAP kinase activation through overexpression and inhibition studies of p74raf-1.
Main Results:
- Lysophosphatidic acid stimulates MAP kinase via a pertussis toxin-sensitive pathway.
- Dominant-negative Ras blocked lysophosphatidic acid-induced MAP kinase activation.
- Overexpression of p74raf-1 potentiated the response, while dominant-negative Raf inhibited it.
Conclusions:
- Lysophosphatidic acid activates MAP kinases through a G-protein-coupled pathway.
- This pathway critically requires both p21ras and p74raf-1.
- p74raf-1 is a necessary component in G-protein-coupled receptor-mediated MAP kinase activation.