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The function of GRB2 in linking the insulin receptor to Ras signaling pathways
1Department of Pharmacology, New York University Medical Center, NY 10016.
Abstract:
Insulin-induced activation of extracellular signal-regulated kinases [ERKs, also known as mitogen-activated protein (MAP) kinases] is mediated by Ras. Insulin activates Ras primarily by increasing the rate of guanine nucleotide-releasing activity. Here, we show that insulin-induced activation of ERKs was enhanced by stable overexpression of growth factor receptor-bound protein 2 (GRB2) but not by overexpression of GRB2 proteins with point mutations in the Src homology 2 and 3 domains. Moreover, a dominant negative form of Ras (with Ser17 substituted with Asn) blocked insulin-induced activation of ERKs in cells that overexpressed GRB2. GRB2 overexpression led to increased formation of a complex between the guanine nucleotide-releasing factor Sos (the product of the mammalian homolog of son of sevenless gene) and GRB2. In response to insulin stimulation, this complex bound to tyrosine-phosphorylated IRS-1 (insulin receptor substrate-1) and Shc. In contrast to the activated epidermal growth factor receptor that binds the GRB2-Sos complex directly, activation of the insulin receptor results in the interaction of GRB2-Sos with IRS-1 and Shc, thus linking the insulin receptor to Ras signaling pathways.
Insights
Insulin activates extracellular signal-regulated kinases (ERKs) via Ras. Growth factor receptor-bound protein 2 (GRB2) enhances this insulin-induced ERK activation by facilitating the formation of a complex that links the insulin receptor to Ras signaling.
Area of Science:
- Cellular signaling
- Molecular biology
- Endocrinology
Background:
- Insulin signaling pathways are crucial for glucose homeostasis and cellular growth.
- Extracellular signal-regulated kinases (ERKs) are key mediators of mitogenic and differentiation signals.
- Ras GTPases are central regulators of these signaling cascades.
Purpose of the Study:
- To elucidate the role of growth factor receptor-bound protein 2 (GRB2) in insulin-induced extracellular signal-regulated kinase (ERK) activation.
- To investigate the interaction of GRB2 with upstream signaling components in response to insulin.
- To understand how the insulin receptor links to Ras signaling pathways via GRB2.
Main Methods:
- Overexpression of wild-type and mutant growth factor receptor-bound protein 2 (GRB2) in cultured cells.
- Introduction of a dominant-negative Ras mutant to block downstream signaling.
- Analysis of protein-protein interactions using co-immunoprecipitation assays.
- Assessment of ERK activation by Western blotting.
Main Results:
- Stable overexpression of GRB2 enhanced insulin-induced ERK activation, while mutated GRB2 proteins did not.
- A dominant-negative Ras mutant abrogated insulin-induced ERK activation in GRB2-overexpressing cells.
- GRB2 overexpression increased the formation of a GRB2-Sos complex.
- This GRB2-Sos complex associated with tyrosine-phosphorylated insulin receptor substrate-1 (IRS-1) and Shc upon insulin stimulation.
Conclusions:
- Growth factor receptor-bound protein 2 (GRB2) is essential for insulin-induced extracellular signal-regulated kinase (ERK) activation.
- Insulin receptor signaling converges on the GRB2-Sos complex, which then activates Ras.
- This study reveals a critical mechanism linking the insulin receptor to Ras/ERK pathways through IRS-1 and Shc scaffolding proteins.