Related Experiment Videos
An incomplete program of cellular tyrosine phosphorylations induced by kinase-defective epidermal growth factor
J D Wright1, C W Reuter, M J Weber
1Department of Microbiology, University of Virginia Health Sciences Center, Charlottesville 22908, USA.
Abstract:
Although signaling by the epidermal growth factor (EGF) receptor is thought to be dependent on receptor tyrosine kinase activity, it is clear that mitogen-activated protein (MAP) kinase can be activated by receptors lacking kinase activity. Since analysis of the signaling pathways used by kinase-defective receptors could reveal otherwise masked capabilities, we examined in detail the tyrosine phosphorylations and enzymes of the MAP kinase pathway induced by kinase-defective EGF receptors. Following EGF stimulation of B82L cells expressing a kinase-defective EGF receptor mutant (K721M), we found that ERK2 and ERK1 MAP kinases, as well as MEK1 and MEK2 were all activated, and SHC became prominently tyrosine-phosphorylated. By contrast, kinase-defective receptors failed to induce detectable phosphorylations of GAP (GTPase-activating protein), p62, JAK1, or p91STAT1, all of which were robustly phosphorylated by wild-type receptors. These data demonstrate that kinase-defective receptors induce several protein tyrosine phosphorylations, but that these represent only a subset of those seen with wild-type receptors. This suggests that kinase-defective receptors activate a heterologous tyrosine kinase with a specificity different from the EGF receptor. We found that kinase-defective receptors induced ErbB2/c-Neu enzymatic activation and ErbB2/c-Neu binding to SHC at a level even greater than that induced by wild-type receptors. Thus, heterodimerization with and activation of endogenous ErbB2/c-Neu is a possible mechanism by which kinase-defective receptors stimulate the MAP kinase pathway.
Insights
Kinase-defective epidermal growth factor (EGF) receptors activate mitogen-activated protein (MAP) kinase pathways by phosphorylating SHC and activating ErbB2/c-Neu. This suggests a mechanism independent of EGF receptor tyrosine kinase activity.
Area of Science:
- Cellular signaling pathways
- Receptor tyrosine kinases
- MAP kinase cascade
Background:
- Epidermal growth factor (EGF) receptor signaling typically relies on its tyrosine kinase activity.
- Mitogen-activated protein (MAP) kinase can be activated by receptors lacking kinase activity, suggesting alternative signaling mechanisms.
Purpose of the Study:
- To investigate the tyrosine phosphorylations and MAP kinase pathway activation induced by kinase-defective EGF receptors.
- To elucidate the specific signaling molecules and pathways engaged by non-functional EGF receptors.
Main Methods:
- Utilized B82L cells expressing a kinase-defective EGF receptor mutant (K721M).
- Stimulated cells with EGF and analyzed protein tyrosine phosphorylations and MAP kinase pathway components (ERK1/2, MEK1/2).
- Compared signaling events induced by kinase-defective receptors with those induced by wild-type EGF receptors.
Main Results:
- Kinase-defective EGF receptors activated ERK1/2 and MEK1/2, and induced SHC tyrosine phosphorylation.
- These receptors failed to phosphorylate GAP, p62, JAK1, or p91STAT1, unlike wild-type receptors.
- Kinase-defective receptors significantly increased ErbB2/c-Neu enzymatic activation and its binding to SHC.
Conclusions:
- Kinase-defective EGF receptors trigger a subset of tyrosine phosphorylations and activate the MAP kinase pathway.
- Activation of a heterologous tyrosine kinase, specifically ErbB2/c-Neu, is a likely mechanism for this signaling.
- ErbB2/c-Neu heterodimerization and activation by kinase-defective EGF receptors mediate MAP kinase pathway stimulation.