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Epidermal growth factor stimulates a protein tyrosine kinase which is separable from the epidermal growth factor
O Filhol1, E M Chambaz, G N Gill
1Institut National de la Santé et de la Recherche Medicale, DBMS/BRCE, Centre d'Etudes Nucléaires, Grenoble, France.
Abstract:
The intrinsic protein tyrosine kinase activity of the epidermal growth factor (EGF) receptor is necessary for ligand-induced signaling. To determine whether cellular protein tyrosine kinases are substrates for EGF-activated receptors, phosphotyrosine-containing proteins were isolated from EGF-treated cells and assayed for tyrosine kinase activity using peptide substrates. A tyrosine kinase activity that is distinct from the EGF receptor was adsorbed to monoclonal anti-phosphotyrosine antibody columns and eluted with phenyl phosphate. Near-maximal tyrosine phosphorylation of this kinase occurred within 1 min of cell stimulation with an ED50 for EGF of 2.5 nM. The kinase was deactivated by incubation with purified CD45 tyrosine phosphatase in vitro, but activity could be restored by incubation with purified EGF receptor and Mn2+ ATP. These results suggest a cascade of tyrosine kinase signaling analogous to well characterized serine/threonine kinase cascades.
Insights
Epidermal Growth Factor (EGF) receptor signaling involves a distinct tyrosine kinase. This kinase is activated by EGF, suggesting a tyrosine kinase cascade similar to other signaling pathways.
Area of Science:
- Cellular Biology
- Molecular Signaling
- Biochemistry
Background:
- Epidermal Growth Factor (EGF) receptor's intrinsic tyrosine kinase activity is crucial for its signaling.
- Understanding the substrates of EGF-activated receptors is key to elucidating cellular signaling pathways.
Purpose of the Study:
- To investigate whether cellular protein tyrosine kinases act as substrates for EGF-activated receptors.
- To identify and characterize tyrosine kinase activities distinct from the EGF receptor in EGF-treated cells.
Main Methods:
- Isolation of phosphotyrosine-containing proteins from EGF-treated cells.
- Assay of isolated proteins for tyrosine kinase activity using peptide substrates.
- Adsorption to monoclonal anti-phosphotyrosine antibody columns and elution with phenyl phosphate.
Main Results:
- A tyrosine kinase activity, separate from the EGF receptor, was identified.
- This novel kinase exhibited near-maximal phosphorylation within 1 minute of EGF stimulation (ED50 = 2.5 nM).
- The kinase was deactivated by CD45 tyrosine phosphatase but reactivated by purified EGF receptor, Mn2+, and ATP.
Conclusions:
- These findings suggest a novel tyrosine kinase signaling cascade initiated by EGF receptor activation.
- This cascade is analogous to established serine/threonine kinase pathways.
- The results highlight a complex interplay between receptor tyrosine kinases and other cellular kinases.