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Phosphorylation and activation of epidermal growth factor receptors in cells transformed by the src oncogene
W J Wasilenko1, D M Payne, D L Fitzgerald
1Department of Microbiology, University of Virginia School of Medicine, Charlottesville 22908.
Abstract:
Because functionally significant substrates for the tyrosyl protein kinase activity of pp60v-src are likely to include membrane-associated proteins involved in normal growth control, we have tested the hypothesis that pp60v-src could phosphorylate and alter the signaling activity of transmembrane growth factor receptors. We have found that the epidermal growth factor (EGF) receptor becomes constitutively phosphorylated on tyrosine in cells transformed by the src oncogene and in addition displays elevated levels of phosphoserine and phosphothreonine. High-performance liquid chromatography phosphopeptide mapping revealed two predominant sites of tyrosine phosphorylation, both of which differed from the major sites of receptor autophosphorylation; thus, the src-induced phosphorylation is unlikely to occur via an autocrine mechanism. To determine whether pp60v-src altered the signaling activity of the EGF receptor, we analyzed the tyrosine phosphorylation of phospholipase C-gamma, since phosphorylation of this enzyme occurs in response to activation of the EGF receptor but not in response to pp60v-src alone. We found that in cells coexpressing pp60v-src and the EGF receptor, phospholipase C-gamma was constitutively phosphorylated, a result we interpret as indicating that the signaling activity of the EGF receptor was altered in the src-transformed cells. These findings suggest that pp60v-src-induced alterations in phosphorylation and function of growth regulatory receptors could play an important role in generating the phenotypic changes associated with malignant transformation.
Insights
The src oncogene alters epidermal growth factor (EGF) receptor signaling by causing constitutive tyrosine phosphorylation. This impacts cell growth control and may contribute to malignant transformation.
Area of Science:
- Cellular Biology
- Oncology
- Molecular Biology
Background:
- The pp60v-src protein is a tyrosyl protein kinase implicated in cell growth control.
- Membrane-associated proteins, including growth factor receptors, are potential substrates for pp60v-src.
- Understanding pp60v-src's interaction with growth factor receptors is crucial for comprehending malignant transformation.
Purpose of the Study:
- To investigate if pp60v-src phosphorylates and alters the signaling activity of transmembrane growth factor receptors.
- Specifically, to examine the effect of pp60v-src on the epidermal growth factor (EGF) receptor.
Main Methods:
- Cells transformed by the src oncogene were analyzed for EGF receptor phosphorylation.
- High-performance liquid chromatography phosphopeptide mapping was used to identify tyrosine phosphorylation sites.
- Phosphorylation of phospholipase C-gamma was assessed to evaluate EGF receptor signaling activity.
Main Results:
- EGF receptor exhibited constitutive tyrosine phosphorylation in src-transformed cells, alongside elevated phosphoserine and phosphothreonine.
- Src-induced tyrosine phosphorylation sites on the EGF receptor differed from its autophosphorylation sites.
- Phospholipase C-gamma was constitutively phosphorylated in cells coexpressing pp60v-src and EGF receptor, indicating altered receptor signaling.
Conclusions:
- pp60v-src directly phosphorylates the EGF receptor at distinct sites, altering its signaling capacity.
- The src-induced alterations in EGF receptor phosphorylation and function contribute to phenotypic changes observed in malignant transformation.
- pp60v-src-mediated modification of growth regulatory receptors plays a significant role in oncogenesis.