Related Experiment Videos
Regulation of stability and synthesis of EGF-receptor mRNAs encoding for intact and truncated receptor forms
1Department of Biochemistry & Biophysics, University of Pennsylvania School of Medicine, Philadelphia 19104.
Abstract:
The EGF-receptor is a proto-oncogene encoded membrane protein related to the v-erb-B oncogene product of avian erythroblastosis virus. Here we have studied the expression and stability characteristics of two forms of receptor mRNA--a 5.6 kb form that encodes for the intact transmembrane 170 kDa receptor and a 3' deleted 2.6 kb form that encodes for a 100 kDa secreted EGF-receptor. In previous studies we showed that EGF increased the synthesis of its own receptor in human cytotrophoblasts and A431 cells. Now we show that EGF stimulates the transcription of the receptor gene and blocks the degradation of transcripts, and thus elevates the level of receptor mRNA in these cells. To test the role of prior protein synthesis in these phenomena, we examined the effect of cycloheximide, and found that this reagent, either alone or in combination with EGF, leads to an increase in the level of EGF-receptor mRNA. Increases were seen in both size classes of receptor mRNA. Further analysis showed that cycloheximide enhances both synthesis and stability of receptor mRNA. While the intrinsic stability of the 3'-truncated 2.6 kb mRNA is approximately 2-fold higher than that of the 5.6 kb species, it is enhanced by more than 10-fold in the presence of cycloheximide and EGF acting in a synergistic manner. The extent of stabilization was less for the larger mRNA form. The results suggest the involvement of ligand as well as labile proteins in the control of synthesis and stability of EGF-receptor mRNA, and imply the involvement of 3' located structural features in the control of receptor mRNA stability.
Insights
Epidermal Growth Factor (EGF) receptor gene expression is boosted by EGF, which stimulates transcription and blocks mRNA degradation. Cycloheximide further increases EGF receptor mRNA levels by enhancing both synthesis and stability.
Area of Science:
- Molecular Biology
- Cell Biology
- Oncology
Background:
- The epidermal growth factor (EGF) receptor is a proto-oncogene-encoded membrane protein.
- Two forms of EGF receptor mRNA exist: a 5.6 kb form encoding the intact receptor and a 3' deleted 2.6 kb form encoding a secreted receptor.
- Previous studies showed EGF increases its own receptor synthesis in human cytotrophoblasts and A431 cells.
Purpose of the Study:
- To investigate the mechanisms by which EGF affects EGF receptor mRNA levels.
- To determine the role of protein synthesis in EGF-induced changes in EGF receptor mRNA.
- To analyze the differential stability of EGF receptor mRNA forms.
Main Methods:
- Studied expression and stability of 5.6 kb and 2.6 kb EGF receptor mRNA forms.
- Investigated the effects of EGF and cycloheximide on receptor gene transcription and mRNA degradation.
- Analyzed the impact of cycloheximide on mRNA synthesis and stability.
Main Results:
- EGF stimulates EGF receptor gene transcription and blocks mRNA degradation, increasing mRNA levels.
- Cycloheximide, alone or with EGF, increases both size classes of EGF receptor mRNA.
- Cycloheximide enhances both synthesis and stability of EGF receptor mRNA; the 2.6 kb form shows synergistic stabilization with EGF and cycloheximide.
Conclusions:
- Ligand (EGF) and labile proteins regulate the synthesis and stability of EGF receptor mRNA.
- 3' located structural features likely play a role in controlling receptor mRNA stability.
- Differential mRNA stability contributes to the regulation of EGF receptor expression.