Phosphorylation regulates Myc expression via prolonged activation of the mitogen-activated protein kinase pathway
Article first published online: 4 APR 2006
Copyright © 2006 Wiley-Liss, Inc.
Journal of Cellular Physiology
Volume 208, Issue 1, pages 133–140, July 2006
How to Cite
Wang, Z., Ge, L., Wang, M. and Carr, B. I. (2006), Phosphorylation regulates Myc expression via prolonged activation of the mitogen-activated protein kinase pathway. J. Cell. Physiol., 208: 133–140. doi: 10.1002/jcp.20649
- Issue published online: 21 APR 2006
- Article first published online: 4 APR 2006
- Manuscript Accepted: 6 FEB 2006
- Manuscript Received: 27 SEP 2005
- This work is supported in part by NIH grant CA 82723 (B.I.C)
We previously showed that prolonged and strong ERK phosphorylation induced by Compound 5 (Cpd 5), a Cdc25A protein phosphatase inhibitor, was involved in its mechanism of cell growth inhibition. To study the relationship between ERK phosphorylation and cell growth inhibition, we used Cpd 5 as a tool to investigate ERK-regulated c-Myc expression in Hep3B hepatoma cells. We found that ERK phosphorylation caused by Cpd 5 induced c-Myc phosphorylation, but suppressed c-Myc expression at the mRNA and protein levels. Furthermore, Cpd 5 inhibited c-Myc transcriptional activity and DNA binding ability, and this inhibition was antagonized by ERK kinase (MEK) inhibitor U-0126, implying that the ERK pathway was involved in regulating c-Myc expression. Since the participation of c-Myc protein in transcription requires its dimerization with Max protein, we examined the Myc-Max association in Cpd 5-treated cells and found that Cpd 5 suppressed Myc-Max dimerization. Transfection of Hep3B cells with mutated ERK (T188A/Y190F), which has lost its dual-phosphorylation sites, attenuated the actions of Cpd 5 on Myc-Max association. To further demonstrate whether Myc phosphorylation by Cpd 5-induced ERK activation was able to directly regulate c-myc gene expression, a chromatin immunoprecipitation (ChIP) assay was used to examine the binding of phospho-Myc to the c-myc promoter region. We found that phospho-Myc induced by Cpd 5 had lost its ability to bind to the c-myc promoter, whereas MEK inhibitor U-0126 antagonized this inhibitory effect. These data suggest that an increase in c-Myc phosphorylation in response to prolonged ERK phosphorylation negatively auto-regulates c-Myc gene expression, leading to the suppression of its target gene expression and cell cycle block. J. Cell. Physiol. © 2006 Wiley-Liss, Inc.