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Papers In Press, published online ahead of print October 22, 2001
J. Biol. Chem, 10.1074/jbc.M109014200
Submitted on September 18, 2001
Revised on October 16, 2001
Accepted on October 22, 2001

c-Myc mediates activation of the cad promoter via a post-RNA polymerase II recruitment mechanism

Scott R. Eberhardy and Peggy J. Farnham

McArdle Laboratory for Cancer Research, University of Wisconsin Medical School, Madison, WI 53706

Corresponding Author: farnham{at}oncology.wisc.edu

The c-Myc protein is a site-specific DNA binding transcription factor that is upregulated in a number of different cancers. We have previously shown that binding of Myc correlates with increased transcription of the cad promoter. We have now further investigated the mechanism by which Myc mediates transcriptional activation of the cad gene. Using a chromatin immunoprecipitation assay, we found high levels of RNA polymerase II bound to the cad promoter in quiescent NIH 3T3 cells and in differentiated U937 cells, even though the promoter is inactive. However, chromatin immunoprecipitation with an antibody that recognizes the hyperphosphorylated form of the RNA polymerase II carboxy terminal domain (CTD) revealed that phosphorylation of the CTD does correlate with c-Myc binding and cad transcription. We have also found that the c-Myc transactivation domain interacts with cdk9 and cyclin T1, components of the CTD kinase P-TEFb. Furthermore, activator bypass experiments have shown that direct recruitment of cyclin T1 to the cad promoter can substitute for c-Myc to activate the promoter. In summary, our results suggest that c-Myc activates transcription of cad by stimulating promoter clearance and elongation, perhaps via recruitment of P-TEFb.


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