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Papers In Press, published online ahead of print July 6, 2001
J. Biol. Chem, 10.1074/jbc.M103951200
Submitted on May 2, 2001
Revised on June 28, 2001
Accepted on July 5, 2001

5 aza C treatment enhances expression of TGF-beta receptors through down-regulation of Sp3

Sudhakar Ammanamanchi and Michael G. Brattain

Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, NY 14263

Corresponding Author: Michael.Brattain{at}roswellpark.org

We have previously reported that Sp3 acts as a transcriptional repressor of TGF-beta receptors type I (RI) and type II (RII). We now present data suggesting that treatment of MCF-7L breast and GEO colon cancer cells with 5 aza cytidine leads to down-regulation of Sp3 and the concomitant induction of RI and RII. Western blot and gel shift analyses on 5 aza C treated MCF-7L and GEO nuclear extracts indicated reduced Sp3 protein levels and decreased binding of Sp3 protein to radiolabeled consensus Sp1 oligonucleotide. Southwestern analysis detected decreased binding of Sp3 to RI and RII promoters in 5 aza C treated MCF-7L and GEO cells suggesting a correlation between decreased Sp3 binding and enhanced RI and RII expression in these cells. RT-PCR and nuclear run on data from 5 aza C treated MCF-7L and GEO cells indicated down-regulation of Sp3 mRNA as a result of decreased transcription of Sp3. We reported earlier that 5 aza C treatment induces RI and RII expression through increased Sp1 protein levels/activities in these cells. These studies demonstrate that the effect of 5 aza C involves a combination of effects on Sp1 and Sp3.


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