5-AzaC treatment enhances expression of transforming growth factor-beta receptors through down-regulation of Sp3

We have previously reported that Sp3 acts as a transcriptional repressor of transforming growth factor-P receptors type I (RI) and type II (RII). We n ow present data suggesting that treatment of MCF-7L, breast and GEO colon c ancer cells with 5-aza cytidine (5-azaC) leads to down-regulation of Sp3 an d the concomitant induction of RI and RII. Western blot and gel shift analy ses on 5-azaC-treated MCF-7L and GEO nuclear extracts indicated reduced Sp3 protein levels and decreased binding of Sp3 protein to radiolabeled consen sus Sp1 oligonucleotide. Southwestern analysis detected decreased binding o f Sp3 to RI and RH promoters in 5-azaC-treated MCF-7L and GEO cells, sugges ting a correlation between decreased Sp3 binding and enhanced RI and RII ex pression in these cells. Reverse transcription-polymerase chain reaction an d nuclear run-on data from 5-azaC-treated MCF-7L and GEO cells indicated do wn-regulation of Sp3 mRNA as a result of decreased transcription of Sp3. We reported earlier that 5-azaC treatment induces RI and RII expression throu gh increased Sp1 protein levels/activities in these cells. These studies de monstrate that the effect of 5-azaC involves a combination of effects on Sp 1 and Sp3.