THE ESTROGEN-RESPONSIVE ELEMENT OF THE PS2 GENE IS RECOGNIZED BY A METHYLATION SENSITIVE DNA-BINDING PROTEIN

The hu man pS2 gene is specifically expressed is a subclass of estroge n receptor containing human breast cancer cells. In the MCF7 cell line , its induction by estradiol is a primary transcriptional event. The e xact location of its estrogen responsive element has been determined u sing a chimeric recombinant transfected into HeLa cells and a transien t expression assay. In this study we found, using electrophoretic mobi lity shift experiments, that in HeLa cells the estrogen responsive ele ment (ERE) of the pS2 gene is recognized by a methylation sensitive DN A binding protein (MSDBP) different from the estrogen receptor. Compet ition experiments have shown that the binding of this protein requires at least one CpG in the center of the palindromic sequence and that i mperfect palindromic sequences are also recognized. Although the prese nce of CpG is necessary, CpG-rich oligonucleotides, containing consens us sequences for Spl or AP2, do not interfere with its binding to the pS2 oligonucleotide, indicating that the ERE sequence itself participa tes in the specificity of its binding. This protein binds the pS2 sequ ence with a relatively high affinity (apparent K-d = 10(-10) M) and it s binding is strongly reduced by the methylation of the cytosines at C pG sites. UV cross-linking experiments and peptide mapping indicate th at this protein has an apparent molecular weight of 46 kDa and is pres ent in several cell lines, including non-human cell lines. Taken toget her, these data suggest that this protein might have a potential role in regulating gene activity or in chromatin structure of some genes po ssessing an ERE.