Expression of DNA methyl-transferase (DMT) and the cell cycle in human breast cancer cells

Estrogen receptor (ER)-negative breast cancer cells display extensive methy lation of the ER gene CpG island and elevated DNA methyltransferase (DMT) e xpression compared to ER-positive cells. The present study demonstrates tha t DMT protein levels tightly correlate with S phase fraction in ER-positive cells, whereas ER-negative cells express DMT throughout the cell cycle. In addition, levels of p21(CIP1), which disrupts DMT binding to PCNA, are inv ersely correlated with DMT levels, Therefore increased DMT expression in ER -negative cells is not simply due to elevated S-phase fraction, but rather to more complex changes that allow cells to escape normal cell cycle-depend ent controls on DMT expression. Because ER-negative breast tumors often hav e activated growth factor pathways, the impact of these pathways on DMT exp ression was examined in ER-positive cells. Stable transfection with fibrobl ast growth factors (FGFs) 1 and 4 led to increased DMT expression that coul d not be accounted for by a shift in S phase fraction. Elevated DMT protein expression in FGF-transfectants was accompanied by a significant decrease in p21, again suggesting a reciprocal relationship between these two protei ns. However, acquisition of an estrogen-independent phenotype, even in conj unction with elevated DMT levels, was not sufficient to promote ER gene sil encing via methylation, These results indicate that multiple steps are requ ired for de novo methylation of the ER CpG island.