Figure 2.

(A) Downregulation of NT5E mRNA correlates with CpG methylation in ER-positive breast carcinoma cell lines. Expression of NT5E mRNA was determined by qPCR and % methylation by pyrosequencing. Downregulation correlates with methylation. (B) NT5E mRNA is overexpressed in breast carcinoma cell lines lacking CpG island methylation. Average % methylation in the NT5E CpG island was determined by pyrosequencing and expression of NT5E mRNA by qPCR (relative to normal breast epithelium). (C) Demethylation reactivates NT5E expression. MCF-7 cells were treated with AZA and harvested at the indicated times (in hours). Total RNA was isolated and levels of NT5E mRNA determined by qPCR as described in Materials and Methods.