Transcriptional regulation of the glutathione S-transferase P1 gene

Glutathione S-transferases (GSTs) are thought to play a critical role in cellular protection. GSTs are known to neutralize exogenous toxins by catalyzing their conjugation to glutathione. Of particular interest is the human {dollar}pi{dollar} class GST (GSTP1-1). Increased expression of GSTP1 has been observed in numerous human tumors, and is implicated in the development of drug resistance. Consequently, the goal of this thesis is to examine the mechanisms that regulate the expression of GSTP1. To investigate these processes, we have employed model breast cancer cell lines that express (ER{dollar}-{dollar}) and do not express (ER+) GSTP1.; Previously, we showed that authentic full-length GSTP1 mRNA sequences are less stable in ER+, than in ER{dollar}-{dollar} cells. However, mRNA stability or other posttranscriptional processes cannot completely account for differential GSTP1 expression in these cell lines. Accordingly, we examined transcriptional mechanisms that influence GSTP1 expression. Transient transfection of several GSTP1 promoter-reporter genes and EMSA identified regions within the proximal promoter that are important for both basal and differential GSTP1 expression. In particular, a putative silencer element was shown to selectively repress expression of a transiently transfected GSTP1 promoter in ER+ cells. However, repression by the silencer is incomplete. Both transiently transfected GSTP1 promoters and a stably transfected GSTP1 minigene contain the repressor sequences and are expressed in GSTP1 non-expressing cells. We conclude that other factors are responsible for differential GSTP1 expression in breast cancer cell lines.; Previously, it was shown that methylation of GSTP1 promoter sequences is important for GSTP1 repression in GSTP1 non-expressing prostate and prostate cancer cell lines. Therefore, we examined whether differences in the methylation status of the GSTP1 CpG island are associated with cell line-specific expression of GSTP1 in model breast cancer cells. Our studies revealed that GSTP1 is hypermethylated in ER+ cells, but undermethylated in ER{dollar}-{dollar} cells. Moreover, partial demethylation of the GSTP1 CpG island resulted in de novo GSTP1 mRNA and protein synthesis in ER+ cell lines. These data strongly suggest that the methylation status of the GSTP1 promoter contributes significantly to the levels of GSTP1 expressed in ER{dollar}-{dollar} and ER+ breast cancer cells lines.