1. ¹Laboratoire de Biologie Moléculaire Eucaryote, Université de Toulouse, Toulouse, France
2. ²UMR 5099, CNRS, Toulouse, France

Correspondence: Dr K Bystricky, Laboratoire de Biologie Moléculaire Eucaryote, Université de Toulouse, IBCG, 118 route de Narbonne, Toulouse, Midi Pyrenees 31062, France. E-mail: kerstin@ibcg.biotoul.fr

³These authors contributed equally to this work.

Received 28 August 2007; Revised 27 December 2007; Accepted 31 January 2008; Published online 3 March 2008.

Abstract

In breast cancer, approximately one-third of tumors express neither the estrogen receptor (ER) nor estrogen-regulated genes such as the progesterone receptor gene (PR). Our study provides new insights into the mechanism allowing hormone-activated expression of ER target genes silenced in ER-negative mammary tumor cells. In cell lines derived from ER-negative MDA-MB231 cells, stable expression of different levels of ER from a transgene did not result in transcription of PR. A quantitative comparative analysis demonstrates that inhibiting DNA methyltransferases using 5-aza-2'-deoxycytidine or specific disruption of DNMT1 by small interfering RNAs and treatment with the histone-deacetylase inhibitor trichostatin A enabled ER-mediated hormone-dependent expression of endogenous PR. We show that demethylation of a CpG island located in the first exon of PR was a prerequisite for ER binding to these regulatory sequences. Although not a general requirement, DNA demethylation is also necessary for derepression of a subset of ER target genes involved in tumorigenesis. PR transcription did not subsist 4 days after removal of the DNA methyltransferase blocking agents, suggesting that hormone-induced expression of ER target genes in ER-negative tumor cells is transient. Our observations support a model where an epigenetic mark confers stable silencing by precluding ER access to promoters.