The effects of histone acetylation on estrogen responsiveness in MCF-7 cells

Because histone acetylation is implicated in the facilitation of specific g ene transcription, the effect of increasing histone acetylation on the expr ession of an endogenous gene was investigated. The ability of trichostatin A (TSA), a histone deacetylase inhibitor, to potentiate the estradiol (E-2) induction of progesterone receptor (PR) levels in MCF-7 cells was studied. Although TSA alone had no effect on PR synthesis, measured by a whole-cell binding assay with [H-3]R5020, TSA potentiated the effect of 10-(11) M E-2 such that 10 ng of TSA/mL approximately doubled the hormone response, When TSA was removed from the cells after various incubation times (24 and 48 h ) by successive washings with TSA-free medium, it was determined that TSA w as required through out the 96-h incubation period in order to achieve maxi mal potentiation for the E-2 response. In addition, TSA potentiated E-2 ind uction of pS2 mRNA, These results suggested that the estrogen receptor (ER) complex might alter histone acetylation as part of the gene activation pro cess. To test this directly, MCF-7 cells were incubated for 48 h with E-2 f ollowed by incubation with sodium [H-3]acetate for 1 h, From two-dimensiona l polyacrylamide gel electrophoresis, an increase in total acetate incorpor ation into histones in estrogen- treated cells compared to control was obse rved as well as a preferential increase in the mono- and diacetylated histo ne H4. Experiments with lysine-specific antiacetylated H4 antibodies sugges t a preferential increase in acetylation at lysine 16, but not 5, 8, or 12. The results of this study support an important role for histone acetylatio n in the mechanism of action of the ER.