Acetylation of estrogen receptor alpha by p300 at lysines 266 and 268 enhances the deoxyribonucleic acid binding and transactivation activities of the receptor.
Using a variety of biochemical and cell-based approaches, we show that estrogen receptor alpha (ERalpha) is acetylated by the p300 acetylase in a ligand- and steroid receptor coactivator-dependent manner. Using mutagenesis and mass spectrometry, we identified two conserved lysine residues in ERalpha (Lys266 and Lys268) that are the primary targets of p300-mediated acetylation. These residues are acetylated in cells, as determined by immunoprecipitation-Western blotting experiments using an antibody that specifically recognizes ERalpha acetylated at Lys266 and Lys268. The acetylation of ERalpha by p300 is reversed by native cellular deacetylases, including trichostatin A-sensitive enzymes (i.e. class I and II deacetylases) and nicotinamide adenine dinucleotide-dependent/nicotinamide-sensitive enzymes (i.e. class III deacetylases, such as sirtuin 1). Acetylation at Lys266 and Lys268, or substitution of the same residues with glutamine (i.e. K266/268Q), a residue that mimics acetylated lysine, enhances the DNA binding activity of ERalpha in EMSAs. Likewise, substitution of Lys266 and Lys268 with glutamine enhances the ligand-dependent activity of ERalpha in a cell-based reporter gene assay. Collectively, our results implicate acetylation as a modulator of the ligand-dependent gene regulatory activity of ERalpha. Such regulation is likely to play a role in estrogen-dependent signaling outcomes in a variety of estrogen target tissues in both normal and pathological states.
Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA.
Mi Young
MY
Kim
Eileen M
EM
Woo
Yee Ting Esther
YT
Chong
Daria R
DR
Homenko
W Lee
WL
Kraus
D000107:Acetylation; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002415:Cats; D002451:Cell Compartmentation; D002478:Cells, Cultured; D017124:Conserved Sequence; D006224:Cricetinae; D004247:DNA / Q000378:metabolism*; D047628:Estrogen Receptor alpha / Q000378:metabolism*; D047629:Estrogen Receptor beta / Q000378:metabolism; D004967:Estrogens / Q000378:metabolism; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D008239:Lysine / Q000378:metabolism*; D013058:Mass Spectrometry; D051379:Mice; D008969:Molecular Sequence Data; D009536:Niacinamide / Q000494:pharmacology; D051958:Nuclear Receptor Coactivator 2 / Q000378:metabolism; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D051381:Rats; D016415:Sequence Alignment; D037761:Sirtuins / Q000378:metabolism; D015533:Transcriptional Activation; D050880:p300-CBP Transcription Factors / Q000378:metabolism*
D047628:Estrogen Receptor alpha; D047629:Estrogen Receptor beta; D004967:Estrogens; D006877:Hydroxamic Acids; D051958:Nuclear Receptor Coactivator 2; D009536:Niacinamide; C012589:trichostatin A; D004247:DNA; D050880:p300-CBP Transcription Factors; D037761:Sirtuins; D008239:Lysine