Dataset Information

Acetylation of estrogen receptor alpha by p300 at lysines 266 and 268 enhances the deoxyribonucleic acid binding and transactivation activities of the receptor.

ABSTRACT: 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.

SUBMITTER: Kim MY

PROVIDER: S-EPMC1483068 | biostudies-literature | 2006 Jul

REPOSITORIES: biostudies-literature

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Acetylation of estrogen receptor alpha by p300 at lysines 266 and 268 enhances the deoxyribonucleic acid binding and transactivation activities of the receptor.

Kim Mi Young MY Woo Eileen M EM Chong Yee Ting Esther YT Homenko Daria R DR Kraus W Lee WL

Molecular endocrinology (Baltimore, Md.) 20060223 7

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 antib ...[more]

PMID: 16497729

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Project description:Cellular levels of estrogen receptor-alpha (ERalpha) protein are regulated primarily by the ubiquitin-proteasome pathway. Dynamic interactions between ERalpha and the protein degradation machinery facilitate the down-regulation process by targeting receptor lysine residues for polyubiquitination. To date, the lysines that control receptor degradation have not been identified. Two receptor lysines, K302 and K303, located in the hinge-region of ERalpha, serve multiple regulatory functions, and we examined whether these might also regulate receptor polyubiquitination, turnover, and receptor-protein interactions. We used ERalpha-negative breast cancer C4-12 cells to generate cells stably expressing wild-type (wt)ERalpha or ERalpha with lysine-to-alanine substitutions at K302 and K303 (ERalpha-AA). In the unliganded state, ERalpha-AA displayed rapid polyubiquitination and enhanced basal turnover, as compared with wtERalpha, due to its elevated association with the ubiquitin ligase carboxy terminus of Hsc70-interacting protein (CHIP) and the proteasome-associated cochaperone Bag1. Treatment of C4-12 cells with either 17beta-estradiol (E2) or the pure antiestrogen ICI 182,780 (ICI) induced rapid degradation of wtERalpha via the ubiquitin-proteasome pathway; however, in the presence of these ligands, ERalpha-AA was less efficiently degraded. Furthermore, ERalpha-AA was resistant to ICI-induced polyubiquitination, suggesting that these lysines are polyubiquitinated in response to the antiestrogen and demonstrate a novel role for these two lysines in the mechanism of action of ICI-induced receptor down-regulation. The reduced stability of ERalpha-AA in the unliganded state and the increased stability of ERalpha-AA in the liganded state were concordant with reporter gene assays demonstrating that ERalpha-AA has lower basal activity but higher E2 inducibility than wtERalpha. These data provide the first evidence that K302/303 protect ERalpha from basal degradation and are necessary for efficient E2- and ICI-induced turnover in breast cancer cells.

Dataset Information

Acetylation of estrogen receptor alpha by p300 at lysines 266 and 268 enhances the deoxyribonucleic acid binding and transactivation activities of the receptor.

Publications

Acetylation of estrogen receptor alpha by p300 at lysines 266 and 268 enhances the deoxyribonucleic acid binding and transactivation activities of the receptor.

Similar Datasets

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