Phosphorylation of human estrogen receptor alpha by protein kinase A regulates dimerization

Phosphorylation provides an important mechanism by which transcription fact or activity is regulated. Estrogen receptor or (ER alpha) is phosphorylated on multiple sites, and stimulation of a number of growth factor receptors and/or protein kinases leads to ligand-independent and/or synergistic incre ase in transcriptional activation by ER alpha in the presence of estrogen, Here we show that ER alpha is phosphorylated by protein kinase A (PKA) on s erine-236 within the DNA binding domain. Mutation of serine-236 to glutamic acid prevents DNA binding by inhibiting dimerization by ER alpha, whereas mutation to alanine has little effect on DNA binding or dimerization. Furth ermore, PKA overexpression or activation of endogenous PKA inhibits dimeriz ation in the absence of ligand, This inhibition is overcome by the addition of 17 beta-estradiol or the partial agonist 4-hydroxy tamoxifen. Interesti ngly, treatment with the complete antagonist ICI 182,780 does not overcome the inhibitory effect of PKA activation. Our results indicate that in the a bsence of ligand ER alpha forms dimers through interaction between DNA bind ing domains and that dimerization mediated by the ligand binding domain onl y occurs upon ligand binding but that the complete antagonist ICI 182,780 p revents dimerization through the ligand-binding domain. Heterodimer formati on between ER alpha and ER beta is similarly affected by PKA phosphorylatio n of serine 236 of ER alpha, However, 4-hydroxytamoxifen is unable to overc ome inhibition of dimerization by PKA. Thus, phosphorylation of ER alpha in the DNA binding domain provides a mechanism by which dimerization and ther eby DNA binding by the estrogen receptor is regulated.