Akt stabilizes estrogen receptor α with the concomitant reduction in its transcriptional activity

Abstract

We have investigated the effect of Akt on estrogen receptor (ER) α protein level and its transcriptional activity. Transient transfection studies revealed that constitutively active Akt1 up-regulated ERα at the post-transcriptional level. Studies using Akt inhibitor and dominant-negative Akt1 showed that Akt1 kinase activity is required for the up-regulation of ERα. Cycloheximide decay assays and studies with proteasome inhibitor indicated that Akt1-mediated up-regulation of ERα was maintained by inhibiting proteasome-mediated degradation of ERα. When Akt consensus phosphorylation site mutant, ERαS167A was tested for Akt1-mediated up-regulation, increase of ERαS167A by Akt1 was significantly impaired as compared to wild type ERα. In addition, dominant-negative glycogen synthase kinase (GSK) 3β and LiCl could also partially up-regulate ERα protein level, suggesting that concerted action of Akt1-mediated phosphorylation on S167 and kinase activity of Akt-downstream GSK3β could affect ERα protein level. Paradoxically, co-expression of Akt1 could down-regulate transcriptional activity of ERα. The inhibitory effect of Akt1 on ERα transcriptional activity was not attributable to changes in subcellular distribution of ERα. Transfection studies using increasing amount of Akt1 and ERα indicated that the transcriptional activity of ERα was negatively regulated by ERα protein quantities at higher ERα concentrations. Chromatin immunoprecipitation assays revealed that at Akt1 concentration high enough to induce up-regulation of ERα, association of ERα to promoter region of ERα target pS2 gene was impaired. Taken together, these data suggest that Akt1 could increase ERα protein level with simultaneous reduction in its transcriptional activity, possibly by modulating association of ERα to the target gene promoters.