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Glycogen Synthase Kinase-3 Protects Estrogen Receptor from Proteasomal Degradation and Is Required for Full Transcriptional Activity of the Receptor

Hormones and Signal Transduction Group, German Cancer Research Center, 69120 Heidelberg, Germany

Address all correspondence and requests for reprints to: Doris Mayer, Hormones and Signal Transduction Group, German Cancer Research Center, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany. E-mail: d.mayer{at}dkfz.de.

Glycogen synthase kinase-3 (GSK-3) plays a key role in the regulation of transcription factors including steroid receptors. Having identified estrogen receptor- (ER) as substrate for GSK-3, the impact of GSK-3 on ER function and activity upon 17ß-estradiol (E2)-dependent activation remains to be clarified. Here we show by using small interfering technology in combination with immunoblot, gene expression analysis, and luciferase reporter assays that silencing of GSK-3 or GSK-3ß results in the reduction of ER levels and transcriptional activity in ER-positive breast cancer cells. Using MCF-7 cells we demonstrate that reduction of ER levels upon GSK-3 silencing was due to increased proteasomal degradation of ER rather than inhibition of ER protein synthesis. Indeed, under this condition, ER protein was rescued using the proteasome inhibitor MG132 in presence of the protein synthesis inhibitor cycloheximide. In addition, strong accumulation of ubiquitinated ER was obtained after GSK-3 silencing in the presence of MG132. We conclude that GSK-3 protects ER from proteasomal degradation and plays a crucial role in ER protein stabilization and turnover. Furthermore, in vitro kinase assay depicted that GSK-3ß phosphorylates ER at Ser-118. GSK-3 silencing resulted in decrease of E2-induced nuclear ER phosphorylation at Ser-118 and E2-induced estrogen response element-dependent luciferase reporter gene expression. Neither Ser-118 phosphorylation nor luciferase activity was restored by use of MG132. Moreover, the expression of estrogen-responsive genes (pS2 and progesterone receptor) was decreased upon GSK-3 silencing. These findings demonstrated that GSK-3 is required for E2-induced ER phosphorylation at Ser-118 and full transcriptional activity of the receptor upon E2 stimulation.

NURSA Molecule Pages Link:

Nuclear Receptors:   ERα

Ligands:   17β-Estradiol

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