Glycogen synthase kinase-3 interacts with and phosphorylates estrogen receptor alpha and is involved in the regulation of receptor activity.
Like other steroid hormone receptors, estrogen receptor-alpha (ERalpha) is a substrate for protein kinases, and phosphorylation has profound effects on the function and activity of this receptor. A number of different kinases have been implicated in ERalpha regulation. In this report we show by mutational analysis and in vitro kinase assays that ERalpha is a substrate for glycogen synthase kinase-3 (GSK-3) in vitro and is phosphorylated on two sites, the Ser-102, -104, and -106 motif and Ser-118, both located in the N-terminal transcription activation function (AF-1) domain. GSK-3 forms a complex with ERalpha in vivo as demonstrated by co-immunoprecipitation from cell lysates. The GSK-3 inhibitor lithium chloride was used to determine the role of GSK-3 in phosphorylation of Ser-102, -104, and -106 and Ser-118 in vivo and to explore the role of these serines in the regulation of ERalpha function. Treatment of cells with lithium chloride resulted in dephosphorylation of Ser-104 and -106 and Ser-118, which suggests these serine residues as targets for GSK-3 in vivo. Our results further suggest that ERalpha phosphorylation by GSK-3 stabilizes ERalpha under resting conditions and modulates ERalpha transcriptional activity upon ligand binding. Inhibition and constitutive activation of GSK-3, both, resulted in inhibition of ERalpha transcriptional activity, indicating a function of active as well as inactive GSK-3 in ERalpha regulation. These findings uncover a novel mechanism for the regulation of ERalpha-mediated estrogen signaling controlled by a dual action of GSK-3.
Medunjanin
Senad
S
Hormones and Signal Transduction Group, German Cancer Research Center, 69120 Heidelberg, Germany.
Hermani
Alexander
A
De Servi
Barbara
B
Grisouard
Jean
J
Rincke
Gabriele
G
Mayer
Doris
D
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004252:DNA Mutational Analysis; D018076:DNA, Complementary / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D047628:Estrogen Receptor alpha / Q000378:metabolism*; D005786:Gene Expression Regulation; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism*; D006801:Humans; D047468:Immunoprecipitation; D008024:Ligands; D018021:Lithium Chloride / Q000494:pharmacology; D008156:Luciferases / Q000378:metabolism; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry; D015398:Signal Transduction; D013347:Subcellular Fractions / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D047628:Estrogen Receptor alpha; D008024:Ligands; D011994:Recombinant Proteins; D012694:Serine; D008156:Luciferases; D038362:Glycogen Synthase Kinase 3; D018021:Lithium Chloride