The phosphatidylinositol 3-kinase inhibitor LY294002 binds the estrogen receptor and inhibits 17beta-estradiol-induced transcriptional activity of an estrogen sensitive reporter gene.
Estrogen receptors (ERs) are members of the superfamily of ligand-activated transcription factors. In addition to the classical, hormone-mediated activation, ERs may alternatively be activated in a ligand-independent manner by a variety of agents including growth factors, neurotransmitters and cAMP. It has been demonstrated that the phosphatidylinositol 3 (PI3)-dependent kinase/Akt pathway may activate the ER alpha by increasing the activity of both estrogen independent activation function-1 and estrogen-dependent activation function-2 domains. The Akt phosphorylation site in the ER is Ser167. Phosphorylation of this residue is inhibited by LY294002, which blocks the PI3-kinase/Akt pathway. In the course of studies examining the effects of LY294002 on ligand-independent activation of ERs in L cells, we found that LY294002 exhibits antiestrogenic effects in a dose-dependent manner. By competition binding assays, we found that LY294002 specifically displaced radiolabelled estradiol from ERs with an IC(50) of 11+/-0.06 nM, being an estradiol competitor as effective as the antiestrogens ICI182,780 (IC(50), 21+/-0.13) and 4-OH-tamoxifen (IC(50), 15+/-0.09). Further, LY294002 irreversibly blocked estrogen-induced transactivation of an estradiol-sensitive reporter gene. These findings are of particular importance in the interpretation of studies demonstrating ERs inactivation by the PI3-kinase inhibitor. Our studies show that an apparent block of ER activation cannot be dissociated from inhibition of ligand-mediated events. Thus, this effect can be the result of the ability of LY294002 to bind the ERs and inhibit transactivation of estrogen-regulated genes.
Pasapera Limón
A M
AM
Research Unit in Reproductive Medicine, Hospital de Gineco Obstetricia Luis Castelazo Ayala, Instituto Mexicano del Seguro Social, Mexico, D.F., Mexico. annapl@servidor.unam.mx
Herrera-Muñoz
J
J
Gutiérrez-Sagal
R
R
Ulloa-Aguirre
A
A
D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002478:Cells, Cultured; D002867:Chromones / Q000378:metabolism / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D004958:Estradiol / Q000494:pharmacology*; D005260:Female; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D007739:L Cells; D051379:Mice; D009025:Morpholines / Q000378:metabolism / Q000494:pharmacology*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010957:Plasmids / Q000235:genetics; D011725:Pyridines / Q000494:pharmacology; D011960:Receptors, Estrogen / Q000378:metabolism*; D013449:Sulfonamides; D013629:Tamoxifen / Q000031:analogs & derivatives* / Q000494:pharmacology; D014158:Transcription, Genetic
D002867:Chromones; D004791:Enzyme Inhibitors; D007093:Imidazoles; D007546:Isoquinolines; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011725:Pyridines; D011960:Receptors, Estrogen; D013449:Sulfonamides; D013629:Tamoxifen; C016601:afimoxifene; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D004958:Estradiol; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; C093642:SB 203580