TY - JOUR
T1 - Analysis of estrogen receptor α-Sp1 interactions in breast cancer cells by flourescence resonance energy transfer
AU - Kim, Kyounghyun
AU - Barhoumi, Rola
AU - Burghardt, Robert
AU - Safe, Stephen
N1 - Copyright:
Copyright 2008 Elsevier B.V., All rights reserved.
PY - 2005/4
Y1 - 2005/4
N2 - Estrogen-dependent regulation of several genes associated with cell cycle progression, proliferation, and nucleotide metabolism in breast cancer cells is associated with interactions of estrogen receptor (ER)α/Sp1 with GC-rich promoter elements. This study investigates ligand-dependent interactions of ERα and Sp1 in MCF-7 breast cancer cells using fluorescence resonance energy transfer (FRET). Chimeric ERα and Sp1 proteins fused to cyan fluorescent protein or yellow fluorescent protein were transfected into MCF-7 cells, and a FRET signal was induced after treatment with 170-estradiol, 4′-hydroxyitamoxifen, or ICI 182,780. Induction of FRET by these ERα agonists/antagonists was paralleled by their activation of gene expression in cells transfected with a construct (pSp13) containing three tandem Sp1 binding sites linked to a luciferase reporter gene. In contrast, interactions between ERα and Sp1ΔDBD [a DNA binding domain (DBD) deletion mutant of Sp1] are not observed, and this is consistent with the critical role of the C-terminal DBD of Sp1 for interaction with ERα. Results of the FRET assay are consistent with in vitro studies on ERα/Sp1 interactions and transactivation, and confirm that ERα and Sp1 interact in living breast cancer cells.
AB - Estrogen-dependent regulation of several genes associated with cell cycle progression, proliferation, and nucleotide metabolism in breast cancer cells is associated with interactions of estrogen receptor (ER)α/Sp1 with GC-rich promoter elements. This study investigates ligand-dependent interactions of ERα and Sp1 in MCF-7 breast cancer cells using fluorescence resonance energy transfer (FRET). Chimeric ERα and Sp1 proteins fused to cyan fluorescent protein or yellow fluorescent protein were transfected into MCF-7 cells, and a FRET signal was induced after treatment with 170-estradiol, 4′-hydroxyitamoxifen, or ICI 182,780. Induction of FRET by these ERα agonists/antagonists was paralleled by their activation of gene expression in cells transfected with a construct (pSp13) containing three tandem Sp1 binding sites linked to a luciferase reporter gene. In contrast, interactions between ERα and Sp1ΔDBD [a DNA binding domain (DBD) deletion mutant of Sp1] are not observed, and this is consistent with the critical role of the C-terminal DBD of Sp1 for interaction with ERα. Results of the FRET assay are consistent with in vitro studies on ERα/Sp1 interactions and transactivation, and confirm that ERα and Sp1 interact in living breast cancer cells.
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U2 - 10.1210/me.2004-0326
DO - 10.1210/me.2004-0326
M3 - Article
C2 - 15637147
AN - SCOPUS:15544365184
VL - 19
SP - 843
EP - 854
JO - Molecular Endocrinology
JF - Molecular Endocrinology
SN - 0888-8809
IS - 4
ER -