title: Estrogen receptor-α mediates human multidrug resistance associated protein 3 induction by 17α-ethynylestradiol. Role of activator protein-1.
abstract: Previously, we have demonstrated that 17α-ethynylestradiol (EE) induces rat multidrug-resistance associated protein 3 (Mrp3, Abcc3) expression transcriptionally through estrogen receptor-α (ER-α) activation. We explored the effect of EE on MRP3 expression of human origin. HepG2 cells were transfected with ER-α and incubated with EE (1-10-50 μM) for 48 h. MRP3 protein and mRNA levels were measured by Western blotting and Real time PCR, respectively. EE up-regulated MRP3 protein and mRNA at 50 μM only in ER-α(+)-HepG2 cells. The in silico analysis of mrp3 promoter region demonstrated absence of estrogen response elements, but showed several Ap-1 binding sites. We further evaluated the potential involvement of the transcription factors c-JUN and c-FOS (members of Ap-1) in MRP3 up-regulation. ER-α(+) HepG2 cells were incubated with EE and c-FOS and c-JUN levels measured by Western blotting in nuclear extracts. EE up-regulated only c-JUN. Experiments of overexpression and knock-down of c-JUN by siRNA further demonstrated that this transcription factor is indeed implicated in MRP3 upregulation by EE. Co-immunoprecipitation assay demonstrated that EE induces c-JUN/ER-α interaction, and chromatin immunoprecipitation assay showed that this complex is recruited to the AP-1 binding consensus element present at the position (-1300/-1078 bp) of human mrp3 promoter. We conclude that EE induces MRP3 expression through ER-α, with recruitment of ER-α in complex with c-JUN to the human mrp3 promoter.
authors.affiliation: Instituto de Fisiología Experimental (CONICET) - Facultad de Ciencias Bioquímicas y Farmacéuticas (UNR), Suipacha 570, (2000) Rosario, Argentina. ruiz@ifise-conicet.gov.ar
authors.forename: María Laura
authors.initials: ML
authors.lastname: Ruiz
authors.forename: Juan Pablo
authors.initials: JP
authors.lastname: Rigalli
authors.forename: Agostina
authors.initials: A
authors.lastname: Arias
authors.forename: Silvina Stella Maris
authors.initials: SS
authors.lastname: Villanueva
authors.forename: Claudia
authors.initials: C
authors.lastname: Banchio
authors.forename: Mary
authors.initials: M
authors.lastname: Vore
authors.forename: Aldo Domingo
authors.initials: AD
authors.lastname: Mottino
authors.forename: Viviana Alicia
authors.initials: VA
authors.lastname: Catania
mesh_terms: D001483:Base Sequence; D047628:Estrogen Receptor alpha / Q000502:physiology*; D004997:Ethinyl Estradiol / Q000494:pharmacology*; D056945:Hep G2 Cells; D006801:Humans; D008969:Molecular Sequence Data; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000096:biosynthesis* / Q000235:genetics; D018808:Transcription Factor AP-1 / Q000502:physiology*
chemical_list: D047628:Estrogen Receptor alpha; D004997:Ethinyl Estradiol; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D018808:Transcription Factor AP-1; C506487:ESR1 protein, human; C114462:multidrug resistance-associated protein 3
keywords: AP-1; Drug transporter; Estrogen; Multidrug resistance; Nuclear receptor; c-JUN.