title: Recruitment of the NCoA/SRC-1/p160 family of transcriptional coactivators by the aryl hydrocarbon receptor/aryl hydrocarbon receptor nuclear translocator complex.
abstract: The aryl hydrocarbon receptor complex heterodimeric transcription factor, comprising the basic helix-loop-helix-Per-ARNT-Sim (bHLH-PAS) domain aryl hydrocarbon receptor (AHR) and aryl hydrocarbon receptor nuclear translocator (ARNT) proteins, mediates the toxic effects of TCDD (2,3,7,8 tetrachlorodibenzo-p-dioxin). The molecular events underlying TCDD-inducible gene activation, beyond the activation of the AHRC, are poorly understood. The SRC-1/NCoA-1, NCoA-2/GRIP-1/TIF-2, and p/CIP/AIB/ACTR proteins have been shown to act as mediators of transcriptional activation. In this report, we demonstrate that SRC-1, NCoA-2, and p/CIP are capable of independently enhancing TCDD-dependent induction of a luciferase reporter gene by the AHR/ARNT dimer. Furthermore, injection of anti-SRC-1 or anti-p/CIP immunoglobulin G into mammalian cells abolishes the transcriptional activity of a TCDD-dependent reporter gene. We demonstrate by coimmunoprecipitation and by a reporter gene assay that SRC-1 and NCoA-2 but not p/CIP are capable of interacting with ARNT in vivo after transient transfection into mammalian cells, while AHR is capable of interacting with all three coactivators. We confirm the interactions of ARNT and AHR with SRC-1 with immunocytochemical techniques. Furthermore, SRC-1, NCoA-2, and p/CIP all associate with the CYP1A1 enhancer region in a TCDD-dependent fashion, as demonstrated by chromatin immunoprecipitation assays. We demonstrate by yeast two-hybrid, glutathione S-transferase pulldown, and mammalian reporter gene assays that ARNT requires its helix 2 domain but not its transactivation domain to interact with SRC-1. This indicates a novel mechanism of action for SRC-1. SRC-1 does not require its bHLH-PAS domain to interact with ARNT or AHR, but utilizes distinct domains proximal to its p300/CBP interaction domain. Taken together, these data support a role for the SRC family of transcriptional coactivators in TCDD-dependent gene regulation.
authors.affiliation: Department of Pathology and Laboratory Medicine, Molecular Biology Institute, and Jonsson Comprehensive Cancer Center, School of Medicine, University of California-Los Angeles, Los Angeles, California 90095, USA.
authors.forename: Timothy V
authors.initials: TV
authors.lastname: Beischlag
authors.forename: Song
authors.initials: S
authors.lastname: Wang
authors.forename: David W
authors.initials: DW
authors.lastname: Rose
authors.forename: Joseph
authors.initials: J
authors.lastname: Torchia
authors.forename: Suzanne
authors.initials: S
authors.lastname: Reisz-Porszasz
authors.forename: Khurshid
authors.initials: K
authors.lastname: Muhammad
authors.forename: Walter E
authors.initials: WE
authors.lastname: Nelson
authors.forename: Markus R
authors.initials: MR
authors.lastname: Probst
authors.forename: Michael G
authors.initials: MG
authors.lastname: Rosenfeld
authors.forename: Oliver
authors.initials: O
authors.lastname: Hankinson
mesh_terms: D051784:Aryl Hydrocarbon Receptor Nuclear Translocator; D002478:Cells, Cultured; D019363:Cytochrome P-450 CYP1A1 / Q000235:genetics; D004268:DNA-Binding Proteins; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation / Q000187:drug effects; D018257:Helix-Loop-Helix Motifs; D051548:Histone Acetyltransferases; D006801:Humans; D056920:Nuclear Receptor Coactivator 1; D051958:Nuclear Receptor Coactivator 2; D056921:Nuclear Receptor Coactivator 3; D000072317:Polychlorinated Dibenzodioxins / Q000633:toxicity; D011233:Precipitin Tests; D018336:Receptors, Aryl Hydrocarbon / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D013723:Teratogens / Q000633:toxicity; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
chemical_list: C497396:ARNT protein, human; D004268:DNA-Binding Proteins; C498001:NCOA2 protein, human; D051958:Nuclear Receptor Coactivator 2; D000072317:Polychlorinated Dibenzodioxins; D018336:Receptors, Aryl Hydrocarbon; D013723:Teratogens; D015534:Trans-Activators; D014157:Transcription Factors; D051784:Aryl Hydrocarbon Receptor Nuclear Translocator; D019363:Cytochrome P-450 CYP1A1; D051548:Histone Acetyltransferases; C538859:NCOA1 protein, human; C538866:NCOA3 protein, human; D056920:Nuclear Receptor Coactivator 1; D056921:Nuclear Receptor Coactivator 3