title: Ligand and coactivator recruitment preferences of peroxisome proliferator activated receptor alpha.
abstract: The mechanism by which ligands of nuclear receptors show differential effects on gene transcription is not fully understood, but is believed to result in part from the preferential recruitment and/or displacement of coactivators and corepressors. We have explored the interaction of several known ligands and the nuclear receptor (peroxisome proliferator activated receptor alpha, PPARalpha) using scintillation proximity assay (SPA) and the interaction of LXXLL containing peptides derived from three coactivators (SRC-1, CBP and PGC-1) with PPARalpha in the presence of PPARalpha agonist ligands using fluorescence resonance energy transfer (FRET). The EC(50)s of the individual ligands for recruitment showed the same rank order regardless of the coactivator peptide used, with GW2331<WY14643=ciprofibrate<L165041<gemfibrozil. Similarly, for all ligands tested, the rank order of EC(50) for peptide recruitment was CBP<PGC-1<SRC-1. These data suggest that for these LXXLL coactivator peptides, the ligands do not substantially differ in their preferences. Partial agonism was observed with ciprofibrate and PGC-1 and gemfibrozil and CBP giving a lower FRET at saturation than with the other ligands. This suggests that ciprofibrate and gemfibrozil induce a different conformation to the receptor-PGC-1 and receptor-CBP complex, respectively. In cotransfection assays, unexpected differences in potencies and efficacies were observed and the rank order of EC(50)s for activation differed from that predicted by FRET assays. In most cases, the presence of a coactivator peptide led to decrease in the EC(50)s seen in FRET assays compared to the K(i)s observed in binding to receptor only, consistent with the lower EC(50)s obtained in the transfection assays. Our data demonstrate that ligand induced coactivator preferences of PPARalpha contribute to transcription potency and efficacy.
authors.affiliation: Cardiovascular Diseases Research, Bristol-Myers Squibb Company, Experimental Station, E400 Wilmington, DE 19880, USA. ranjan.mukherjee@bms.com
authors.forename: Ranjan
authors.initials: R
authors.lastname: Mukherjee
authors.forename: Shaoxian
authors.initials: S
authors.lastname: Sun
authors.forename: Linda
authors.initials: L
authors.lastname: Santomenna
authors.forename: Bowman
authors.initials: B
authors.lastname: Miao
authors.forename: Harry
authors.initials: H
authors.lastname: Walton
authors.forename: Boshan
authors.initials: B
authors.lastname: Liao
authors.forename: Kenneth
authors.initials: K
authors.lastname: Locke
authors.forename: Ji-Hu
authors.initials: JH
authors.lastname: Zhang
authors.forename: Sonny H
authors.initials: SH
authors.lastname: Nguyen
authors.forename: Li Tao
authors.initials: LT
authors.lastname: Zhang
authors.forename: Kathleen
authors.initials: K
authors.lastname: Murphy
authors.forename: Harold O
authors.initials: HO
authors.lastname: Ross
authors.forename: M X
authors.initials: MX
authors.lastname: Xia
authors.forename: Christopher
authors.initials: C
authors.lastname: Teleha
authors.forename: S-Y
authors.initials: SY
authors.lastname: Chen
authors.forename: Bernard
authors.initials: B
authors.lastname: Selling
authors.forename: Richard
authors.initials: R
authors.lastname: Wynn
authors.forename: Timothy
authors.initials: T
authors.lastname: Burn
authors.forename: Peter R
authors.initials: PR
authors.lastname: Young
mesh_terms: D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004735:Energy Transfer; D004926:Escherichia coli / Q000378:metabolism; D051548:Histone Acetyltransferases; D006801:Humans; D007700:Kinetics; D008024:Ligands; D056920:Nuclear Receptor Coactivator 1; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism*; D013053:Spectrophotometry; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection
chemical_list: D018076:DNA, Complementary; D008024:Ligands; D010455:Peptides; D016271:Proto-Oncogene Proteins c-myc; D018160:Receptors, Cytoplasmic and Nuclear; D014157:Transcription Factors; D051548:Histone Acetyltransferases; C538859:NCOA1 protein, human; D056920:Nuclear Receptor Coactivator 1