Krogsdam Anne-M, Nielsen Curt A F, Neve Søren, Holst Dorte, Helledie Torben, Thomsen Bo, Bendixen Christian, Mandrup Susanne, Kristiansen Karsten
Department of Biochemistry and Molecular Biology, University of Southern Denmark, Odense University, Campusvej 55, DK-5230 Odense M, Denmark.
Biochem J. 2002 Apr 1;363(Pt 1):157-65. doi: 10.1042/0264-6021:3630157.
The nuclear receptor corepressor (NCoR) was isolated as a peroxisome-proliferator-activated receptor (PPAR) delta interacting protein using the yeast two-hybrid system. NCoR interacted strongly with the ligand-binding domain of PPAR delta, whereas interactions with the ligand-binding domains of PPAR gamma and PPAR alpha were significantly weaker. PPAR-NCoR interactions were antagonized by ligands in the two-hybrid system, but were ligand-insensitive in in vitro pull-down assays. Interaction between PPAR delta and NCoR was unaffected by coexpression of retinoid X receptor (RXR) alpha. The PPAR delta-RXR alpha heterodimer bound to an acyl-CoA oxidase (ACO)-type peroxisome-proliferator response element recruited a glutathione S-transferase-NCoR fusion protein in a ligand-independent manner. Contrasting with most other nuclear receptors, PPAR delta was found to interact equally well with interaction domains I and II of NCoR. In transient transfection experiments, NCoR and the related silencing mediator for retinoid and thyroid hormone receptor (SMRT) were shown to exert a marked dose-dependent repression of ligand-induced PPAR delta-mediated transactivation; in addition, transactivation induced by the cAMP-elevating agent forskolin was efficiently reduced to basal levels by NCoR as well as SMRT coexpression. Our results suggest that the transactivation potential of liganded PPAR delta can be fine-tuned by interaction with NCoR and SMRT in a manner determined by the expression levels of corepressors and coactivators.
利用酵母双杂交系统，核受体辅阻遏物（NCoR）作为过氧化物酶体增殖物激活受体（PPAR）δ相互作用蛋白被分离出来。NCoR与PPARδ的配体结合域强烈相互作用，而与PPARγ和PPARα的配体结合域的相互作用则明显较弱。在双杂交系统中，配体可拮抗PPAR-NCoR相互作用，但在体外下拉试验中该相互作用对配体不敏感。视黄酸X受体（RXR）α的共表达不影响PPARδ与NCoR之间的相互作用。PPARδ-RXRα异二聚体与酰基辅酶A氧化酶（ACO）型过氧化物酶体增殖物反应元件结合，以不依赖配体的方式募集谷胱甘肽S-转移酶-NCoR融合蛋白。与大多数其他核受体不同，发现PPARδ与NCoR的相互作用结构域I和II的相互作用同样良好。在瞬时转染实验中，NCoR和相关的类视黄醇和甲状腺激素受体沉默介质（SMRT）对配体诱导的PPARδ介导的反式激活具有明显的剂量依赖性抑制作用；此外，并表达NCoR以及SMRT可将cAMP升高剂福斯可林诱导的反式激活有效地降低至基础水平。我们的结果表明，结合配体的PPARδ 的反式激活潜能可通过与NCoR和SMRT的相互作用进行微调，这种微调方式取决于辅阻遏物和辅激活剂的表达水平。
