Vu-Dac N, Schoonjans K, Kosykh V, Dallongeville J, Heyman R A, Staels B, Auwerx J
Unite 325 Institut National de la Santé et de la Recherche Médicale, Département d'Athérosclerose, Institut Pasteur de Lille, France.
Mol Cell Biol. 1996 Jul;16(7):3350-60. doi: 10.1128/MCB.16.7.3350.
Considering the link between plasma high-density lipoprotein (HDL) cholesterol levels and a protective effect against coronary artery disease as well as the suggested beneficial effects of retinoids on the production of the major HDL apolipoprotein (apo), apo A-I, the goal of this study was to analyze the influence of retinoids on the expression of apo A-II, the other major HDL protein. Retinoic acid (RA) derivatives have a direct effect on hepatic apo A-II production, since all-trans (at) RA induces apo A-II mRNA levels and apo A-II secretion in primary cultures of human hepatocytes. In the HepG2 human hepatoblastoma cell line, both at-RA and 9-cis RA as well as the retinoid X receptor (RXR)-specific agonist LGD 1069, but not the RA receptor (RAR) agonist ethyl-p-[(E)-2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthyl)-l-pro penyl]-benzoic acid (TTNPB), induce apo A-II mRNA levels. Transient-transfection experiments with a reporter construct driven by the human apo A-II gene promoter indicated that 9-cis RA and at-RA, as well as the RXR agonists LGD 1069 and LG 100268, induced apo A-II gene expression at the transcriptional level. Only minimal effects of the RAR agonist TTNPB were observed on the apo A-II promoter reporter construct. Unilateral deletions and site-directed mutagenesis identified the J site of the apo A-II promoter mediating the responsiveness to RA. This element contains two imperfect half-sites spaced by 1 oligonucleotide. Cotransfection assays in combination with the use of RXR or RAR agonists showed that RXR but not RAR transactivates the apo A-II promoter through this element. By contrast, RAR inhibits the inductive effects of RXR on the apo A-II J site in a dose-dependent fashion. Gel retardation assays demonstrated that RXR homodimers bind, although with a lower affinity than RAR-RXR heterodimers, to the AH-RXR response element. In conclusion, retinoids induce hepatic apo A-II production at the transcriptional level via the interaction of RXR with an element in the J site containing two imperfect half-sites spaced by 1 oligonucleotide, thereby demonstrating an important role of RXR in controlling human lipoprotein metabolism. Since the J site also confers responsiveness of the apo A-II gene to fibrates and fatty acids via the activation of peroxisome proliferator-activated receptor-RXR heterodimers, this site can be considered a plurimetabolic response element.
鉴于血浆高密度脂蛋白（HDL）胆固醇水平与对冠状动脉疾病的保护作用之间的联系，以及类视黄醇对主要HDL载脂蛋白（apo）即apo A-I产生的有益作用，本研究的目的是分析类视黄醇对另一种主要HDL蛋白apo A-II表达的影响。视黄酸（RA）衍生物对肝脏apo A-II的产生有直接作用，因为全反式（at）RA可诱导人肝细胞原代培养物中apo A-II mRNA水平和apo A-II分泌。在HepG2人肝癌细胞系中，at-RA和9-顺式RA以及视黄醇X受体（RXR）特异性激动剂LGD 1069，但不是RA受体（RAR）激动剂乙基-p-[(E)-2-(5,6,7,8-四氢-5,5,8,8-四甲基-2-萘基)-1-丙烯基]-苯甲酸（TTNPB），可诱导apo A-II mRNA水平。用人apo A-II基因启动子驱动的报告构建体进行的瞬时转染实验表明，9-顺式RA和at-RA以及RXR激动剂LGD 1069和LG 100268在转录水平上诱导apo A-II基因表达。在apo A-II启动子报告构建体上仅观察到RAR激动剂TTNPB的最小作用。单侧缺失和定点诱变确定了apo A-II启动子的J位点介导对RA的反应性。该元件包含两个由1个寡核苷酸隔开的不完全半位点。结合使用RXR或RAR激动剂的共转染分析表明，RXR而非RAR通过该元件反式激活apo A-II启动子。相比之下，RAR以剂量依赖性方式抑制RXR对apo A-II J位点的诱导作用。凝胶阻滞分析表明，RXR同二聚体结合到AH-RXR反应元件上，尽管其亲和力低于RAR-RXR异二聚体。总之，类视黄醇通过RXR与J位点中包含两个由1个寡核苷酸隔开的不完全半位点的元件相互作用，在转录水平上诱导肝脏apo A-II的产生，从而证明RXR在控制人类脂蛋白代谢中起重要作用。由于J位点还通过过氧化物酶体增殖物激活受体-RXR异二聚体的激活赋予apo A-II基因对贝特类药物和脂肪酸的反应性，因此该位点可被视为多代谢反应元件。
