Nuclear receptors RXRα:RARα are repressors for human MRP3 expression
Nuclear receptors RXRα:RARα are repressors for human MRP3 expression
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DOI：
10.1152/ajpgi.00191.2006
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发表时间：
2007-05-01
影响因子：
4.5
通讯作者：
Boyer, James L.
中科院分区：
文献类型：
--
作者：
Chen, Wensheng;Cai, Shi-Ying;Boyer, James L.
多药耐药相关蛋白MRP3/MRP3(ABCC3)在胆汁淤积症中上调，这是一种适应性反应，可能保护肝脏免受有毒化合物的堆积，如胆盐和胆红素结合物。然而，这种上调的机制却鲜为人知。我们和其他人之前已经报道了胎儿蛋白转录因子/肝脏受体同源1是MRP3/MRP3表达的激活剂。在寻找人类MRP3启动子中的其他调控元件时，我们现在已经确定核受体维甲酸X受体-α：维甲酸受体-α(RXRα：RARα)是转录因子Sp1激活MRP3的抑制因子。荧光素酶报告分析表明，共转染转录因子Sp1可以刺激MRP3启动子的活性，RXRα：RARα的加入以剂量依赖的方式抑制了这种激活。位点突变和凝胶位移分析已经确定在MRP3翻译起始点上游-113到-108个核苷酸处有一个Sp1结合的GC盒基序，其中RXRα：RARα特异性地减少了Sp1与该位点的结合。GC盒的突变也降低了MRP3启动子的活性。RXRα：RARα作为MRP3表达的抑制因子，在HepG2细胞中被RARα小干扰RNA敲除，从而上调内源性MRP3的表达，进一步证实了RXRα的功能作用。综上所述，我们的结果表明，激活子Sp1和抑制子RXRα：RARα协同作用调节MRP3的表达。由于RXRα：RARα的表达因淤胆性肝损伤而降低，RXRα：RARα的缺失可能导致这些疾病中MRP3/MRP3的表达上调。
Multidrug resistanceassociated protein MRP3/Mrp3 ( ABCC3) is upregulated in cholestasis, an adaptive response that may protect the liver from accumulation of toxic compounds, such as bile salts and bilirubin conjugates. However, the mechanism of this upregulation is poorly understood. We and others have previously reported that fetoprotein transcription factor/ liver receptor homolog- 1 is an activator of MRP3/ Mrp3 expression. In searching for additional regulatory elements in the human MRP3 promoter, we have now identified nuclear receptor retinoic X receptor-alpha: retinoic acid receptor-alpha ( RXR alpha: RAR alpha) as a repressor of MRP3 activation by transcription factor Sp1. A luciferase reporter assay demonstrated that cotransfection of transcription factor Sp1 stimulates the MRP3 promoter activity and that additions of RXR alpha: RAR alpha abrogated this activation in a dose- dependent manner. Site mutations and gel shift assays have identified a Sp1 binding GC box motif at -113 to -108 nts upstream from the MRP3 translation start site, where RXR alpha: RAR alpha specifically reduced Sp1 binding to this site. Mutation of the GC box also reduced MRP3 promoter activity. The functional role of RXR alpha: RAR alpha as a repressor of MRP3 expression was further confirmed by RAR alpha small- interfering RNA knockdown in HepG2 cells, which upregulated endogenous MRP3 expression. In summary, our results indicate that activator Sp1 and repressor RXR alpha: RAR alpha act in concert to regulate MRP3 expression. Since RXR alpha: RAR alpha expression is diminished by cholestatic liver injury, loss of RXR alpha: RAR alpha may lead to upregulation of MRP3/ Mrp3 expression in these disorders.