Shan Ying, Lambrecht Richard W, Ghaziani Tahereh, Donohue Susan E, Bonkovsky Herbert L
Department of Medicine and Pharmacology, the General Clinical Research Center of the University of Connecticut Health Center, Farmington, Connecticut 06030, USA.
J Biol Chem. 2004 Dec 10;279(50):51769-74. doi: 10.1074/jbc.M409463200. Epub 2004 Oct 1.
Heme oxygenase-1 is an antioxidant defense enzyme that converts heme to biliverdin, iron, and carbon monoxide. Bach-1 is a bZip protein that forms heterodimers with small Maf proteins and was reported recently to down-regulate the HO-1 gene in mice. Using small interfering RNAs targeted to human Bach-1 mRNA, we investigated whether modulation of human hepatic Bach-1 expression by small interfering (si)RNA technology influences heme oxygenase-1 gene expression. We found that Bach-1 siRNAs transfected into Huh-7 cells significantly reduced Bach-1 mRNA and protein levels approximately 80%, compared with non siRNA-treated cells. In contrast, transfection with the same amounts of nonspecific control duplexes or LaminB2-duplex did not reduce Bach-1 mRNA or protein levels, confirming the specificity of Bach-1 siRNA. Expression of the heme oxygenase-1 gene in Bach-1 siRNA-transfected cells was up-regulated 7-fold, compared with cells without Bach-1 siRNA. The effect of increasing concentrations of heme to up-regulate levels of heme oxygenase-1 was more pronounced when Bach-1 siRNA was present. Taken together, these results indicated that Bach-1 has a specific and selective ability to repress expression of human hepatic heme oxygenase-1. Silencing of Bach-1 by siRNAs is a useful method for up-regulating HO-1 gene expression. Exogenous heme produces additional up-regulation, beyond that produced by Bach-1 siRNAs, suggesting that heme does not act solely through its effects on Bach-1.
血红素加氧酶-1是一种抗氧化防御酶，可将血红素转化为胆绿素、铁和一氧化碳。Bach-1是一种bZip蛋白，可与小Maf蛋白形成异源二聚体，最近有报道称其可下调小鼠中的HO-1基因。我们使用靶向人Bach-1 mRNA的小干扰RNA，研究了通过小干扰（si）RNA技术调节人肝脏Bach-1表达是否会影响血红素加氧酶-1基因表达。我们发现，与未用siRNA处理的细胞相比，转染到Huh-7细胞中的Bach-1 siRNA可使Bach-1 mRNA和蛋白水平显著降低约80%。相比之下，转染相同量的非特异性对照双链体或LaminB2双链体不会降低Bach-1 mRNA或蛋白水平，从而证实了Bach-1 siRNA的特异性。与未转染Bach-1 siRNA的细胞相比，转染Bach-1 siRNA的细胞中血红素加氧酶-1基因的表达上调了7倍。当存在Bach-1 siRNA时，增加血红素浓度上调血红素加氧酶-1水平的作用更为明显。综上所述，这些结果表明Bach-1具有特异性和选择性抑制人肝脏血红素加氧酶-1表达的能力。通过siRNA使Bach-1沉默是上调HO-1基因表达的一种有用方法。外源性血红素产生的上调作用超出了Bach-1 siRNA所产生的作用，这表明血红素并非仅通过其对Bach-1的作用发挥作用。
