Novel role of antioxidant-1 (Atox1) as a copper-dependent transcription factor involved in cell proliferation
Novel role of antioxidant-1 (Atox1) as a copper-dependent transcription factor involved in cell proliferation
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DOI：
10.1074/jbc.m709463200
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发表时间：
2008-04-04
影响因子：
4.8
通讯作者：
Fukai, Tohru
中科院分区：
文献类型：
--
作者：
Itoh, Shinichi;Kim, Ha Won;Fukai, Tohru
铜在调节细胞生长方面起着至关重要的作用。许多类型的人类癌症组织中的铜含量高于正常组织。铜还能诱导基因表达。然而，在哺乳动物中，介导铜诱导细胞增殖的转录因子尚未被确定。在此我们表明，抗氧化蛋白1（Atox1），先前被认为是一种铜伴侣蛋白，是一种新型的铜依赖性转录因子，可介导铜诱导的细胞增殖。用铜刺激小鼠胚胎成纤维细胞（MEFs）可显著增加细胞增殖、细胞周期蛋白D1的表达以及进入S期，而在Atox1缺失（-/-）的MEFs中这些现象完全消失。启动子分析和电泳迁移率变动分析（EMSA）显示，铜刺激Atox1与细胞周期蛋白D1启动子中一个先前未描述的顺式元件结合。染色质免疫沉淀（ChIP）分析证实铜在体内刺激Atox1与DNA结合。将Atox1与Gal4的DNA结合结构域融合后进行转染，在包括内皮细胞和癌细胞在内的多种细胞类型中显示出铜依赖性的反式激活作用。此外，Atox1通过其高度保守的C末端KKTGK基序和N末端铜结合位点响应铜而转移至细胞核。最后，通过观察在Atox1缺失（-/-）的MEFs中重新表达核靶向的Atox1可挽救有缺陷的铜诱导细胞增殖，证明了核Atox1的功能作用。因此，Atox1作为一种新型转录因子发挥作用，当被铜激活时，它会发生核转位、DNA结合和反式激活，从而促进细胞增殖。
Copper plays a fundamental role in regulating cell growth. Many types of human cancer tissues have higher copper levels than normal tissues. Copper can also induce gene expression. However, transcription factors that mediate copper-induced cell proliferation have not been identified in mammals. Here we show that antioxidant-1 (Atox1), previously appreciated as a copper chaperone, represents a novel copper-dependent transcription factor that mediates copper-induced cell proliferation. Stimulation of mouse embryonic fibroblasts (MEFs) with copper markedly increased cell proliferation, cyclin D1 expression, and entry into S phase, which were completely abolished in Atox1(-/-) MEFs. Promoter analysis and EMSA revealed that copper stimulates the Atox1 binding to a previously undescribed cis element in the cyclin D1 promoter. The ChIP assay confirms that copper stimulates Atox1 binding to the DNA in vivo. Transfection of Atox1 fused to the DNA-binding domain of Gal4 demonstrated a copper-dependent transactivation in various cell types, including endothelial and cancer cells. Furthermore, Atox1 translocated to the nucleus in response to copper through its highly conserved C-terminal KKTGK motif and N-terminal copper-binding sites. Finally, the functional role of nuclear Atox1 is demonstrated by the observation that re-expression of nuclear-targeted Atox1 in Atox1(-/-) MEFs rescued the defective copper- induced cell proliferation. Thus, Atox1 functions as a novel transcription factor that, when activated by copper, undergoes nuclear translocation, DNA binding, and transactivation, thereby contributing to cell proliferation.