A far-upstream AP-1/Smad binding box regulates human NOX4 promoter activation by transforming growth factor-β.
A far-upstream AP-1/Smad binding box regulates human NOX4 promoter activation by transforming growth factor-β.
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DOI：
10.1016/j.gene.2014.02.026
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发表时间：
2014-04-25
期刊：
影响因子：
3.5
通讯作者：
Thannickal VJ
中科院分区：
文献类型：
--
作者：
Bai G;Hock TD;Logsdon N;Zhou Y;Thannickal VJ
NADPH氧化酶4（NOX 4）是NADPH氧化酶基因家族的成员，其调节细胞分化、先天免疫和组织纤维化。已知转化生长因子-β（TGF-β1）可诱导间充质细胞中NOX 4 mRNA的表达。然而，NOX 4的转录调控机制还不清楚。在这项研究中，我们研究了TGF-β1对人肺成纤维细胞中NOX 4的转录调控。构建了5个启动子-报告基因构建体，分别含有人NOX 4基因转录起始位点（TSS）上游0.74 kb、1.35 kb、1.84 kb、3.97 kb和4.76 kb的DNA片段，并分析了它们对TGF-β1的相对反应性。TGF-β1诱导的NOX 4基因启动子激活需要−3.97 kb和−4.76 kb之间的区域。生物信息学分析表明，该区域存在一个15 bp的AP-1/Smad结合元件。AP-1或Smad元件的突变或缺失减弱了-4.76 kb NOX 4启动子的TGF-β1反应性。此外，AP-1/Smad盒的插入使TGF-β1诱导非响应性-3.97 kb NOX 4启动子构建体。染色质免疫沉淀分析表明磷酸化Smad 3和cJun以TGF-β1诱导的方式与该元件结合。这些结果表明，AP-1/Smad box位于NOX 4启动子TSS位点上游3.97 kb和4.76 kb之间，是TGF-β1诱导人肺成纤维细胞中NOX 4基因转录所必需的。我们的研究提供了对NOX 4基因表达的分子机制的见解，为干扰以TGF-β1/NOX 4通路激活为特征的疾病中NOX 4上调提供了新的治疗方法。
NADPH oxidase 4 (NOX4) is a member of the NADPH oxidase gene family that regulates cellular differentiation, innate immunity and tissue fibrosis. Transforming growth factor-β (TGF-β1) is known to induce expression of NOX4 mRNA in mesenchymal cells. However, the mechanisms of transcriptional regulation of NOX4 are not well understood. In this study, we examined the transcriptional regulation of NOX4 in human lung fibroblasts by TGF-β1. Five promoter-reporter constructs containing DNA fragments of 0.74 kb, 1.35 kb, 1.84 kb, 3.97 kb and 4.76 kb upstream from the transcriptional start site (TSS) of the human NOX4 gene were generated and their relative responsiveness to TGF-β1 analyzed. TGF-β1-induced NOX4 gene promoter activation requires a region between −3.97 kb and −4.76 kb. Bioinformatics analysis revealed a 15 bp AP-1/Smad binding element in this region. Mutation or deletion of either the AP-1 or the Smad element attenuated TGF-β1 responsiveness of the −4.76 kb NOX4 promoter. Furthermore, insertion of this AP-1/Smad box conferred TGF-β1 inducibility to the non-responsive −3.97 kb NOX4 promoter construct. Chromatin immunoprecipitation analysis indicated phospho-Smad3 and cJun associate with this element in a TGF-β1-inducible manner. These results demonstrate that the AP-1/Smad box located between 3.97 kb and 4.76 kb upstream of the TSS site of the NOX4 promoter is essential for NOX4 gene transcription induced by TGF-β1 in human lung fibroblasts. Our study provides insights into the molecular mechanisms of NOX4 gene expression, informing novel therapeutic approaches to interfere with upregulation of NOX4 in diseases characterized by activation of the TGF-β1/NOX4 pathway.
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影响因子：
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作者：
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通讯作者：
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DOI：
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影响因子：
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