The p65 isoform of Nrf1 is a dominant negative inhibitor of ARE-mediated transcription
The p65 isoform of Nrf1 is a dominant negative inhibitor of ARE-mediated transcription
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DOI：
10.1074/jbc.m700159200
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发表时间：
2007-08-24
影响因子：
4.8
通讯作者：
Chan, Jefferson Y.
中科院分区：
文献类型：
--
作者：
Wang, Weiping;Kwok, Annie M.;Chan, Jefferson Y.
氧化应激反应转录部分通过被称为抗氧化反应元件（ARE）的顺式活性序列进行调节。通过ARE激活涉及碱性亮氨酸拉链蛋白的cnc亚家族成员，包括Nrf1和Nrf2。特别是，Nrf2已被证明可以协调诱导编码抗氧化和2期代谢酶的基因，以响应亲电化合物和暴露于异种生物的刺激。在这里，我们表明Nrf1基因的65-kDa亚型作为Nrf2的抑制因子。p65Nrf1的瞬时表达抑制了nrf2介导的are依赖性报告基因在细胞中的激活。内源性re -基因的诱导在稳定表达p65Nrf1的Hepa1c1c7细胞中被阻断，导致细胞死亡增加。与这些发现一致，在Nrf1（-/-）成纤维细胞中，p65Nrf1功能的丧失增强了are基因表达的亲电激活，而在稳定表达p65Nrf1的Nrf1（-/-）细胞中，氧化预处理和are基因表达的保护作用被阻断。凝胶转移实验表明，p65Nrf1以异源二聚体形式与小maf蛋白结合抗氧化反应元件。免疫沉淀研究表明，p65Nrf1在体内与Nrf2竞争与小maf蛋白的相互作用和与抗氧化反应元件的结合。综上所述，这些结果表明p65Nrf1可能在调节氧化应激反应中发挥重要作用，其功能是作为nrf2介导的are依赖性基因转录激活的跨显性抑制因子。
Oxidative stress-responsive transcription is regulated in part through cis-active sequences known as antioxidant response elements (ARE). Activation through the ARE involves members of the CNC-subfamily of basic leucine zipper proteins including Nrf1 and Nrf2. In particular, Nrf2 has been shown to coordinate induction of genes encoding antioxidant and phase 2 metabolizing enzymes in response to stimulation with electrophilic compounds and exposure to xenobiotics. Here we show that the 65-kDa isoform of the Nrf1 gene functions as a repressor of Nrf2. Transient expression of p65Nrf1 suppressed Nrf2-mediated activation of ARE-dependent reporter genes in cells. Induction of endogenous ARE-genes is blocked in Hepa1c1c7 cells stably expressing p65Nrf1 leading to increased cell death. Consistent with these findings, electrophilic activation of ARE-gene expression is augmented by loss of p65Nrf1 function in Nrf1(-/-) fibroblasts, and the protective effects of oxidative preconditioning and ARE-gene expression are blocked in Nrf1(-/-) cells stably expressing p65Nrf1. Gel shift experiments demonstrated that p65Nrf1 binds the antioxidant response element as a heterodimer with small-Maf protein. Immunoprecipitation studies demonstrated that p65Nrf1 competes with Nrf2 for interaction with small-Maf protein and binding to the antioxidant response element in vivo. Together, these results demonstrate that p65Nrf1 has the potential to play an important role in modulating the response to oxidative stress by functioning as a transdominant repressor of Nrf2-mediated activation of ARE-dependent gene transcription.