NF1, Sp1 and HSF1 are synergistically involved in sulfide-induced sqr activation in echiuran worm Urechis unicinctus
NF1, Sp1 and HSF1 are synergistically involved in sulfide-induced sqr activation in echiuran worm Urechis unicinctus
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NF1、Sp1 和 HSF1 协同参与 echiuran 蠕虫 Urechis unicinctus 中硫化物诱导的 sqr 激活
DOI：
10.1016/j.aquatox.2016.04.002
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发表时间：
2016-06-01
影响因子：
4.5
通讯作者：
Zhang, Zhifeng
中科院分区：
文献类型：
--
作者：
Liu, Xiaolong;Qin, Zhenkui;Zhang, Zhifeng
背景：硫化物是一种众所周知的环境有毒物质。线粒体硫化物氧化是生物体内硫化物解毒的主要机制之一，而硫化物：醌氧化还原酶（SQR）是参与硫化物向泛醌传递电子并将硫化物转化为硫代硫酸盐的关键酶。已有研究表明单环刺鳅线粒体存在SQR介导的硫化物氧化反应，且SQR mRNA水平在硫化物暴露后显著升高。在本研究中，我们试图揭示转录因子对硫诱导的sqr转录的协同调节。unicinctus.Methods:采用ChIP和EMSA技术检测sqr近端启动子（-391 ~+194 bp）与转录因子NF 1（核因子1）和Sp1（特异性蛋白1）的相互作用。定点突变和转染实验进一步揭示了它们在sqr转录中的结合位点和HSF 1、NF 1和Sp1的协同作用。当你。用150 μ M的硫化物处理unicinctus，Western blotting检测NF 1和Sp1的表达水平和核含量，ChIP检测NF 1和Sp1与sqr启动子的结合量。证实转录因子NF 1和Sp1与sqr近端启动子相互作用，NF 1和Sp1的结合位点分别在75 ~ 69 bp和210 ~ 201 bp之间。转染实验表明NF 1和Sp1结合位点的突变分别使sqr启动子活性降低50%和73%。此外，我们证明了三个转录因子NF 1，Sp1和HSF 1协同增强sqr转录活性。此外，当蠕虫暴露于150 μ M硫化物时，后肠中与sqr近端区域结合的NF 1或Spl的含量显著增加。结论：转录因子NF 1、Spl和HSF 1均参与了硫诱导sqr基因的转录。硫化物不仅可以通过增加它们的表达水平，而且可以促进它们进入细胞核并与sqr启动子结合来激活sqr转录。NF 1和Spl参与基础和硫化物诱导的sqr转录，而HSF 1主要在硫化物诱导的sqr转录中起作用。(C)2016爱思唯尔B. V.保留所有权利。
Background: Sulfide is a well-known environmental toxic substance. Mitochondrial sulfide oxidation is a main mechanism of sulfide detoxification in organisms, and sulfide: quinone oxidoreductase (SQR) is a key enzyme which is involved in transferring electrons from sulfide to ubiquinone and converting sulfide into thiosulfate. Previous studies have revealed the SQR-mediated mitochondrial sulfide oxidation exists in the echiuran worm Urechis unicinctus, and its sqr mRNA level increased significantly when the worm is exposed to sulfide. In this study, we attempt to reveal the synergistic regulation of transcription factors on sulfide-induced sqr transcription in U. unicinctus.Methods: ChIP and EMSA were used to identify the interactions between sqr proximal promoter (from -391 to +194 bp) and transcription factors NF1 (nuclear factor 1) and Sp1 (specificity protein 1). Site directed mutation and transfection assays further revealed their binding sites and synergistic roles of HSF1, NF1 and Sp1 in the sqr transcription. When U. unicinctus were exposed to 150 mu M sulfide, the expression levels and nuclear contents of NF1 and Sp1 were examined by Western blotting, and the binding contents between NF1 or Sp1 and the sqr promoter were also detected by ChIP.Results: Transcription factors NF1 and Spl were confirmed to interact with the sqr proximal promoter, and their binding sites were identified in 75 to 69 by for NF1 and 210 to 201 bp for Spl. Transfection assays showed mutation of NF1 or Sp1 binding site significantly decreased the sqr promoter activity by 50% or 73%, respectively. Moreover, we demonstrated three transcription factors NF1, Spl and HSF1 enhanced synergistically the activity of sqr transcription. Furthermore, contents of NF1 or Spl binding to the sqr proximal region increased significantly in the hindgut when the worms were exposed to 150 mu M sulfide. Similar changes of NF1 or Spl levels and nuclear NF1 or Spl levels were also presented.Conclusion: Transcription factors NF1, Spl and HSF1 are all involved in sulfide-induced sqr transcription. Sulfide can activate sqr transcription by not only increasing their expression levels, but also promoting them entering nucleus and binding to the sqr promoter. NF1 and Spl participate in both basal and sulfide induced sqr transcription, while HSF1 functions mainly in sulfide-induced sqr transcription. (C) 2016 Elsevier B.V. All rights reserved.