Chromate Reductase YieF from Escherichia coli Enhances Hexavalent Chromium Resistance of Human HepG2 Cells.
Chromate Reductase YieF from Escherichia coli Enhances Hexavalent Chromium Resistance of Human HepG2 Cells.
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DOI：
10.3390/ijms160611892
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发表时间：
2015-05-26
影响因子：
5.6
通讯作者：
Liu P
中科院分区：
文献类型：
--
作者：
Liu X;Wu G;Zhang Y;Wu D;Li X;Liu P
六价铬（Cr(VI)）是一种严重的环境污染物和人体毒物。哺乳动物细胞对铬酸盐非常敏感，因为它们缺乏有效的铬酸盐解毒策略，例如原核生物中广泛存在的铬酸盐还原基因。为了测试将原核铬酸盐还原基因引入哺乳动物细胞是否可以产生更高的铬酸盐抗性，将大肠杆菌铬酸盐还原基因yieF转染到人HepG2细胞中。通过定量RT-PCR测量稳定转染的HepG2-YieF细胞中yieF的表达，发现Cr(VI)诱导后yieF的表达上调3.89倍。在铬酸盐还原能力测试中，携带还原酶的HepG2-YieF细胞显示出比HepG2对照细胞显着更高的Cr(VI)还原能力。 HepG2-YieF 细胞粗提取物去除 Cr(VI) 能力增强的证据进一步支持了这一结果。此外，在 Cr(VI) 胁迫下，HepG2-YieF 的活力提高了 10%，GSH 合成酶的表达降低。 YieF 的亚细胞定位是通过追踪通过激光共聚焦显微镜在细胞核和细胞质中检测到的 GFP-YieF 融合蛋白来确定的。总之，该研究成功证明了原核Cr(VI)还原基因yieF的表达赋予哺乳动物细胞HepG2增强的铬酸盐抗性，这为哺乳动物细胞中Cr(VI)解毒带来了新的见解。
Hexavalent chromium (Cr(VI)) is a serious environmental pollutant and human toxicant. Mammalian cells are very sensitive to chromate as they lack efficient chromate detoxifying strategy, e.g., chromate-reducing genes that are widely present in prokaryotes. To test whether introduction of prokaryotic chromate-reducing gene into mammalian cells could render higher chromate resistance, an Escherichia coli chromate-reducing gene yieF was transfected into human HepG2 cells. The expression of yieF was measured in stably transfected cells HepG2-YieF by quantitative RT-PCR and found up-regulated by 3.89-fold upon Cr(VI) induction. In chromate-reducing ability test, HepG2-YieF cells that harbored the reductase showed significantly higher reducing ability of Cr(VI) than HepG2 control cells. This result was further supported by the evidence of increased Cr(VI)-removing ability of crude cell extract of HepG2-YieF. Moreover, HepG2-YieF demonstrated 10% higher viability and decreased expression of GSH synthesizing enzymes under Cr(VI) stress. Subcellular localization of YieF was determined by tracing GFP-YieF fusion protein that was detected in both nucleus and cytoplasm by laser confocal microscopy. Altogether, this study successfully demonstrated that the expression of a prokaryotic Cr(VI)-reducing gene yieF endowed mammalian cell HepG2 with enhanced chromate resistance, which brought new insight of Cr(VI) detoxification in mammalian cells.
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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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