Effects of glutathione on chromium-induced DNA crosslinking and DNA polymerase arrest
Effects of glutathione on chromium-induced DNA crosslinking and DNA polymerase arrest
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谷胱甘肽对铬诱导的 DNA 交联和 DNA 聚合酶阻滞的影响
DOI：
10.1007/978-1-4615-0793-2_20
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发表时间：
2001
影响因子：
4.3
通讯作者：
S. Patierno
中科院分区：
文献类型：
--
作者：
T. O'brien;Jian Xu;S. Patierno
六价铬（Cr（VI））在细胞内被抗坏血酸（Asc）、半胱氨酸和谷胱甘肽（GSH）还原为Cr（V）、Cr（IV）和Cr（HI）。这些代谢物诱导一系列基因组DNA损伤，导致DNA复制抑制。我们以前的研究表明，用Cr（III）或Cr（VI）加Asc处理DNA导致DNA-CrDNA交联（Cr-DDC）的形成和原核和哺乳动物DNA聚合酶的鸟嘌呤特异性停滞。GSH不仅作为Cr（VI）的还原剂，而且通过Cr与DNA交联，因此，本研究的重点是研究GSH在Cr诱导的DNA损伤和聚合酶停滞中的作用。在存在GSH的情况下，Cr（III）与质粒DNA共孵育导致GSH与DNA交联。GSH与Cr（III）共处理也导致Cr诱导的DNA链间交联程度相对于单独Cr（III）降低，而不影响总Cr DNA结合。DNA聚合酶的逮捕，观察治疗后的DNA与Cr（III）单独，但显着减少GSH加入到反应混合物中。预先形成的聚合酶阻止病变（铬DDC）没有被随后添加GSH去除。在GSH存在下，用Cr（VI）处理DNA导致GSH与DNA交联，但未能产生可检测的DNA链间交联或聚合酶停滞。使用定量PCR（QPCR）分析，在人类基因组DNA中进一步证实了GSH对Cr诱导的聚合酶停滞的抑制作用。用Cr（III）处理基因组DNA，可明显抑制Tag聚合酶对p53基因1.6kb靶片段的扩增。这是几乎完全防止与GSH和Cr（III）的共同治疗。这些结果表明，铬诱导的DNA链间交联，而不是DNA-Cr-GSH交联，是负责阻断DNA复制的主要病变。此外，DNA-Cr-GSH交联的形成实际上可能阻止聚合酶抑制病变的形成。(Mol Cell Biochem222：173-182，2001）
Hexavalent chromium (Cr (VI)) is reduced intracellularly to Cr (V), Cr (IV) and Cr (HI) by ascorbate (Asc), cysteine and glutathione (GSH). These metabolites induce a spectrum of genomic DNA damage resulting in the inhibition of DNA replication. Our previous studies have shown that treatment of DNA with Cr (III) or Cr (VI) plus Asc results in the formation of DNA-CrDNA crosslinks (Cr-DDC) and guanine-specific arrests of both prokaryotic and mammalian DNA polymerases. GSH not only acts as a reductant of Cr (VI) but also becomes crosslinked to DNA by Cr, thus, the focus of the present study was to examine the role of GSH in Cr-induced DNA damage and polymerase arrests. Co-incubation of Cr (III) with plasmid DNA in the presence of GSH led to the crosslinking of GSH to DNA. GSH co-treatment with Cr (III) also led to a decrease in the degree of Cr-induced DNA interstrand crosslinks relative to Cr(III)alone, without affecting total Cr DNA binding. DNA polymerase arrests were observed following treatment of DNA with Cr (III) alone, but were markedly reduced when GSH was added to the reaction mixture. Pre-formed polymerase-arresting lesions (Cr-DDC) were not removed by subsequent addition of GSH. Treatment of DNA with Cr (VI), in the presence of GSH, resulted in crosslinking of GSH to DNA, but failed to produce detectable DNA interstrand crosslinks or polymerase arrests. The inhibitory effect of GSH on Cr-induced polymerase arrest was further confirmed in human genomic DNA using quantitative PCR (QPCR) analysis. Treatment of genomic DNA with Cr (III) resulted in a marked inhibition of the amplification of a 1.6 kb target fragment of the p53 gene byTagpolymerase. This was almost completely prevented by co-treatment with GSH and Cr (III). These results indicate that Cr-induced DNA interstrand crosslinks, and not DNA-Cr-GSH crosslinks, are the principal lesions responsible for blocking DNA replication. Moreover, the formation of DNA-Cr-GSH crosslinks may actually preclude the formation of the polymerase arresting lesions. (Mol Cell Biochem222:173-182, 2001)
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