Critical role of chromium (Cr)-DNA interactions in the formation of Cr-induced polymerase arresting lesions
Critical role of chromium (Cr)-DNA interactions in the formation of Cr-induced polymerase arresting lesions
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DOI：
10.1021/bi020452j
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发表时间：
2002-10-15
期刊：
影响因子：
2.9
通讯作者：
Patierno, SR
中科院分区：
文献类型：
--
作者：
O'Brien, T;Mandel, HG;Patierno, SR
与六价铬[Cr（VI）]代谢还原相关的遗传毒性很复杂，可阻碍DNA聚合酶介导的体外复制。铬诱导的聚合酶抑制病变（帕尔斯）的确切生物化学性质尚不清楚，但据信涉及Cr-DNA链间交联（ICL）的形成。本研究的目的是确定直接铬-DNA相互作用的依赖性上的发展PAL处理的DNA与三价铬[铬（III）]或与铬（VI）的存在下，抗坏血酸（Asc），一个主要的细胞内还原剂，使用在体外，非细胞系统。Cr-DNA加合物、ICLs和帕尔斯的形成在Asc：Cr（VI）摩尔比为0.5-2时最大，但在更高的比例下逐渐减少。EDTA，铬（III）螯合剂，显着降低Cr-DNA结合和ICL和PAL的形成。用Cr（VI）/Asc和甘露醇（Cr（V）螯合剂）共处理DNA，选择性地抑制Cr（VI）还原产生的单/双功能DNA加合物和帕尔斯的形成，但对Cr（III）-DNA结合或Cr（III）诱导的聚合酶阻滞没有影响。阻断Cr-DNA磷酸盐相互作用的预孵育的DNA与氯化镁废除DNA结合和ICL和PAL的生产。DNA链断裂和脱碱基位点可能会导致DNA聚合酶在体外被捕，但是，我们未能检测到显着增加的频率，这些病变后铬（VI）/天冬氨酸治疗。这些数据表明，双功能加合铬的DNA磷酸（ICLs）构成了一个主要的PAL。此外，通过Cr（VI）还原产生的DNA链断裂和脱碱基位点不足以解释体外观察到的帕尔斯。
The genotoxicity associated with the metabolic reduction of hexavalent chromium [Cr(VI)] is complex and can impede DNA polymerase-mediated replication in vitro. The exact biochemical nature of Cr-induced polymerase arresting lesions (PALs) is not understood, but is believed to involve the formation of Cr-DNA interstrand cross-links (ICLs). The aim of this investigation was to determine the dependence of direct Cr-DNA interactions on the development of PALs in DNA treated with trivalent Cr [Cr(III)] or with Cr(VI) in the presence of ascorbic acid (Asc), a major intracellular reductant, using an in vitro, acellular system. The formation of Cr-DNA adducts, ICLs, and PALs was maximal at Asc:Cr(VI) molar ratios of 0.5-2, but gradually decreased at higher ratios. EDTA, a Cr(III) chelator, significantly decreased Cr-DNA binding and ICL and PAL formation. Co-treatment of DNA with Cr(VI)/Asc and mannitol, a Cr(V) chelator, selectively inhibited the formation of mono/bifunctional DNA adducts and PALs produced by Cr(VI) reduction, but had no effect on Cr(III)-DNA binding or Cr(III)-induced polymerase arrest. Blocking Cr-DNA phosphate interaction by preincubation of DNA with MgCl2 abrogated DNA binding and ICL and PAL production. DNA strand breaks and abasic sites may lead to the in vitro arrest of DNA polymerases; however, we failed to detect significant increases in the frequency of these lesions following Cr(VI)/Asc treatment. These data indicate that the bifunctional adduction of Cr to DNA phosphates (ICLs) constitutes a major PAL. Furthermore, the generation of DNA strand breaks and abasic sites by Cr(VI) reduction is insufficient to explain PALs observed in vitro.