Incorporation of dUMP into DNA is a major source of spontaneous DNA damage, while excision of uracil is not required for cytotoxicity of fluoropyrimidines in mouse embryonic fibroblasts
Incorporation of dUMP into DNA is a major source of spontaneous DNA damage, while excision of uracil is not required for cytotoxicity of fluoropyrimidines in mouse embryonic fibroblasts
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DOI：
10.1093/carcin/bgh347
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发表时间：
2005-03-01
期刊：
影响因子：
4.7
通讯作者：
Nilsen, H
中科院分区：
文献类型：
--
作者：
Andersen, S;Heine, T;Nilsen, H
DNA中的尿嘧啶可能是由于胞嘧啶脱氨或在复制过程中掺入dUMP而不是dTMP而产生的。我们研究了尿嘧啶-DNA糖基化酶缺陷小鼠的原代细胞和小鼠胚胎成纤维细胞（MEF）细胞系DNA中尿嘧啶的稳态水平。结果表明，尿嘧啶在Ung（-/-）细胞DNA中的水平强烈依赖于增殖，表明尿嘧啶残基主要来源于复制过程中的错误掺入。用5-氟-2 '-脱氧尿苷（5-FdUrd）或5-氟尿嘧啶（5-FU）处理引起Ung（-/-）MEF中尿嘧啶的剂量依赖性增加（高达1.5倍），但在野生型细胞中没有。有趣的是，Ung（-/-）MEF响应于5-FdUrd而不是5-FU积累AP位点以及尿嘧啶。这种修复中间体的积累表明在缺乏Ung的情况下失去了紧密协调的修复，并且与Ung（-/-）MEFs中与野生型细胞相比对5-FdUrd而不是5-FU的细胞增殖的更强抑制相关。然而，这些氟嘧啶的其他细胞毒性作用在野生型和Ung缺陷型细胞中相当，表明通过Ung尿嘧啶-DNA糖基化酶从DNA中切除尿嘧啶不是获得细胞毒性的先决条件。
Uracil may arise in DNA as a result of deamination of cytosine or through incorporation of dUMP instead of dTMP during replication. We have studied the steady-state levels of uracil in the DNA of primary cells and mouse embryonic fibroblast (MEF) cell lines from mice deficient in the Ung uracil-DNA glycosylase. The results show that the levels of uracil in the DNA of Ung(-/-) cells strongly depend on proliferation, indicating that the uracil residues originate predominantly from misincorporation during replication. Treatment with 5-fluoro-2'-deoxyuridine (5-FdUrd) or 5-fluorouracil (5-FU) gives rise to a dose-dependent increase of uracil in Ung(-/-) MEFs (up to 1.5-fold) but not in wild-type cells. Interestingly, Ung(-/-) MEFs accumulate AP-sites as well as uracil in response to 5-FdUrd but not to 5-FU. This accumulation of repair intermediates suggests a loss of tightly co-ordinated repair in the absence of Ung, and correlates with stronger inhibition of cell proliferation in response to 5-FdUrd, but not to 5-FU, in Ung(-/-) MEFs compared with wild-type cells. However, other cytotoxic effects of these fluoropyrimidines are comparable in both wild-type and Ung-deficient cells, demonstrating that excision of uracil from DNA by the Ung uracil-DNA glycosylase is not a prerequisite for obtaining cytotoxicity.