Trypanosomes lacking uracil-DNA glycosylase are hypersensitive to antifolates and present a mutator phenotype
Trypanosomes lacking uracil-DNA glycosylase are hypersensitive to antifolates and present a mutator phenotype
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DOI：
10.1016/j.biocel.2012.06.014
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发表时间：
2012-09-01
影响因子：
4
通讯作者：
Gonzalez-Pacanowska, Dolores
中科院分区：
文献类型：
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作者：
Castillo-Acosta, Victor M.;Aguilar-Pereyra, Fernando;Gonzalez-Pacanowska, Dolores
细胞在DNA中含有少量尿嘧啶，这可能是复制过程中dUTP错误掺入或胞嘧啶脱氨基的结果。在碱基切除修复途径中消除尿嘧啶产生脱碱基位点，除非修复，否则其具有潜在的致突变性。布氏锥虫基因组呈现单个尿嘧啶-DNA糖基化酶，其负责从DNA去除尿嘧啶。在这里，我们建立了没有切除活性检测U：G，U：A对或单链尿嘧啶的DNA尿嘧啶-DNA糖基化酶无效突变体细胞提取物，表明备份尿嘧啶切除活动的情况下。虽然原环形式可以在DNA中含有适量尿嘧啶的情况下存活，但突变细胞中突变率和光谱的分析揭示了超变子表型，其中主要事件是GC至AT转换和插入。通过碱基切除修复途径消除尿嘧啶的缺陷引起对抗叶酸剂和氧化应激的超敏反应，并增加DNA链断裂的数量，表明替代DNA修复途径的激活。最后，我们表明，尿嘧啶-DNA糖基化酶缺陷的细胞表现出降低体内感染性证明，有效的尿嘧啶消除是重要的哺乳动物宿主内的生存。(C)2012爱思唯尔有限公司保留所有权利。
Cells contain low amounts of uracil in DNA which can be the result of dUTP misincorporation during replication or cytosine deamination. Elimination of uracil in the base excision repair pathway yields an abasic site, which is potentially mutagenic unless repaired. The Trypanosoma brucei genome presents a single uracil-DNA glycosylase responsible for removal of uracil from DNA. Here we establish that no excision activity is detected on U:G, U:A pairs or single-strand uracil-containing DNA in uracil-DNA glycosylase null mutant cell extracts, indicating the absence of back-up uracil excision activities. While procyclic forms can survive with moderate amounts of uracil in DNA, an analysis of the mutation rate and spectra in mutant cells revealed a hypermutator phenotype where the predominant events were GC to AT transitions and insertions. Defective elimination of uracil via the base excision repair pathway gives rise to hypersensitivity to antifolates and oxidative stress and an increased number of DNA strand breaks, suggesting the activation of alternative DNA repair pathways. Finally, we show that uracil-DNA glycosylase defective cells exhibit reduced infectivity in vivo demonstrating that efficient uracil elimination is important for survival within the mammalian host. (C) 2012 Elsevier Ltd. All rights reserved.