Processing of a single ribonucleotide embedded into DNA by human nucleotide excision repair and DNA polymerase η
Processing of a single ribonucleotide embedded into DNA by human nucleotide excision repair and DNA polymerase η
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通过人核苷酸切除修复和 DNA 聚合酶 η 处理嵌入 DNA 的单个核糖核苷酸
DOI：
10.1038/s41598-019-50421-8
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发表时间：
2019
影响因子：
4.6
通讯作者：
Ura Kiyoe
中科院分区：
文献类型：
--
作者：
Sassa Akira;Tada Haruto;Takeishi Ayuna;Harada Kaho;Suzuki Megumi;Tsuda Masataka;Sasanuma Hiroyuki;Takeda Shunichi;Sugasawa Kaoru;Yasui Manabu;Honma Masamitsu;Ura Kiyoe
DNA聚合酶通常掺入非规范核苷酸，即，核糖核苷三磷酸进入基因组DNA。核糖核苷酸在基因组中的异常积累导致各种细胞异常。在这里，我们展示了人类核苷酸切除修复（NER）和DNA聚合酶η（Pol η）在嵌入DNA的单个核糖核苷酸加工中的可能作用。我们发现重组的NER系统可以切除质粒DNA上的氧化核糖核苷酸。结合Pol η精确绕过核糖核苷酸的证据，即，为了进一步研究核糖鸟苷（rG）或其氧化衍生物（8-oxo-rG）在缺乏NER或Pol η的人细胞中的致突变潜力，我们在体外对包埋的核糖核苷酸进行了研究。报告基因上的单个rG主要诱导大的缺失突变。嵌入的8-oxo-rG在3′-邻近碱基处引起碱基替换突变，而不是野生型细胞中的大缺失。XPA是NER或Pol η的重要因子，其破坏导致8-oxo-rG突变频率增加。此外，8-oxo-rG介导的大缺失的频率会因Pol η的缺失而增加，但XPA的缺失则不会。总的来说，我们的研究结果表明，嵌入的核糖核苷酸的碱基氧化能够通过替代DNA修复和损伤耐受途径来处理核糖核苷酸。
DNA polymerases often incorporate non-canonical nucleotide, i.e., ribonucleoside triphosphates into the genomic DNA. Aberrant accumulation of ribonucleotides in the genome causes various cellular abnormalities. Here, we show the possible role of human nucleotide excision repair (NER) and DNA polymerase η (Pol η) in processing of a single ribonucleotide embedded into DNA. We found that the reconstituted NER system can excise the oxidized ribonucleotide on the plasmid DNA. Taken together with the evidence that Pol η accurately bypasses a ribonucleotide, i.e., riboguanosine (rG) or its oxidized derivative (8-oxo-rG)in vitro, we further assessed the mutagenic potential of the embedded ribonucleotide in human cells lacking NER or Pol η. A single rG on thesupFreporter gene predominantly induced large deletion mutations. An embedded 8-oxo-rG caused base substitution mutations at the 3′-neighboring base rather than large deletions in wild-type cells. The disruption ofXPA, an essential factor for NER, or Pol η leads to the increased mutant frequency of 8-oxo-rG. Furthermore, the frequency of 8-oxo-rG-mediated large deletions was increased by the loss of Pol η, but notXPA. Collectively, our results suggest that base oxidation of the embedded ribonucleotide enables processing of the ribonucleotide via alternative DNA repair and damage tolerance pathways.