Yeast Tdp1 regulates the fidelity of nonhomologous end joining
Yeast Tdp1 regulates the fidelity of nonhomologous end joining
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DOI：
10.1073/pnas.0909917107
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发表时间：
2010-03-02
影响因子：
11.1
通讯作者：
Nitiss, John L.
中科院分区：
文献类型：
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作者：
Bahmed, Karim;Nitiss, Karin C.;Nitiss, John L.
酪氨酰-DNA-磷酸二酯酶1（Tdp 1）能使与DNA共价结合的肽段解离。我们评估了Tdp 1在非同源末端连接（NHEJ）中的作用，发现具有5'延伸的线性DNA分子在Delta tdp 1细胞中显示出高频率的错误修复。Deltatdp 1细胞中的连接错误主要是2-4个核苷酸的插入。具有3'延伸或平端的末端没有显示出增加的错误频率，尽管Delta tdp 1细胞修复平端DNA末端的效率高于WT细胞。我们发现，插入需要Ku 80和DNA连接酶IV，以及聚合酶IV。我们的研究结果表明，酵母Tdp 1是NHEJ途径的一个组成部分。我们认为Tdp 1 p 3'核苷酶活性通过产生3'磷酸调节DNA末端的加工，从而限制聚合酶和其他酶作用于DNA末端的能力。为了支持这一模型，我们发现酵母DNA 3'磷酸酶Tpp 1的过表达也导致更高频率的插入，这表明3'磷酸的产生是NHEJ期间Tdp 1介导的错误预防的关键步骤。
Tyrosyl-DNA-phosphodiesterase 1 (Tdp1) can disjoin peptides covalently bound to DNA. We assessed the role of Tdp1 in nonhomologous end joining (NHEJ) and found that linear DNA molecules with 5' extensions showed a high frequency of misrepair in Delta tdp1 cells. The joining errors in Delta tdp1 cells were predominantly 2-4 nucleotide insertions. Ends with 3' extensions or blunt ends did not show enhanced frequencies of errors, although Delta tdp1 cells repaired blunt DNA ends with greater efficiency than WT cells. We found that insertions required Ku80 and DNA ligase IV, as well as polymerase IV. Our results show that yeast Tdp1 is a component of the NHEJ pathway. We suggest that Tdp1p 3' nucleosidase activity regulates the processing of DNA ends by generating a 3' phosphate, thereby restricting the ability of polymerases and other enzymes from acting at DNA ends. In support of this model, we found that overexpression of Tpp1, a yeast DNA 3' phosphatase, also leads to a higher frequency of insertions, suggesting that the generation of a 3' phosphate is a key step in Tdp1-mediated error prevention during NHEJ.