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首页> 外文期刊>Proceedings of the National Academy of Sciences of the United States of America >A role for FEN-1 in nonhomologous DNA end joining: The order of strand annealing and nucleolytic processing events
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A role for FEN-1 in nonhomologous DNA end joining: The order of strand annealing and nucleolytic processing events

机译：FEN-1在非同源DNA末端连接中的作用：链退火和核酸分解加工事件的顺序

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摘要

Eukaryotic repair of double-strand DNA breaks can occur either by homologous recombination or by nonhomologous DNA end joining (NHEJ). NHEJ relies on Ku70/86, XRCC4, DNA ligase IV, and DNA-dependent protein kinase. NHEJ involves a synapsis step in which the two ends are maintained in proximity, processing steps in which nucleases and polymerases act on the ends, an alignment step in which a few nudeotides of terminal homology guide the ends into pre- ferred alignments, and a ligation step. Some of the steps, such as Iigation, rely on a single enzymatic component. However, the processing steps begin and end with a wide array of alternative substrates and products, respectively, and likely involve multiple nucleases and polymerases. Given the alternative pathways that can be catalyzed by the remaining nucleases and polymerases, no one of these processing enzymes is likely to be essential. The only requirement for the processing enzymes, as a collective, is to generate a ligatable configuration, namely a ligatable nick on each strand. Here, we have tested the two major known 5' -spccific nucleases of Sacchoromyces cereivisiae for involvement in NHM. Whereas EXI1 does not appear to be involved to any detectable level, deleting R.AD27 (FEN-I of yeast) leads to a 4.4-fold reduction specifically of those NHEJ events predicted to proceed by means of 5' flap intermediates. Because Rail27/FEN-l acts specifically at 5' flap structures, thesE results suggest that the NHEJ alignment step precedes nucleolytic processing steps in a significant fraction of NHEJ events.
机译：双链DNA断裂的真核修复可通过同源重组或通过非同源DNA末端连接（NHEJ）进行。 NHEJ依赖Ku70 / 86，XRCC4，DNA连接酶IV和DNA依赖性蛋白激酶。 NHEJ涉及一个突触步骤，在该步骤中，两端保持邻近状态；加工步骤中，核酸酶和聚合酶作用于该末端；一个比对步骤，其中一些末端同源的裸核苷酸将这些末端引导为优选的比对；以及连接步。某些步骤（例如，导入）依赖于单个酶组分。然而，加工步骤分别以各种各样的替代底物和产物开始和结束，并且可能涉及多种核酸酶和聚合酶。考虑到剩余的核酸酶和聚合酶可以催化的替代途径，这些加工酶中的任何一种都可能不是必不可少的。总之，对加工酶的唯一要求是产生可连接的构型，即在每条链上可连接的切口。在这里，我们已经测试了酿酒酵母的两种主要已知的5'-特异性核酸酶是否参与NHM。尽管EXI1似乎未涉及任何可检测的水平，但删除R.AD27（酵母的FEN-1）特别导致那些通过5'襟翼中间体预测发生的NHEJ事件降低了4.4倍。因为Rail27 / FEN-1特异性地作用于5'襟翼结构，所以该sE结果表明，在相当一部分NHEJ事件中，NHEJ比对步骤先于核酸裂解步骤。

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