Department of Life and Environmental System Science, Graduate School of Nanobioscience, Yokohama City University, Yokohama 236-0027, Japan.
Department of Biology, Graduate School of Science, Chiba University, Chiba 263-8522, Japan.
Nat Commun. 2017 Jul 11;8:16112. doi: 10.1038/ncomms16112.
Homologous recombination-mediated gene targeting has greatly contributed to genetic analysis in a wide range of species, but is highly inefficient in human cells because of overwhelmingly frequent random integration events, whose molecular mechanism remains elusive. Here we show that DNA polymerase θ, despite its minor role in chromosomal DNA repair, substantially contributes to random integration, and that cells lacking both DNA polymerase θ and DNA ligase IV, which is essential for non-homologous end joining (NHEJ), exhibit 100% efficiency of spontaneous gene targeting by virtue of undetectable levels of random integration. Thus, DNA polymerase θ-mediated end joining is the sole homology-independent repair route in the absence of NHEJ and, intriguingly, their combined absence reveals rare Alu-Alu recombination events utilizing a stretch of homology. Our findings provide new insights into the mechanics of foreign DNA integration and the role of DNA polymerase θ in human genome maintenance.
同源重组介导的基因靶向在广泛的物种中极大地促进了遗传分析，但由于随机整合事件的频率极高，在人类细胞中效率非常低，其分子机制仍不清楚。在这里，我们表明，尽管 DNA 聚合酶θ在染色体 DNA 修复中作用较小，但它对随机整合有很大贡献，并且缺乏 DNA 聚合酶θ和 DNA 连接酶 IV 的细胞（后者对于非同源末端连接（NHEJ）是必不可少的），由于随机整合水平无法检测到，自发基因靶向的效率达到 100%。因此，在没有 NHEJ 的情况下，DNA 聚合酶θ介导的末端连接是唯一的非同源依赖性修复途径，有趣的是，它们的共同缺失揭示了利用同源性的罕见 Alu-Alu 重组事件。我们的发现为外源 DNA 整合的机制和 DNA 聚合酶θ在人类基因组维持中的作用提供了新的见解。
