Dual roles for DNA polymerase theta in alternative end-joining repair of double-strand breaks in Drosophila.
Dual roles for DNA polymerase theta in alternative end-joining repair of double-strand breaks in Drosophila.
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DOI：
10.1371/journal.pgen.1001005
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发表时间：
2010-07-01
期刊：
影响因子：
4.5
通讯作者：
McVey M
中科院分区：
文献类型：
--
作者：
Chan SH;Yu AM;McVey M
DNA双链断裂通过多种机制修复，这些机制大致分为同源性定向修复和非同源性末端连接两类。末端连接修复可以进一步分类为经典的非同源末端连接，其需要DNA连接酶4，或“替代”末端连接，其不需要。可变末端连接与基因组缺失和易位有关，但其分子机制在很大程度上尚未表征。在这里，我们报告说，果蝇DNA聚合酶θ（pol θ），编码的mus308基因和以前牵连在DNA链间交联修复，起着至关重要的作用，在DNA连接酶4独立的替代末端连接。在缺乏pol θ的情况下，末端连接受损，并且残余修复通常在断裂位点侧翼产生大的缺失。对带有mus308功能分离等位基因的果蝇的断裂修复连接的分析表明，pol theta促进了替代末端连接过程中长微同源性的使用，并增加了复杂插入事件的可能性。我们的研究结果建立pol θ作为一个关键蛋白在果蝇的替代末端连接，并建议一个潜在的机械连接替代末端连接和链间交联修复。DNA双链断裂，其中DNA双螺旋的两条链都被切断，必须被识别并准确修复，以促进细胞存活并防止突变的积累。然而，即使在可以进行准确修复的情况下，也会偶尔发生容易出错的修复。我们已经调查了一个易错的断裂修复机制称为替代结束连接的遗传要求。我们以前已经表明，替代末端连接是经常使用的果蝇，果蝇。在这里，我们证明了一种名为DNA聚合酶θ的果蝇蛋白质是这种不准确修复机制的关键参与者。遗传分析表明，聚合酶θ可能对与交替末端连接相关的两个过程很重要：（1）短互补DNA序列的退火，以及（2）在断裂修复位点产生小插入的DNA合成。在没有聚合酶theta的情况下，揭示了经常导致大染色体缺失的备份修复机制。由于DNA聚合酶theta在许多类型的人类癌症中高度表达，我们的研究结果为进一步研究聚合酶theta如何参与可能促进癌症发展的修复过程奠定了基础。
DNA double-strand breaks are repaired by multiple mechanisms that are roughly grouped into the categories of homology-directed repair and non-homologous end joining. End-joining repair can be further classified as either classical non-homologous end joining, which requires DNA ligase 4, or “alternative” end joining, which does not. Alternative end joining has been associated with genomic deletions and translocations, but its molecular mechanism(s) are largely uncharacterized. Here, we report that Drosophila melanogaster DNA polymerase theta (pol theta), encoded by the mus308 gene and previously implicated in DNA interstrand crosslink repair, plays a crucial role in DNA ligase 4-independent alternative end joining. In the absence of pol theta, end joining is impaired and residual repair often creates large deletions flanking the break site. Analysis of break repair junctions from flies with mus308 separation-of-function alleles suggests that pol theta promotes the use of long microhomologies during alternative end joining and increases the likelihood of complex insertion events. Our results establish pol theta as a key protein in alternative end joining in Drosophila and suggest a potential mechanistic link between alternative end joining and interstrand crosslink repair. DNA double-strand breaks, in which both strands of the DNA double helix are cut, must be recognized and accurately repaired in order to promote cell survival and prevent the accumulation of mutations. However, error-prone repair occasionally occurs, even when accurate repair is possible. We have investigated the genetic requirements of an error-prone break-repair mechanism called alternative end joining. We have previously shown that alternative end joining is frequently used in the fruit fly, Drosophila melanogaster. Here, we demonstrate that a fruit fly protein named DNA polymerase theta is a key player in this inaccurate repair mechanism. Genetic analysis suggests that polymerase theta may be important for two processes associated with alternative end joining: (1) annealing at short, complementary DNA sequences, and (2) DNA synthesis that creates small insertions at break-repair sites. In the absence of polymerase theta, a backup repair mechanism that frequently results in large chromosome deletions is revealed. Because DNA polymerase theta is highly expressed in many types of human cancers, our findings lay the groundwork for further investigations into how polymerase theta is involved in repair processes that may promote the development of cancer.
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