The major roles of DNA polymerases epsilon and delta at the eukaryotic replication fork are evolutionarily conserved.
The major roles of DNA polymerases epsilon and delta at the eukaryotic replication fork are evolutionarily conserved.
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DOI：
10.1371/journal.pgen.1002407
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发表时间：
2011-12
期刊：
影响因子：
4.5
通讯作者：
Carr AM
中科院分区：
文献类型：
--
作者：
Miyabe I;Kunkel TA;Carr AM
真核基因组的协调是不对称的，在不连续的滞后链中，我们提供了两种类型的证据，表明在裂变酵母中，这两个生物合成任务是由两个不同的复制酶执行的。编码POLδ-L591M突变器等位基因的菌株，位于相反方向的报告基因中的碱基取代相对于特征良好的复制来源，是特定于链特异性的，并以模式分布，这意味着POLδ主要参与滞后的链复制，其次是编码POLε-M630F等位基因的菌株在RNase H2中，在新生的前导链DNA中有选择地观察到RNMP。通常，以RNase H2依赖性方式去除。保守。
重要的是要了解DNA复制机制的结构，以及所有组织中的最新工作是否是酿酒酵母的最新工作。使用类似的遗传测定法证明，在高度进化的酵母菌sichizosacachomyces pombe中，POLδ相似重要的是，我们建立了一种新颖的物理测定法，将RNMP掺入新复制的DNA中，这表明POLε负责领先的链综合，并且没有显着贡献滞后的链条复制，并巩固了对先前遗传数据的解释。并表明聚合酶之间的劳动分裂是通过进化来保存的。
Coordinated replication of eukaryotic genomes is intrinsically asymmetric, with continuous leading strand synthesis preceding discontinuous lagging strand synthesis. Here we provide two types of evidence indicating that, in fission yeast, these two biosynthetic tasks are performed by two different replicases. First, in Schizosaccharomyces pombe strains encoding a polδ-L591M mutator allele, base substitutions in reporter genes placed in opposite orientations relative to a well-characterized replication origin are strand-specific and distributed in patterns implying that Polδ is primarily involved in lagging strand replication. Second, in strains encoding a polε-M630F allele and lacking the ability to repair rNMPs in DNA due to a defect in RNase H2, rNMPs are selectively observed in nascent leading strand DNA. The latter observation demonstrates that abundant rNMP incorporation during replication can be tolerated and that they are normally removed in an RNase H2-dependent manner. This provides strong physical evidence that Polε is the primary leading strand replicase. Collectively, these data and earlier results in budding yeast indicate that the major roles of Polδ and Polε at the eukaryotic replication fork are evolutionarily conserved. It is important to understand the architecture of the DNA replication machinery and whether this is common to all organisms. Recent work in Saccharomyces cerevisiae has genetically assigned specific DNA polymerases to leading and lagging strand DNA synthesis, Polε and Polε respectively. In this manuscript, we use a similar genetic assay to demonstrate that, in the highly evolutionarily diverged yeast Schizosaccharomyces pombe, Polδ is similarly responsible for lagging strand synthesis. Importantly, we establish a novel physical assay, the incorporation of rNMPs into newly replicated DNA, which demonstrates that Polε is responsible for leading strand synthesis and does not contribute significantly to lagging strand replication. These data strongly support and consolidate the interpretation of previous genetic data and suggest that the division of labour between polymerases is conserved through evolution.
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