Holliday junction-containing DNA structures persist in cells lacking Sgs1 or Top3 following exposure to DNA damage
Holliday junction-containing DNA structures persist in cells lacking Sgs1 or Top3 following exposure to DNA damage
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DOI：
10.1073/pnas.1014240108
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发表时间：
2011-03-22
影响因子：
11.1
通讯作者：
Hickson, Ian D.
中科院分区：
文献类型：
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作者：
Mankouri, Hocine W.;Ashton, Thomas M.;Hickson, Ian D.
Sgs 1-Rmi 1-Top3“溶解体”是维持基因组稳定性所必需的，并且涉及在DNA复制期间产生的各种类型的DNA结构的加工。以前的研究表明，未加工的（X形）同源重组修复（HRR）的中间体持续存在时，S-相被干扰使用甲基甲磺酸盐（MMS）在酿酒酵母细胞受损的Sgs 1或Top3。然而，这些持久的DNA结构的确切性质仍然很差的特点。在这里，我们报告，异位表达的两个异源和结构无关的霍利迪连接（HJ）分解酶，大肠杆菌RusA或人类GEN 1（1-527），促进这些X-结构在体内的去除。此外，其他类型的DNA复制中间体，包括停滞的复制叉和非HRR依赖性X结构，对RusA或GEN 1（1-527）是难治的，证明了这些HJ解离酶对MMS诱导的体内X结构的特异性。这些数据表明，X-结构持续存在于受损的Sgs 1或Top3细胞含有HJ。此外，我们表明，Sgs 1直接促进X-结构的去除，因为在Sgs 1缺陷菌株中产生的持久结构被消除时，Sgs 1在体内被重新激活。我们建议，HJ resolvases和Sgs 1-Top3-Rmi 1包括两个独立的过程来处理HRR过程中产生的含HJ的DNA中间体在S-阶段。
The Sgs1-Rmi1-Top3 "dissolvasome" is required for the maintenance of genome stability and has been implicated in the processing of various types of DNA structures arising during DNA replication. Previous investigations have revealed that unprocessed (X-shaped) homologous recombination repair (HRR) intermediates persist when S-phase is perturbed by using methyl methanesulfonate (MMS) in Saccharomyces cerevisiae cells with impaired Sgs1 or Top3. However, the precise nature of these persistent DNA structures remains poorly characterized. Here, we report that ectopic expression of either of two heterologous and structurally unrelated Holliday junction (HJ) resolvases, Escherichia coli RusA or human GEN1(1-527), promotes the removal of these X-structures in vivo. Moreover, other types of DNA replication intermediates, including stalled replication forks and non-HRR-dependent X-structures, are refractory to RusA or GEN1(1-527), demonstrating specificity of these HJ resolvases for MMS-induced X-structures in vivo. These data suggest that the X-structures persisting in cells with impaired Sgs1 or Top3 contain HJs. Furthermore, we demonstrate that Sgs1 directly promotes X-structure removal, because the persistent structures arising in Sgs1-deficient strains are eliminated when Sgs1 is reactivated in vivo. We propose that HJ resolvases and Sgs1-Top3-Rmi1 comprise two independent processes to deal with HJ-containing DNA intermediates arising during HRR in S-phase.