Replication-induced DNA secondary structures drive fork uncoupling and breakage.
Replication-induced DNA secondary structures drive fork uncoupling and breakage.
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DOI：
10.15252/embj.2023114334
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发表时间：
2023-11-15
期刊：
影响因子：
11.4
通讯作者：
Coster, Gideon
中科院分区：
文献类型：
--
作者：
Williams, Sophie L.;Casas-Delucchi, Corella S.;Raguseo, Federica;Guneri, Dilek;Li, Yunxuan;Minamino, Masashi;Fletcher, Emma E.;Yeeles, Joseph T. P.;Keyser, Ulrich F.;Waller, Zoe A. E.;Di Antonio, Marco;Coster, Gideon
关键词：
形成DNA二级结构的序列，如G-四链体（G4）和插入基序（iM），在人类基因组中丰富，并发挥各种生理作用。然而，它们也可以干扰复制并威胁基因组稳定性。多种证据表明G4抑制复制，但其潜在机制仍不清楚。此外，缺乏iMs如何影响复制体的证据。在这里，我们重建了生理衍生的结构形成序列的复制，发现单个G4或iM阻止了DNA复制。固态纳米孔内的直接单分子结构检测揭示了复制后形成的结构。结合遗传和生物物理特征确定结构稳定性和结构形成的概率是复制体停滞的关键决定因素。从机制上讲，复制停滞是由合成受损引起的，导致解旋酶-聚合酶解偶联。值得注意的是，iM还诱导新生DNA的断裂。最后，停滞的分叉只能被一种专门的解旋酶Pif 1拯救，而不能被Rrm 3、Sgs 1、Chl 1或Hrq 1拯救。总之，我们提供了一种机制，四链体结构的形成和分辨率在复制过程中，并强调G4和iMs作为内源性来源的复制应力。 体外重建表明，一个单一的生理G4或iM二级结构通过抑制前导链合成来阻止真核复制体。
Sequences that form DNA secondary structures, such as G‐quadruplexes (G4s) and intercalated‐Motifs (iMs), are abundant in the human genome and play various physiological roles. However, they can also interfere with replication and threaten genome stability. Multiple lines of evidence suggest G4s inhibit replication, but the underlying mechanism remains unclear. Moreover, evidence of how iMs affect the replisome is lacking. Here, we reconstitute replication of physiologically derived structure‐forming sequences to find that a single G4 or iM arrest DNA replication. Direct single‐molecule structure detection within solid‐state nanopores reveals structures form as a consequence of replication. Combined genetic and biophysical characterisation establishes that structure stability and probability of structure formation are key determinants of replisome arrest. Mechanistically, replication arrest is caused by impaired synthesis, resulting in helicase‐polymerase uncoupling. Significantly, iMs also induce breakage of nascent DNA. Finally, stalled forks are only rescued by a specialised helicase, Pif1, but not Rrm3, Sgs1, Chl1 or Hrq1. Altogether, we provide a mechanism for quadruplex structure formation and resolution during replication and highlight G4s and iMs as endogenous sources of replication stress. In vitro reconstitution shows that a single physiological G4 or iM secondary structure stalls the eukaryotic replisome by inhibiting leading strand synthesis.
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影响因子：
3.3
作者：
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Balasubramanian S
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Yeeles JTP
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通讯作者：
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通讯作者：
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