Yeast Nucleolin Nsr1 Impedes Replication and Elevates Genome Instability at an Actively Transcribed Guanine-Rich G4 DNA-Forming Sequence.
Yeast Nucleolin Nsr1 Impedes Replication and Elevates Genome Instability at an Actively Transcribed Guanine-Rich G4 DNA-Forming Sequence.
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DOI：
10.1534/genetics.120.303736
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发表时间：
2020-12
期刊：
影响因子：
3.3
通讯作者：
Kim N
中科院分区：
文献类型：
--
作者：
Singh S;Berroyer A;Kim M;Kim N
基因组不稳定性的显著增加与含鸟嘌呤的DNA链构象转变为四链G-四链体（G4）DNA有关。G4 DNA在体内形成后的重组和基因组重排的机制一直难以阐明，但通过几种关键的G4 DNA结合蛋白的鉴定和功能表征已经变得更清楚。哺乳动物核仁素（NCL）是一种高度特异性的G4 DNA结合蛋白，在基因启动子处具有相关G4 DNA形成序列基序的基因的转录调控中具有明确的作用。G4 DNA和核仁素之间的体内相互作用对基因组不稳定性的影响以前没有研究过。我们在这里表明，酵母核仁素Nsr 1是丰富的G4 DNA形成序列在体内，是一个主要因素，诱导基因组不稳定性与共转录形成的G4 DNA在酵母基因组中。我们还表明，Nsr 1的结果阻碍复制过去这样的G4 DNA形成序列。G4相关的基因组不稳定性和G4 DNA在体内的结合需要位于Nsr 1蛋白C末端的甘氨酸-甘氨酸-甘氨酸（RGG）重复序列。缺失RGG结构域的Nsr 1支持正常细胞生长，并足以发挥其前rRNA加工功能。然而，Nsr 1的RGG结构域的截短显著削弱了其与G4 DNA在体内的相互作用，并恢复了不受阻碍的复制，总体上导致与富含鸟嘌呤的G4 DNA形成序列相关的基因组不稳定性急剧降低。我们的数据表明，Nsr 1与完整的RGG重复序列和G4 DNA之间的相互作用损害基因组的稳定性，排除访问G4-解决蛋白和阻碍复制。
A significant increase in genome instability is associated with the conformational shift of a guanine-run-containing DNA strand into the four-stranded G-quadruplex (G4) DNA. The mechanism underlying the recombination and genome rearrangements following the formation of G4 DNA in vivo has been difficult to elucidate but has become better clarified by the identification and functional characterization of several key G4 DNA-binding proteins. Mammalian nucleolin (NCL) is a highly specific G4 DNA-binding protein with a well-defined role in the transcriptional regulation of genes with associated G4 DNA-forming sequence motifs at their promoters. The consequence of the in vivo interaction between G4 DNA and nucleolin in respect to the genome instability has not been previously investigated. We show here that the yeast nucleolin Nsr1 is enriched at a G4 DNA-forming sequence in vivo and is a major factor in inducing the genome instability associated with the cotranscriptionally formed G4 DNA in the yeast genome. We also show that Nsr1 results in impeding replication past such a G4 DNA-forming sequence. The G4-associated genome instability and the G4 DNA-binding in vivo require the arginine-glycine-glycine (RGG) repeats located at the C-terminus of the Nsr1 protein. Nsr1 with the deletion of RGG domain supports normal cell growth and is sufficient for its pre-rRNA processing function. However, the truncation of the RGG domain of Nsr1 significantly weakens its interaction with G4 DNA in vivo and restores unhindered replication, overall resulting in a sharp reduction in the genome instability associated with a guanine-rich G4 DNA-forming sequence. Our data suggest that the interaction between Nsr1 with the intact RGG repeats and G4 DNA impairs genome stability by precluding the access of G4-resolving proteins and impeding replication.