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The interplay of RNA:DNA hybrid structure and G-quadruplexes determines the outcome of R-loop-replisome collisions
eLife ( IF 6.4 ) Pub Date : 2021-09-08 , DOI: 10.7554/elife.72286
Charanya Kumar 1 , Sahil Batra 1 , Jack D Griffith 2 , Dirk Remus 1
Affiliation  

R-loops are a major source of genome instability associated with transcription-induced replication stress. However, how R-loops inherently impact replication fork progression is not understood. Here, we characterize R-loop-replisome collisions using a fully reconstituted eukaryotic DNA replication system. We find that RNA:DNA hybrids and G-quadruplexes at both co-directional and head-on R-loops can impact fork progression by inducing fork stalling, uncoupling of leading strand synthesis from replisome progression, and nascent strand gaps. RNase H1 and Pif1 suppress replication defects by resolving RNA:DNA hybrids and G-quadruplexes, respectively. We also identify an intrinsic capacity of replisomes to maintain fork progression at certain R-loops by unwinding RNA:DNA hybrids, repriming leading strand synthesis downstream of G-quadruplexes, or utilizing R-loop transcripts to prime leading strand restart during co-directional R-loop-replisome collisions. Collectively, the data demonstrates that the outcome of R-loop-replisome collisions is modulated by R-loop structure, providing a mechanistic basis for the distinction of deleterious from non-deleterious R-loops.

中文翻译:

RNA:DNA 杂交结构和 G-四链体的相互作用决定了 R-loop-replisome 碰撞的结果

R 环是与转录诱导的复制应激相关的基因组不稳定性的主要来源。然而,R 环如何固有地影响复制叉进程尚不清楚。在这里,我们使用完全重组的真核 DNA 复制系统来表征 R 环复制体碰撞。我们发现,共向和正面 R 环上的 RNA:DNA 杂合体和 G-四链体可以通过诱导叉停滞、前导链合成与复制体进展的解偶联以及新生链间隙来影响叉进展。RNase H1 和 Pif1 分别通过解析 RNA:DNA 杂交体和 G-四链体来抑制复制缺陷。我们还确定了复制体通过解开 RNA:DNA 杂交体、重新启动 G-四链体下游的前导链合成来维持某些 R 环的叉进展的内在能力,或利用 R 环转录本在同向 R 环-复制体碰撞期间引发前导链重新启动。总的来说,数据表明 R-loop-replisome 碰撞的结果受 R-loop 结构的调节,为区分有害和非有害 R-loop 提供了机制基础。
更新日期:2021-09-08
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