Kronenberg Michael, Carey Michael F
Department of Biological Chemistry, UCLA David Geffen School of Medicine, Los Angeles, CA 90095, USA.
Molecular Biology Institute, UCLA, Los Angeles, CA 90024, USA.
iScience. 2022 Dec 10;26(1):105791. doi: 10.1016/j.isci.2022.105791. eCollection 2023 Jan 20.
Head-on (HO) collisions between the DNA replication machinery and RNA polymerase over R-loop forming sequences (RLFS) are genotoxic, leading to replication fork blockage and DNA breaks. Current models suggest that HO collisions are avoided through replication initiation site (RIS) positioning upstream of active genes, ensuring co-orientation of replication fork movement and genic transcription. However, this model does not account for pervasive transcription, or intragenic RIS. Moreover, pervasive transcription initiation and CG-rich DNA is a feature of RIS, suggesting that HO transcription units (HO TUs) capable of forming R-loops might occur. Through mining phased GRO-seq data, and developing an informatics strategy to stringently identify RIS, we demonstrate that HO TUs containing RLFS occur at RIS in MCF-7 cells, and are downregulated at the G1/S phase boundary. Our analysis reveals a novel spatiotemporal relationship between transcription and replication, and supports the idea that HO collisions are avoided through transcriptional regulatory mechanisms.
DNA复制机器与RNA聚合酶在R环形成序列（RLFS）上发生的正面（HO）碰撞具有基因毒性，会导致复制叉阻滞和DNA断裂。目前的模型表明，通过将复制起始位点（RIS）定位在活跃基因的上游，可以避免HO碰撞，从而确保复制叉移动和基因转录的同向性。然而，该模型无法解释普遍存在的转录现象或基因内的RIS。此外，普遍存在的转录起始和富含CG的DNA是RIS的一个特征，这表明可能会出现能够形成R环的HO转录单元（HO TU）。通过挖掘阶段性的GRO-seq数据，并开发一种信息学策略来严格识别RIS，我们证明了含有RLFS的HO TU在MCF-7细胞的RIS处出现，并在G1/S期边界处下调。我们的分析揭示了转录与复制之间一种新的时空关系，并支持通过转录调控机制避免HO碰撞的观点。
