Replication timing of siRNA control and siRNA PolQRKO cells. Homo sapiens

The human A-family DNA polymerase θ (Pol q) is a large, multidomain enzyme whose physiological function is still unclear despite its in vitro translesion synthesis capacity in front of DNA damage and its involvement in some features of DNA repair after external stress. Here we present evidence that Pol q holds a novel role in the absence of external stress as a critical determinant of the replication timing program in human cells. Pol q binds to chromatin at early G1 and is required for proper formation of pre-replicative complexe and replication origin activation. Pol q-depleted cells show modified spatial organization of chromatin-loop structures at replication factories. Genome-wide analysis of replication timing shows delayed replication of a part of early replicating domains and advanced replication of a part of late replicating domains following Pol q depletion. Our results identify Pol q as one of the first critical human factors discovered in the replication timing programme. Overall design: Two-condition experiment, siRNA control vs. siRNA polQ cells. Biological replicates: 2 control replicates, 2 transfected replicates.

人类A家族DNA聚合酶θ（DNA polymerase θ，Pol q）是一种大型多结构域酶。尽管其在体外具备绕过DNA损伤的跨损伤合成能力，并参与外源应激后的部分DNA修复过程，但其生理功能至今仍未明确。本研究提供证据表明，在无外源应激的条件下，Pol q在人类细胞的复制时序程序中发挥全新的关键调控作用。Pol q于早G1期结合染色质，对复制前复合物的正确组装以及复制起点的激活不可或缺。Pol q敲低细胞中，复制工厂处的染色质环结构空间组织发生异常改变。全基因组复制时序分析显示，敲低Pol q后，部分早期复制区域的复制进程出现延迟，而部分晚期复制区域的复制进程则提前。本研究结果将Pol q鉴定为人类复制时序程序中首个被发现的关键调控因子之一。实验整体设计：开展双条件对照实验，即小干扰RNA（siRNA）阴性对照组与siRNA polQ敲低组。生物学重复设置为：对照组2个生物学重复，转染组2个生物学重复。