Sriramachandran Annie M, Petrosino Giuseppe, Méndez-Lago María, Schäfer Axel J, Batista-Nascimento Liliana S, Zilio Nicola, Ulrich Helle D
Institute of Molecular Biology (IMB), Ackermannweg 4, 55128 Mainz, Germany.
Institute of Molecular Biology (IMB), Ackermannweg 4, 55128 Mainz, Germany.
Mol Cell. 2020 Jun 4;78(5):975-985.e7. doi: 10.1016/j.molcel.2020.03.027. Epub 2020 Apr 21.
DNA single-strand breaks (SSBs) are among the most common lesions in the genome, arising spontaneously and as intermediates of many DNA transactions. Nevertheless, in contrast to double-strand breaks (DSBs), their distribution in the genome has hardly been addressed in a meaningful way. We now present a technique based on genome-wide ligation of 3'-OH ends followed by sequencing (GLOE-Seq) and an associated computational pipeline designed for capturing SSBs but versatile enough to be applied to any lesion convertible into a free 3'-OH terminus. We demonstrate its applicability to mapping of Okazaki fragments without prior size selection and provide insight into the relative contributions of DNA ligase 1 and ligase 3 to Okazaki fragment maturation in human cells. In addition, our analysis reveals biases and asymmetries in the distribution of spontaneous SSBs in yeast and human chromatin, distinct from the patterns of DSBs.
DNA单链断裂（SSB）是基因组中最常见的损伤类型之一，可自发产生，也是许多DNA交易过程中的中间体。然而，与双链断裂（DSB）不同，其在基因组中的分布几乎未得到有意义的研究。我们现在提出一种基于全基因组3'-OH末端连接并测序的技术（GLOE-Seq）以及一个相关的计算流程，该技术专为捕获SSB而设计，但通用性足以应用于任何可转化为游离3'-OH末端的损伤。我们证明了其在无需事先进行大小选择的情况下绘制冈崎片段图谱的适用性，并深入了解了DNA连接酶1和连接酶3对人类细胞中冈崎片段成熟的相对贡献。此外，我们的分析揭示了酵母和人类染色质中自发SSB分布的偏差和不对称性，这与DSB的模式不同。
