Department of Pathology, University of Michigan Medical School, Ann Arbor, MI, 48109, USA.
Department of Human Genetics, University of Michigan Medical School, Ann Arbor, MI, 48109, USA.
Nat Commun. 2024 Nov 6;15(1):9582. doi: 10.1038/s41467-024-53917-8.
Genomic structural variants (SVs) greatly impact human health, but much is unknown about the mechanisms that generate the largest class of nonrecurrent alterations. Common fragile sites (CFSs) are unstable loci that provide a model for SV formation, especially large deletions, under replication stress. We study SV junction formation as it occurs in human cell lines by applying error-minimized capture sequencing to CFS DNA harvested after low-dose aphidicolin treatment. SV junctions form throughout CFS genes at a 5-fold higher rate after cells pass from G2 into M-phase. Neither SV formation nor CFS expression depend on mitotic DNA synthesis (MiDAS), an error-prone form of replication active at CFSs. Instead, analysis of tens of thousands of de novo SV junctions combined with DNA repair pathway inhibition reveal a primary role for DNA polymerase theta (POLQ)-mediated end-joining (TMEJ). We propose an important role for mitotic TMEJ in nonrecurrent SV formation genome wide.
基因组结构变异（SVs）对人类健康有重大影响，但对于产生最大一类非重复改变的机制知之甚少。常见脆弱位点（CFSs）是不稳定的基因座，在复制应激下为 SV 形成提供了模型，特别是大的缺失。我们通过应用最小化错误的捕获测序来研究人类细胞系中 SV 连接点的形成，这些 SV 连接点是在低剂量阿霉素处理后从 CFS DNA 中提取的。SV 连接点在细胞从 G2 进入 M 期后，在 CFS 基因中以 5 倍的速率形成。SV 的形成和 CFS 的表达都不依赖于有丝分裂 DNA 合成（MiDAS），这是 CFS 中活跃的易错复制形式。相反，对数万条新出现的 SV 连接点的分析，结合 DNA 修复途径抑制，揭示了 DNA 聚合酶θ（POLQ）介导的末端连接（TMEJ）的主要作用。我们提出有丝分裂 TMEJ 在全基因组中非重复 SV 形成中的重要作用。
