Mitotic inheritance of the DNA methylome is a challenging task for the maintenance of cell identity. Whether DNA methylation pattern in different genomic contexts can all be faithfully maintained is an open question. A replication-coupled DNA methylation maintenance model was proposed decades ago, but some observations suggest that a replication-uncoupled maintenance mechanism exists. However, the capacity and the underlying molecular events of replication-uncoupled maintenance are unclear. By measuring maintenance kinetics at the single-molecule level and assessing mutant cells with perturbation of various mechanisms, we found that the kinetics of replication-coupled maintenance are governed by the UHRF1–Ligase 1 and PCNA–DNMT1 interactions, whereas nucleosome occupancy and the interaction between UHRF1 and methylated H3K9 specifically regulate replication-uncoupled maintenance. Surprisingly, replication-uncoupled maintenance is sufficiently robust to largely restore the methylome when replication-coupled maintenance is severely impaired. However, solo-WCGW sites and other CpG sites displaying aging- and cancer-associated hypomethylation exhibit low maintenance efficiency, suggesting that although quite robust, mitotic inheritance of methylation is imperfect and that this imperfection may contribute to selective hypomethylation during aging and tumorigenesis.

DNA 甲基化组的有丝分裂遗传是维持细胞身份的一项具有挑战性的任务。不同基因组环境中的 DNA 甲基化模式是否都能忠实地保持是一个悬而未决的问题。几十年前提出了复制耦合的 DNA 甲基化维持模型，但一些观察表明存在复制非耦合维持机制。然而，复制非耦合维持的能力和潜在分子事件尚不清楚。通过测量单分子水平的维持动力学并评估各种机制扰动的突变细胞，我们发现复制耦合维持的动力学受 UHRF1-Ligase 1 和 PCNA-DNMT1 相互作用的控制，而核小体占据以及 UHRF1 和甲基化 H3K9 之间的相互作用专门调节复制非耦合维持。令人惊讶的是，当复制耦合维护严重受损时，复制非耦合维护足够强大以在很大程度上恢复甲基化组。然而，显示衰老和癌症相关低甲基化的 solo-WCGW 位点和其他 CpG 位点表现出低维持效率，这表明尽管甲基化的有丝分裂遗传非常稳健，但这种缺陷可能有助于衰老和肿瘤发生过程中的选择性低甲基化。