Bi-allelic hypomorphic mutations in DNMT3B disrupt DNA methyltransferase activity and lead to immunodeficiency, centromeric instability, facial anomalies syndrome, type 1 (ICF1). Although several ICF1 phenotypes have been linked to abnormally hypomethylated repetitive regions, the unique genomic regions responsible for the remaining disease phenotypes remain largely uncharacterized. Here we explored two ICF1 patient–derived induced pluripotent stem cells (iPSCs) and their CRISPR-Cas9-corrected clones to determine whether DNMT3B correction can globally overcome DNA methylation defects and related changes in the epigenome. Hypomethylated regions throughout the genome are highly comparable between ICF1 iPSCs carrying different DNMT3B variants, and significantly overlap with those in ICF1 patient peripheral blood and lymphoblastoid cell lines. These regions include large CpG island domains, as well as promoters and enhancers of several lineage-specific genes, in particular immune-related, suggesting that they are premarked during early development. CRISPR-corrected ICF1 iPSCs reveal that the majority of phenotype-related hypomethylated regions reacquire normal DNA methylation levels following editing. However, at the most severely hypomethylated regions in ICF1 iPSCs, which also display the highest increases in H3K4me3 levels and/or abnormal CTCF binding, the epigenetic memory persists, and hypomethylation remains uncorrected. Overall, we demonstrate that restoring the catalytic activity of DNMT3B can reverse the majority of the aberrant ICF1 epigenome. However, a small fraction of the genome is resilient to this rescue, highlighting the challenge of reverting disease states that are due to genome-wide epigenetic perturbations. Uncovering the basis for the persistent epigenetic memory will promote the development of strategies to overcome this obstacle.

中文翻译：

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DNMT3B 突变的 ICF1 患者 iPSC 的异常表观基因组易于纠正，但具有基于 H3K4me3 和/或 CTCF 的表观遗传记忆的区域子集除外

DNMT3B的双等位基因亚型突变会破坏 DNA 甲基转移酶活性，导致免疫缺陷、着丝粒不稳定、1 型面部异常综合征 (ICF1)。尽管一些ICF1表型与异常低甲基化的重复区域有关，但导致其余疾病表型的独特基因组区域在很大程度上仍然未知。在这里，我们探索了两种 ICF1 患者衍生的诱导多能干细胞 (iPSC) 及其 CRISPR-Cas9 校正的克隆，以确定DNMT3B校正是否可以全局克服 DNA 甲基化缺陷和表观基因组的相关变化。整个基因组的低甲基化区域在携带不同DNMT3B变体的 ICF1 iPSC 之间具有高度可比性，并且与 ICF1 患者外周血和类淋巴母细胞系中的低甲基化区域显着重叠。这些区域包括大的 CpG 岛结构域，以及几个谱系特异性基因的启动子和增强子，特别是与免疫相关的基因，表明它们在早期发育过程中已被预先标记。CRISPR 校正的 ICF1 iPSC 显示，大多数表型相关的低甲基化区域在编辑后重新获得正常的 DNA 甲基化水平。然而，在 ICF1 iPSC 中最严重的低甲基化区域，这些区域也表现出 H3K4me3 水平的最高增加和/或异常的 CTCF 结合，表观遗传记忆仍然存在，低甲基化仍未得到纠正。总的来说，我们证明恢复 DNMT3B 的催化活性可以逆转大部分异常的 ICF1 表观基因组。然而，基因组的一小部分对这种拯救具有弹性，突显了恢复由于全基因组表观遗传扰动而导致的疾病状态的挑战。揭示持久表观遗传记忆的基础将促进克服这一障碍的策略的发展。