Early mammalian development entails genome-wide epigenome remodeling, including DNA methylation erasure and reacquisition, which facilitates developmental competence. To uncover the mechanisms that orchestrate DNA methylation dynamics, we coupled a single-cell ratiometric DNA methylation reporter with unbiased CRISPR screening in murine embryonic stem cells (ESCs). We identify key genes and regulatory pathways that drive global DNA hypomethylation, and characterize roles for Cop1 and Dusp6. We also identify Dppa2 and Dppa4 as essential safeguards of focal epigenetic states. In their absence, developmental genes and evolutionarily young LINE1 elements, which are specifically bound by DPPA2, lose H3K4me3 and gain ectopic de novo DNA methylation in pluripotent cells. Consequently, lineage-associated genes and LINE1 acquire a repressive epigenetic memory, which renders them incompetent for activation during future lineage specification. Dppa2/4 thereby sculpt the pluripotent epigenome by facilitating H3K4me3 and bivalency to counteract de novo methylation, a function co-opted by evolutionarily young LINE1 to evade epigenetic decommissioning.

早期哺乳动物发育需要全基因组表观基因组重塑，包括 DNA 甲基化擦除和重新获取，这有助于发育能力。为了揭示协调 DNA 甲基化动力学的机制，我们将单细胞比率 DNA 甲基化报告基因与小鼠胚胎干细胞 (ESC) 中的无偏 CRISPR 筛选相结合。我们确定了驱动全球 DNA 低甲基化的关键基因和调控途径，并描述了Cop1和Dusp6的作用。我们还识别Dppa2和Dppa4作为焦点表观遗传状态的基本保障。在它们缺失的情况下，与 DPPA2 特异性结合的发育基因和进化上年轻的 LINE1 元素会失去 H3K4me3 并在多能细胞中获得异位从头 DNA 甲基化。因此，谱系相关基因和 LINE1 获得了一种抑制性的表观遗传记忆，这使得它们在未来的谱系规范中无法激活。Dppa2 / 4从而通过促进 H3K4me3 和二价性来抵消从头甲基化来塑造多能表观基因组，这是进化上年轻的 LINE1 为逃避表观遗传退役而选择的一种功能。