Dppa2 and Dppa4 directly regulate the Dux-driven zygotic transcriptional program
Dppa2 and Dppa4 directly regulate the Dux-driven zygotic transcriptional program
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DOI：
10.1101/431890
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发表时间：
2018-10
影响因子：
10.5
通讯作者：
M. Eckersley-Maslin;Celia Alda-Catalinas;Marloes Blotenburg;Elisa Kreibich;C. Krueger;W. Reik
中科院分区：
文献类型：
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作者：
M. Eckersley-Maslin;Celia Alda-Catalinas;Marloes Blotenburg;Elisa Kreibich;C. Krueger;W. Reik
哺乳动物合子基因组激活（ZGA）的分子调控仍然知之甚少。引发的小鼠胚胎干细胞含有罕见的“2C 样”细胞子集，它们在表观遗传和转录上与两细胞胚胎相似，因此代表了研究 ZGA 转录调控的理想系统。最近，仅在 ZGA 小波中表达的转录因子 Dux 被描述为激活许多下游 ZGA 转录物。然而，尚不清楚哪些上游母体因素以 Dux 依赖或独立方式启动 ZGA。在这里，我们进行了基于候选的过度表达筛选，其中鉴定了发育多能性相关 2 (Dppa2) 和 4 (Dppa4) 作为 2C 样细胞和 ZGA 转录的正调节因子。在种系中，启动子 DNA 去甲基化与 Dppa2 和 Dppa4 的上调同时发生，Dppa2 和 Dppa4 保持表达直至 E7.5，此时其启动子被重新甲基化。此外，Dppa2 和 Dppa4 也在 iPSC 重编程过程中表达，此时 2C 样 ZGA 转录瞬时达到峰值。通过结合过表达、敲低、敲除和拯救实验以及转录分析，我们表明 Dppa2 和 Dppa4 直接调节 2C 样细胞群和相关转录本，包括 Dux 和 Zscan4 簇。重要的是，我们梳理了 2C 样转录程序启动和稳定的分子层次结构。 Dppa2 和 Dppa4 需要 Dux 启动类似 2C 的 ZGA 转录，这表明它们通过直接调节 Dux 发挥上游作用。 ChIP-seq 分析显示 Dppa2 和 Dppa4 与 Dux 启动子和基因体结合并驱动其表达，支持了这一点。 Zscan4c 还能够在野生型细胞中诱导 2C 样细胞，但与 Dux 不同，在 Dppa2/4 双敲除细胞中不再能诱导 2C 样细胞，这表明它可能起到稳定转录网络的作用，而不是驱动转录网络。我们的研究结果提出了一个模型，其中 Dppa2/4 与 Dux 启动子的结合导致 Dux 上调并激活 2C 样转录程序，随后 Zscan4c 增强了该程序。
The molecular regulation of zygotic genome activation (ZGA) in mammals remains poorly understood. Primed mouse embryonic stem cells contain a rare subset of “2C-like” cells that are epigenetically and transcriptionally similar to the two cell embryo and thus represent an ideal system for studying ZGA transcription regulation. Recently, the transcription factor Dux, expressed exclusively in the minor wave of ZGA, was described to activate many downstream ZGA transcripts. However, it remains unknown what upstream maternal factors initiate ZGA either in a Dux dependent or independent manner. Here we performed a candidate-based overexpression screen, identifying, amongst others, Developmental Pluripotency Associated 2 (Dppa2) and 4 (Dppa4) as positive regulators of 2C-like cells and ZGA transcription. In the germ line, promoter DNA demethylation coincides with upregulation of Dppa2 and Dppa4 which remain expressed until E7.5 when their promoters are remethylated. Furthermore, Dppa2 and Dppa4 are also expressed during iPSC reprogramming at the time 2C-like ZGA transcription transiently peaks. Through a combination of overexpression, knockdown, knockout and rescue experiments, together with transcriptional analyses, we show that Dppa2 and Dppa4 directly regulate the 2C-like cell population and associated transcripts, including Dux and the Zscan4 cluster. Importantly, we tease apart the molecular hierarchy in which the 2C-like transcriptional programme is initiated and stabilised. Dppa2 and Dppa4 require Dux to initiate 2C-like ZGA transcription, suggesting they act upstream by directly regulating Dux. Supporting this, ChIP-seq analysis revealed Dppa2 and Dppa4 bind to the Dux promoter and gene body and drive its expression. Zscan4c is also able to induce 2C-like cells in wild type cells, but, in contrast to Dux, can no longer do so in Dppa2/4 double knockout cells, suggesting it may act to stabilise rather than drive the transcriptional network. Our findings suggest a model in which Dppa2/4 binding to the Dux promoter leads to Dux upregulation and activation of the 2C-like transcriptional programme which is subsequently reinforced by Zscan4c.