Single cell RNA-Seq of Hira-KO and control MII oocytes [Zp3-Cre]

The integrity of chromatin, which provides a dynamic template for all DNA-related processes in eukaryotes, is maintained through replication-dependent and -independent assembly pathways. To address the role of replication-independent histone deposition, we deleted the histone H3.3 chaperone Hira in developing mouse oocytes. We show that chromatin of non-replicative developing oocytes is highly dynamic, and that lack of continuous H3.3/H4 deposition alters chromatin structure, resulting in increased DNase I sensitivity, the accumulation of DNA damage, and, ultimately, a severe fertility phenotype. On the molecular level, abnormal chromatin structure leads to a dramatic decrease in the dynamic range of gene expression, the appearance of spurious transcripts, and inefficient de novo DNA methylation. In contrast to the only minor transcriptional phenotype observed in mouse pluripotent cells, we unequivocally show the importance of histone replacement and chromatin homeostasis for transcriptional regulation and normal developmental progression in an in vivo context. RNA-Seq on 4 Hiraf/f and 4 Hiraf/f, Zp3-Cre single MII oocytes.染色质（chromatin）作为真核生物（eukaryotes）所有DNA相关进程的动态模板，其完整性经由依赖复制与不依赖复制的组装通路维持。为探究不依赖复制的组蛋白沉积的功能，我们在发育中小鼠卵母细胞中敲除了组蛋白H3.3分子伴侣Hira。本研究证实，非复制型发育卵母细胞的染色质具有高度动态特性；若持续缺失H3.3/H4沉积，则会改变染色质结构，引发脱氧核糖核酸酶I（DNase I）敏感性升高、DNA损伤积累，并最终导致严重的生育表型。在分子层面，异常染色质结构会造成基因表达动态范围大幅收窄、异常转录本出现，以及从头DNA甲基化效率低下。与小鼠多能细胞中仅观察到轻微转录表型的情况不同，本研究明确证实了组蛋白替换与染色质稳态在体内环境中对转录调控及正常发育进程的重要性。对4个Hiraf/f及4个Hiraf/f; Zp3-Cre基因型小鼠的单个MII期卵母细胞开展RNA测序（RNA-Seq）。