Reprogramming of H3K9me3-dependent heterochromatin during mammalian early embryo development [RNA-seq]. Mus musculus

H3K9me3-dependent heterochromatin is considered as one of the major barriers for cell fate changes, and must be reprogrammed during fertilization to reactivate highly specialized paternal and maternal genome to establish totipotency. However, the molecular details are lacked for early embryos due to the limited materials. Here we map the genome-wide distribution of H3K9me3 modification in the early embryo as well as in the cell fate determined embryonic tissues after implantation. We find that H3K9me3 exhibits distinct dynamic features in promoters and retro-transposons. Both maternal and paternal genome undergo large scale of H3K9me3 reestablishment after fertilization, and the imbalance of maternal H3K9me3 signal over paternal last until the blastocyst stage. The rebuilding of H3K9me3 on LTR retro-transposons maintains its repression state after the global DNA demethylation, and we further discover that Chaf1a is essential for the establishment of H3K9me3 on LTRs and the loss function of Chaf1a leads to embryo development failure. Finally, we find that lineage specific H3K9me3 is established after lineage commitment in post-implantation embryos. Thus, our data demonstrate that H3K9me3-dependent heterochromatin undergoes dramatic reprogramming during early embryo development and the establishment of H3K9me3 on LTRs is essential for proper embryo development. Overall design: We mapped the H3K9me3 modifications on embryos from zygote to 8.5 day stage (with separated ICM and TE, Epi and Exe). The mouse metaphase II (MII) oocytes, sperm, as well as mouse embryonic stem cells (mESCs) and mouse trophoblast stem cells (mTSCs) were also analyzed. In the ChIP-seq analysis on pre-implantation embryos, 500 cells were used for per reaction and two or three replicates were performed for each stage. This series contains the RNA-seq data on embryos from embryos from zygote to 6.5 day stage (with separated ICM and TE, Epi and Exe) including morula stage with siRNA.

依赖组蛋白H3赖氨酸9三甲基化（H3K9me3）的异染色质被认为是细胞命运转变的主要障碍之一，在受精过程中必须被重编程，以重新激活高度特化的父本与母本基因组，从而建立全能性。但受限于样本稀缺，早期胚胎中的相关分子细节至今仍未被充分解析。本研究绘制了早期胚胎以及着床后细胞命运定型的胚胎组织中H3K9me3修饰的全基因组分布图谱。研究发现，H3K9me3在启动子区域与逆转座子（retro-transposons）中呈现出截然不同的动态变化特征。受精后，父本与母本基因组均经历了大规模的H3K9me3重建立过程，且母本H3K9me3信号相较于父本的失衡状态会持续至囊胚阶段。在全基因组DNA去甲基化后，长末端重复序列（Long Terminal Repeat, LTR）逆转座子上的H3K9me3重建过程维持了其转录抑制状态；进一步研究发现，染色质组装因子1a（Chaf1a）是LTR区域H3K9me3建立的必需因子，其功能缺失会导致胚胎发育失败。此外，我们发现着床后胚胎的细胞谱系定型过程中，会建立谱系特异性的H3K9me3修饰。综上，本研究数据表明，依赖H3K9me3的异染色质在早期胚胎发育过程中经历了剧烈的重编程，且LTR区域H3K9me3的建立对于胚胎正常发育至关重要。

实验设计：本研究对受精卵至胚胎发育8.5天阶段的样本（包含分离的内细胞团（Inner Cell Mass, ICM）、滋养外胚层（Trophectoderm, TE）、上胚层（Epiblast, Epi）以及胚外外胚层（Extraembryonic Ectoderm, Exe））进行了H3K9me3修饰的染色质免疫共沉淀测序（Chromatin Immunoprecipitation sequencing, ChIP-seq）分析。此外，本研究还分析了小鼠第二次减数分裂中期（metaphase II, MII）卵母细胞、精子，以及小鼠胚胎干细胞（mouse embryonic stem cells, mESCs）和小鼠滋养层干细胞（mouse trophoblast stem cells, mTSCs）样本。在着床前胚胎的ChIP-seq分析中，每个反应使用500个细胞，每个发育阶段设置2至3次生物学重复。本数据集同时包含了受精卵至胚胎发育6.5天阶段的胚胎（包含桑椹胚期及小干扰RNA（siRNA）干扰组样本，且分离得到ICM、TE、Epi与Exe）的RNA测序（RNA-seq）数据。