The dynamic reprogramming of H3K9me3 at hominoid-specific retrotransposons during early human development. The dynamic reprogramming of H3K9me3 at hominoid-specific retrotransposons during early human development

Reprogramming of H3K9me3-dependent heterochromatin is required for early development. How H3K9me3 is involved in early human development is, however, largely unclear. Here, we resolve the temporal landscape of H3K9me3 during human preimplantation development and its regulation for diverse hominoid-specific retrotransposons. At the 8-cell stage, H3K9me3 reprogramming at hominoid-specific retrotransposons termed SINE-VNTR-Alu (SVA) facilitates interaction between certain promoters and SVA-derived enhancers, facilitating the zygotic genome activation. In trophectoderm, de novo H3K9me3 domains prohibit pluripotent transcription factors from binding on hominoid-specific retrotransposons-derived regulatory elements for inner cell mass (ICM)-specific genes. H3K9me3 re-establishment at SVA elements in ICM is associated with higher transcription of DNA damage repair genes, compared to naïve human pluripotent stem cells. Our data demonstrate that species-specific reorganization of H3K9me3-dependent heterochromatin at hominoid-specific retrotransposons plays important roles during early human development, shedding light on how the epigenetic regulatory network for early development has evolved in mammals. Overall design: We investigated 1) H3K9me3 landscape on the genome of human preimplantation embryos using advanced ultra-low input ChIP-seq (AULIChIP-seq). AULIChIP-seq was performed in two biological replicates of human 4-cell embryo, 8-cel embryo, morula, inner cell mass (ICM) and trophectoderm (TE). 2) H3K9me3 or H3K27me3 landscape on the genome of hESCs and hTSCs using advanced ultra-low input ChIP-seq (AULIChIP-seq) or conventional ChIP-seq. AULIChIP-seq was performed in two biological replicates of hTSCs. 3) transcriptome of abnormal human embryos using SMART-seq2. 4) We investigated H3K9me3 landscape and gene expression of hESCs, prEpiSC and human preimplantation embryos in which the H3K9me3 modification were deposited on SVAs by epigenetic editing system (dCas9-KRAB). 5) genome of abnormal human embryos using WGS.

依赖组蛋白H3赖氨酸9三甲基化（H3K9me3）的异染色质重编程是早期发育不可或缺的过程。然而，H3K9me3如何参与人类早期发育，目前在很大程度上仍不明确。本研究解析了人类植入前发育过程中H3K9me3的动态修饰图谱，及其对多种类人猿特异性逆转录转座子的调控作用。在8细胞阶段，于被称为短散在核元件-可变数目串联重复序列-Alu（SINE-VNTR-Alu, SVA）的类人猿特异性逆转录转座子处发生的H3K9me3重编程，可促进特定启动子与SVA衍生增强子之间的相互作用，进而推动合子基因组激活。在滋养外胚层中，新发H3K9me3结构域可阻止多能转录因子结合至类人猿特异性逆转录转座子衍生的、调控内细胞团（ICM）特异性基因的调控元件处。与初始态人类多能干细胞相比，内细胞团中SVA元件处的H3K9me3重建与DNA损伤修复基因的更高转录水平相关。本研究数据表明，在类人猿特异性逆转录转座子处发生的、依赖H3K9me3的异染色质物种特异性重塑，在人类早期发育中发挥关键作用，为解析哺乳动物早期发育表观遗传调控网络的演化历程提供了新视角。

实验设计总览：
1. 利用先进超低投入量染色质免疫共沉淀测序（AULIChIP-seq），解析人类植入前胚胎基因组上的H3K9me3修饰图谱。分别对4细胞期、8细胞期、桑椹胚、内细胞团（ICM）以及滋养外胚层的人类胚胎开展两次生物学重复的AULIChIP-seq检测。
2. 利用先进超低投入量染色质免疫共沉淀测序（AULIChIP-seq）或常规染色质免疫共沉淀测序（conventional ChIP-seq），解析人类胚胎干细胞（hESCs）与人类滋养层干细胞（hTSCs）基因组上的H3K9me3或H3K27me3修饰图谱。其中对hTSCs完成两次生物学重复的AULIChIP-seq检测。
3. 采用SMART-seq2技术检测异常人类胚胎的转录组。
4. 针对人类胚胎干细胞、prEpiSC以及人类植入前胚胎的H3K9me3修饰情况展开研究：通过表观遗传编辑系统（dCas9-KRAB）将H3K9me3修饰沉积于SVA元件处，并检测其H3K9me3修饰图谱与基因表达水平。
5. 利用全基因组测序（WGS）分析异常人类胚胎的基因组。