Establishing Chromatin Regulatory Landscape during Mouse Preimplantation Development

Upon fertilization, parental chromatin undergoes extensive reprogramming to generate the highly dynamic chromatin of the inner cell mass (ICM) cells from which an organism is derived. How the chromatin regulatory landscape is established during this process has remained a mystery due to the technical difficulties in chromatin analysis using limited cell numbers. Using a low-input DNase I sequencing (liDNase-seq) method, we generated DNase I-hypersensitive site (DHS) maps of mouse preimplantation embryos. We found that the DHSs are progressively established during development, with Oct4 contributing to a drastic gain of DHSs at the 8-cell stage. In addition, single nucleotide polymorphism analysis revealed that imprinted gene promoters harbor allele-specific DHSs before the onset of allelic gene expression. Furthermore, Nfya is required for 2-cell promoter DHS formation and zygotic genome activation. Thus, our study not only reveals chromatin regulatory landscapes, but also identifies key transcription factors important for DHS establishment in mammalian embryos. Overall design: In this study, we modified scDNase-seq method to achieve de novo genome-wide DHS mapping using as few as 30 cells. Using this low-input DNase-seq (liDNase-seq) method, we have generated DHS maps of mouse preimplantation embryos from 1-cell to the morula stage, revealing the dynamics of chromatin accessibility during the natural developmental process. We constructed a total of 32 liDNase-seq datasets, including: two 1-cell embryo libraries, two 2-cell embryo libraries, two 4-cell embryo libraries, two 8-cell embryo libraries, two morula embryo libraries, two mouse ESC libraries, two 14 day growing oocyte library, two sperm library, two maternal PN3 pronuclei library, two paternal PN3 pronuclei library, two maternal PN5 pronuclei library, two paternal PN5 pronuclei library, two siRNA control 2-cell embryo libraries, two Nfya KD (siNfya#1 and siNfya#2) 2-cell embryo libraries, two siRNA control 8-cell embryo libraries and two Oct4 KD 8-cell (siOct4#1 and siOct4#2) embryo libraries. Five RNA-seq datasets were generated including two siRNA control 2-cell embryo libraries, two Nfya KD (siNfya#1 and siNfya#2) 2-cell embryo libraries and one 1-cell embryo library.