Nucleosome organization of mouse embryos during pre-implantation development

Abstract

Sexual reproduction begins with sperm-oocyte fusion to form a zygote, where chromatin undergoes dramatic reorganization to establish totipotency. Although nucleosomes- the basic units of eukaryotic chromatin and key epigenetic regulators- are extensively remodeled during early embryogenesis, their dynamic repositioning mechanisms and biological implications remain unclear. Here, we employed single-cell MNase sequencing (scMNase-seq) to map genome-wide nucleosome positioning and chromatin accessibility in individual mammalian embryos. We found that nucleosome positioning mirrored somatic cell patterns until the 4-cell stage, with nucleosome depletion and phasing at CTCF sites not fully established until morula formation. By integrating H3K4me3 localization and transcriptomic data, we revealed that nucleosome sparsity at transcription start sites (TSS) and flanking regions correlated with expression levels of genes critical for preimplantation development. Notably, these nucleosome-depleted regions likely serve as regulatory hubs influencing histone modification dynamics. Our study systematically delineates nucleosome reorganization principles during mammalian embryogenesis and provides a high-resolution resource for understanding chromatin remodeling in early development.

Keywords: Chromatin organization; Embryos; H3K4me3; Nucleosome position; scMNase-seq.

Fig. 1

Preimplantation embryos scMNase-seq library construction. (A) Schematic diagram of preimplantation embryo acquisition and scMNase-seq library construction. (B) Fragment-length density of scMNase-seq data in preimplantation embryos.

Fig. 2

Profiles of nucleosome and subnucleosome-sized particles around the TSS of all genes in preimplantation embryos.

Fig. 3

Profiles of nucleosome and subnucleosome-sized particles profiles around the stage specifc CTCF binding sites in preimplantation embryos. ChipSeq sequencing data (PRJCA018124) of preimplantation embryos at each stage were used to obtain the midpoint information of stage-specific CTCF binding sites. Plot2DO was used to generate V-plots to determine the distribution of scMNase-seq fragments at different developmental stages before implantation. The arrangement of nucleosomes and subnucleosome-sized particles is highlighted, with red lines representing nucleosomes and blue subnucleosome-sized particles illustrating their respective arrangement near the midpoint of the phase-specific CTCF binding site.

Fig. 4

Profiles of nucleosome and subnucleosome-sized particles around the common CTCF binding sites of ESC in preimplantation embryos. Heat map of correlation of ChipSeq sequencing results of ESCs CTCF from GSM3771259, ENCSR000CCB, GSM2533853, GSM691014, GSM918748. Venn diagram of CTCF peaks in GSM3771259, ENCSR000CCB, GSM2533853, GSM691014 and GSM918748 to obtain shared peaks. Plot2DO was used to generate V-plots to clarify the distribution of scMNase-seq fragments around the center of these common CTCF binding sites at different stages of preimplantation embryo development. The arrangement of nucleosomes and subnucleosome-sized particles is highlighted, with red lines representing nucleosomes and blue subnucleosome-sized particles, illustrating their respective arrangement near the midpoint of the CTCF binding site.

Fig. 5

Nucleosome profiles around H3K4me3/H3K27me3 marked genes. (A) Nucleosome profiles around H3K4me3 marked genes in preimplantation embryos. (B) Nucleosome profiles around H3K27me3 marked genes in preimplantation embryos. (C) Nucleosome profiles around bivalent marked genes in preimplantation embryos. (D) Nucleosome profiles around nonmarked genes in preimplantation embryos.

Fig. 6

Nucleosome profiles around genes with different length of the H3K4me3 domains. H3K4me3 ULI-NChIP–seq datasets of pre-implantation embryos (GSE73952) were employed to obtain the gene information of the control, normal, narrow, broad H3K4me3 peaks, The arrangement of nucleosome fragments within the 300 bp range upstream and downstream of these genes in the corresponding stage of pre-implantation embryos was shown.

Fig. 7

Nucleosome profiles around genes with different FPKM values and H3K4me3 modification. The H3K4me3 ULI-NChIP–seq datasets of pre-implantation embryos (GSE73952) were utilized to obtain the gene information of the H3K4me3 peaks, and the RNA-seq datasets of pre-implantation embryos (GSE66582) were employed to obtain the gene expression levels of pre-implantation embryos. The genes at each stage were classified according to the FPKM values in the RNA-seq data, and the intersection with the genes marked by H3K4me3 was taken to draw the Venn diagram. The arrangement of nucleosome fragments within the 300 bp range upstream and downstream of genes with or without H3K4me3 modification at different expression levels was analyzed respectively. Gene body was normalized to a length of 300 bp. Blue/red text as results of GO analyses.