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Single-Cell Analysis Elucidates Epigenetic Changes in Early Human Development

NEW YORK (GenomeWeb) – By analyzing single cells, researchers in China have begun to put together a picture of the shifting epigenetic landscape of early human development.

Upon fertilization, the former sperm and egg undergo DNA methylation reprogramming and chromatin remodeling to move from being differentiated gametes to a totipotent cell.

Researchers led by Peking University's Jie Qiao used single-cell chromatin overall omic-scale landscape sequencing (scCOOL-seq) to study human preimplantation embryos' chromatin state, DNA methylation, CNVs, and more, all at the same time. As they reported today in Nature Cell Biology, the researchers found that genes with high variation in DNA methylation tended to be distinct from those with high variation in chromatin accessibility. They also observed a feedback mechanism between transcription and maintaining open chromatin.

"Our work paves the way for dissecting the complex, yet highly coordinated, epigenetic reprogramming during human preimplantation development," Qiao and her colleagues wrote.

They used scCOOL-seq to analyze human cells' epigenetic states during preimplantation development as well as in gametes and embryonic stem cells. Overall, they found the chromatin of oocytes to be more accessible than that of sperm, though oocytes had fewer nucleosome-depleted regions than sperm. After fertilization, chromatin accessibility decreased, hitting a low of accessibility at the eight-cell stage. After zygotic gene activation, global chromatin accessibility increased, reaching a high at the morula stage.

Based on SNPs they identified from the donors' own genomes, the researchers tracked the epigenetic reprogramming of each parental genome in the cells. While the paternal genome was initially more highly methylated, they found it to become quickly demethylated after fertilization — by the two-cell stage, the maternal genome was more highly methylated. The paternal genome chromatin also became more open than the maternal genome until the four-cell stage when their levels of openness became similar.

That differs from what has been seen in mice, the researchers noted. They also uncovered other differences between mouse and human early development — such as an overall difference in chromatin accessibility and differences in timing of chromatin openness — which suggested species-specific features.

Additionally, the researchers reported that oocytes exhibited the lowest variations in DNA methylation. However, upon fertilization, variations in DNA methylation emerged at exons and introns as well as at repeat elements and enhancers. But some spots like promoters, CpG islands, proximal nucleosome-depleted regions (NDRs), and RNA polymerase II-binding sites were only lowly methylated.

There appeared, the researchers said, to be two different sets of genes based on whether they exhibited high variations in DNA methylation or high variations in chromatin accessibility. Those with high chromatin accessibility variation were enriched for genes associated with chromosome organization, chromatin modification, and the cell cycle. At the same time, those with high DNA methylation variations were enriched for involvement in the inflammatory response and the detection of chemical stimulus.

The researchers also noticed that the number of wide proximal NDRs increased slightly after fertilization, going from 1.2 percent in mature metaphase II oocytes to 7.6 percent at the mid-zygotic stage, and then jumped to 21 percent at the eight-cell stage. As the wide NDRs had higher levels of GCH methylation and an enrichment for RNA II polymerase, the researchers suspected they could be associated with transcription.

When they then blocked RNA polymerase II-mediated transcription in zygotes, they found that most of the wide proximal NDRs — especially near genes with key roles in embryogenesis — could not be maintained. This suggested to the researchers that continual transcription is necessary for a number of zygotic genes to maintain their promoters' openness as a feedback mechanism.

"Our study provides insights toward a deeper understanding of epigenetic reprogramming during human preimplantation development," the authors wrote.