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High-throughput functional characterization of enhancers in totipotent-like cells

Yang, L.; Peng, T.; Zhu, Y.; Zhai, X.; Huang, B.; Li, L.; Zhang, T.; Chen, J.; Liang, D.; He, J.; Zhang, M.

2025-11-04 developmental biology
10.1101/2025.11.03.686172 bioRxiv
Show abstract

Zygotic genome activation (ZGA) marks the initial transcription event in embryogenesis, yet the cis-regulatory mechanism remains unclear. Here, utilizing massively parallel reporter assays, we functionally dissect enhancers across mouse genome in DUX-induced 2C-like cells (2CLCs). Integrated analysis with epigenomic and transcriptomic data from 2CLCs and 2-cell (2C) embryos, active enhancers in totipotent cells are depicted. Among them, a notable proportion of promoters exhibit enhancer activities, showing elevated active chromatin features and correlating with enhanced gene expression during ZGA. Furthermore, only half of the MT2_Mm exhibit enhancer activities in 2CLCs. In addition, 2CLC enhancers augment transcription in 2C embryos. Finally, deleting enhancer regions in both 2CLCs and 2C embryos highlighted their crucial role in facilitating transcription of ZGA genes. These findings advance our comprehension of the cis-regulatory mechanism governing ZGA process.

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