Methylome Dynamics of Bovine Gametes and in vivo Early Embryos.

Jingyue Ellie Duan,Jingbo Chen,Xinbao Zheng,Ion Mandoiu,Sadie L Marjani,Xiuchun Cindy Tian,Fahad Alqahtani,Zongliang Carl Jiang,Hong Dong

doi:10.3389/fgene.2019.00512

Abstract

DNA methylation undergoes drastic fluctuation during early mammalian embryogenesis. The dynamics of global DNA methylation in bovine embryos, however, have mostly been studied by immunostaining. We adopted the whole genome bisulfite sequencing (WGBS) method to characterize stage-specific genome-wide DNA methylation in bovine sperm, immature oocytes, oocytes matured in vivo and in vitro, as well as in vivo developed single embryos at the 2-, 4-, 8-, and 16-cell stages. We found that the major wave of genome-wide DNA demethylation was complete by the 8-cell stage when de novo methylation became prominent. Sperm and oocytes were differentially methylated in numerous regions (DMRs), which were primarily intergenic, suggesting that these non-coding regions may play important roles in gamete specification. DMRs were also identified between in vivo and in vitro matured oocytes, suggesting environmental effects on epigenetic modifications. In addition, virtually no (less than 1.5%) DNA methylation was found in mitochondrial DNA. Finally, by using RNA-seq data generated from embryos at the same developmental stages, we revealed a weak inverse correlation between gene expression and promoter methylation. This comprehensive analysis provides insight into the critical features of the bovine embryo methylome, and serves as an important reference for embryos produced in vitro, such as by in vitro fertilization and cloning. Lastly, these data can also provide a model for the epigenetic dynamics in human early embryos.

Highlights

Cytosine methylation plays essential roles in mammalian development, including gene expression, transposon silencing, cell differentiation, genomic imprinting, and X chromosome inactivation (Hackett and Surani, 2013)
Using whole genome bisulfite sequencing (WGBS), we analyzed a total of 35 samples of sperm, germinal vesicle (GV) oocytes, in vivo and in vitro matured oocytes and cleavage stage in vivo embryos
Our results showed that non-CpG methylation peaked when high expression of pluripotency genes and embryonic genome activation (EGA) occurred in bovine pre-implantation embryos (Jiang et al, 2014), indicating an active regulatory role of non-CpG methylation in pluripotent gene expression as speculated earlier (Ramsahoye et al, 2000)

Summary

INTRODUCTION

Cytosine methylation plays essential roles in mammalian development, including gene expression, transposon silencing, cell differentiation, genomic imprinting, and X chromosome inactivation (Hackett and Surani, 2013). In the mouse, reduced representation bisulfite sequencing (RRBS) revealed rapid genome-wide demethylation in zygotes (Smith et al, 2012). In primates, this major demethylation event did not occur until the 2-cell stage (Guo et al, 2014; Gao et al, 2017; Zhu et al, 2018). These data will provide the gold standard reference that can lead to improvements in assisted reproductive technologies and provide evolutionary insights across species. Bovine embryos, which are more similar to human embryos than mouse embryos are, in terms of gene expression profiles and developmental timing, can serve as a great model for understanding human development, especially since human in vivo embryos are not available for research

MATERIALS AND METHODS

RESULTS AND DISCUSSION

ETHICS STATEMENT