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De novo DNA methylation during monkey pre-implantation embryogenesis

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ABSTRACT

Critical epigenetic regulation of primate embryogenesis entails DNA methylome changes. Here we report genome-wide composition, patterning, and stage-specific dynamics of DNA methylation in pre-implantation rhesus monkey embryos as well as male and female gametes studied using an optimized tagmentation-based whole-genome bisulfite sequencing method. We show that upon fertilization, both paternal and maternal genomes undergo active DNA demethylation, and genome-wide de novo DNA methylation is also initiated in the same period. By the 8-cell stage, remethylation becomes more pronounced than demethylation, resulting in an increase in global DNA methylation. Promoters of genes associated with oxidative phosphorylation are preferentially remethylated at the 8-cell stage, suggesting that this mode of energy metabolism may not be favored. Unlike in rodents, X chromosome inactivation is not observed during monkey pre-implantation development. Our study provides the first comprehensive illustration of the 'wax and wane' phases of DNA methylation dynamics. Most importantly, our DNA methyltransferase loss-of-function analysis indicates that DNA methylation influences early monkey embryogenesis.

No MeSH data available.


dnmt3a and dnmt3b knockdown improves developmental potential of the embryos. (A) Morpholinos can significantly knock down endogenous dnmt3a and dnmt3b expression in monkey embryos (n = 3, * represents P ≤ 0.05). (B) Bar graph showing percentages of embryos that reach the blastocyst stage after injection of dnmt3a and dnmt3b morpholinos or negative control morpholino (Three independent experiments. Totally 15 embryos were used in control and knockdown experiments, respectively. Error bars represent SD. * represents P ≤ 0.05).
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fig6: dnmt3a and dnmt3b knockdown improves developmental potential of the embryos. (A) Morpholinos can significantly knock down endogenous dnmt3a and dnmt3b expression in monkey embryos (n = 3, * represents P ≤ 0.05). (B) Bar graph showing percentages of embryos that reach the blastocyst stage after injection of dnmt3a and dnmt3b morpholinos or negative control morpholino (Three independent experiments. Totally 15 embryos were used in control and knockdown experiments, respectively. Error bars represent SD. * represents P ≤ 0.05).

Mentions: During in vitro fertilization (IVF) of rhesus monkey, developmental failure often occurs at the transition from the 8-cell stage to the morula stage. As accurate epigenetic reprogramming is essential in normal development, the failure may result from incomplete demethylation or remethylation caused by imperfect in vitro culture condition. On the basis of the observation we described above, we hypothesized that blockade of de novo DNA methylation during the pre-implantation period could influence embryonic development. To test this hypothesis we injected morpholinos against dnmt3a and dnmt3b at the zygote stage; and found that the number of embryos arrested at the transition from the 8-cell to the morula stage was indeed significantly reduced: 70% of embryos injected with dnmt3a and dnmt3b morpholinos developed into blastocysts, whereas only about 40% of the negative control embryos developed into the blastocyst stage, which is expected from IVF in monkey (Figure 6A and 6B).


De novo DNA methylation during monkey pre-implantation embryogenesis
dnmt3a and dnmt3b knockdown improves developmental potential of the embryos. (A) Morpholinos can significantly knock down endogenous dnmt3a and dnmt3b expression in monkey embryos (n = 3, * represents P ≤ 0.05). (B) Bar graph showing percentages of embryos that reach the blastocyst stage after injection of dnmt3a and dnmt3b morpholinos or negative control morpholino (Three independent experiments. Totally 15 embryos were used in control and knockdown experiments, respectively. Error bars represent SD. * represents P ≤ 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5385613&req=5

fig6: dnmt3a and dnmt3b knockdown improves developmental potential of the embryos. (A) Morpholinos can significantly knock down endogenous dnmt3a and dnmt3b expression in monkey embryos (n = 3, * represents P ≤ 0.05). (B) Bar graph showing percentages of embryos that reach the blastocyst stage after injection of dnmt3a and dnmt3b morpholinos or negative control morpholino (Three independent experiments. Totally 15 embryos were used in control and knockdown experiments, respectively. Error bars represent SD. * represents P ≤ 0.05).
Mentions: During in vitro fertilization (IVF) of rhesus monkey, developmental failure often occurs at the transition from the 8-cell stage to the morula stage. As accurate epigenetic reprogramming is essential in normal development, the failure may result from incomplete demethylation or remethylation caused by imperfect in vitro culture condition. On the basis of the observation we described above, we hypothesized that blockade of de novo DNA methylation during the pre-implantation period could influence embryonic development. To test this hypothesis we injected morpholinos against dnmt3a and dnmt3b at the zygote stage; and found that the number of embryos arrested at the transition from the 8-cell to the morula stage was indeed significantly reduced: 70% of embryos injected with dnmt3a and dnmt3b morpholinos developed into blastocysts, whereas only about 40% of the negative control embryos developed into the blastocyst stage, which is expected from IVF in monkey (Figure 6A and 6B).

View Article: PubMed Central - PubMed

ABSTRACT

Critical epigenetic regulation of primate embryogenesis entails DNA methylome changes. Here we report genome-wide composition, patterning, and stage-specific dynamics of DNA methylation in pre-implantation rhesus monkey embryos as well as male and female gametes studied using an optimized tagmentation-based whole-genome bisulfite sequencing method. We show that upon fertilization, both paternal and maternal genomes undergo active DNA demethylation, and genome-wide de novo DNA methylation is also initiated in the same period. By the 8-cell stage, remethylation becomes more pronounced than demethylation, resulting in an increase in global DNA methylation. Promoters of genes associated with oxidative phosphorylation are preferentially remethylated at the 8-cell stage, suggesting that this mode of energy metabolism may not be favored. Unlike in rodents, X chromosome inactivation is not observed during monkey pre-implantation development. Our study provides the first comprehensive illustration of the 'wax and wane' phases of DNA methylation dynamics. Most importantly, our DNA methyltransferase loss-of-function analysis indicates that DNA methylation influences early monkey embryogenesis.

No MeSH data available.