Dnmt1s in donor cells is a barrier to SCNT-mediated DNA methylation reprogramming in pigs.
Dnmt1s in donor cells is a barrier to SCNT-mediated DNA methylation reprogramming in pigs.
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供体细胞中的 Dnmt1s 是猪 SCNT 介导的 DNA 甲基化重编程的障碍
DOI：
10.18632/oncotarget.16507
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发表时间：
2017-05-23
期刊：
影响因子：
--
通讯作者：
Huan Y
中科院分区：
文献类型：
--
作者：
Song X;Liu Z;He H;Wang J;Li H;Li J;Li F;Jiang Z;Huan Y
体细胞核移植胚胎发育缓慢可能是由于DNA甲基化重编程不完全所致，供体细胞中存在的Dnmt1可能是DNA甲基化重编程中断的原因之一。然而，Dnmt1s的重编程模式及其对克隆胚胎DNA甲基化重编程的影响仍然知之甚少。在此，我们发现，随着Dnmt1在合子基因激活阶段的表达显著增加，基因组甲基化水平显著上调，与体外受精相比，基因组甲基化停滞率显著升高。然后，我们证明在克隆胚胎中，从1-细胞到4-细胞阶段，Dnmt1s的甲基化水平和表达水平显著高于Dnmt1s，而不是Dnmt1o，而在囊胚期显著低于Dnmt1s。当供体细胞中的Dnmt1s被适当去除时，更多的克隆胚胎进入合子基因激活期，囊胚率显著提高。此外，Dnmt1s基因敲除显著改善了自身和克隆胚胎基因组甲基化重建。最后，我们发现Dnmt1s的去除显著促进了克隆胚胎中多能基因的去甲基化和表达。综上所述，这些数据表明，供体细胞中的Dnmt1是体细胞核移植介导的DNA甲基化重编程的关键障碍，损害了克隆胚胎的发育。
Low development of somatic cell nuclear transfer embryos could be due to the incomplete DNA methylation reprogramming, and Dnmt1s existing in donor cells may be one cause of this disrupted DNA methylation reprogramming. However, the reprogramming pattern of Dnmt1s and its effect on DNA methylation reprogramming in cloned embryos remain poorly understood. Here, we displayed that along with the significantly higher Dnmt1 expression at the zygotic gene activation stage of cloned embryos, genomic methylation level was markedly upregulated, and the arrested rate was significantly higher compared with their in vitro fertilization counterparts. Then, we demonstrated that Dnmt1s, not Dnmt1o, methylation and expression levels in cloned embryos were significantly higher from the 1-cell to 4-cell stage but markedly lower at the blastocyst stage. When Dnmt1s in donor cells was appropriately removed, more cloned embryos passed through the zygotic gene activation stage and the blastocyst rate significantly increased. Furthermore, Dnmt1s knockdown significantly improved itself and genomic methylation reconstruction in cloned embryos. Finally, we found that Dnmt1s removal significantly promoted the demethylation and expression of pluripotent genes in cloned embryos. Taken together, these data suggest that Dnmt1s in donor cells is a critical barrier to somatic cell nuclear transfer mediated DNA methylation reprogramming, impairing the development of cloned embryos.