Sydney Medical School, Centre for Developmental and Regenerative Medicine, Kolling Institute for Medical Research, University of Sydney, Sydney, Australia.
PLoS One. 2012;7(1):e30687. doi: 10.1371/journal.pone.0030687. Epub 2012 Jan 26.
Normal development of the mammalian embryo requires epigenetic reprogramming of the genome. The level of cytosine methylation of CpG-rich (5meC) regions of the genome is a major epigenetic regulator and active global demethylation of 5meC throughout the genome is reported to occur within the first cell-cycle following fertilization. An enzyme or mechanism capable of catalysing such rapid global demethylation has not been identified. The mouse is a widely used model for studying developmental epigenetics. We have reassessed the evidence for this phenomenon of genome-wide demethylation following fertilisation in the mouse. We found when using conventional methods of immunolocalization that 5meC showed a progressive acid-resistant antigenic masking during zygotic maturation which gave the appearance of demethylation. Changing the unmasking strategy by also performing tryptic digestion revealed a persistence of a methylated state. Analysis of methyl binding domain 1 protein (MBD1) binding confirmed that the genome remained methylated following fertilisation. The maintenance of this methylated state over the first several cell-cycles required the actions of DNA methyltransferase activity. The study shows that any 5meC remodelling that occurs during early development is not explained by a global active loss of 5meC staining during the cleavage stage of development and global loss of methylation following fertilization is not a major component of epigenetic reprogramming in the mouse zygote.
哺乳动物胚胎的正常发育需要对基因组进行表观遗传重编程。基因组中富含胞嘧啶的 CpG 区域（5meC）的胞嘧啶甲基化水平是主要的表观遗传调节剂，据报道，受精后第一个细胞周期内，基因组内的 5meC 会发生广泛的去甲基化。目前尚未鉴定出能够催化这种快速全基因组去甲基化的酶或机制。小鼠是研究发育表观遗传学的常用模型。我们重新评估了受精后小鼠基因组全范围去甲基化这一现象的证据。我们发现，当使用免疫定位的常规方法时，5meC 在胚胎成熟过程中表现出逐渐的酸抗性抗原性屏蔽，这给人一种去甲基化的表观。通过改变去屏蔽策略，同时进行胰蛋白酶消化，显示出甲基化状态的持续存在。甲基结合域 1 蛋白（MBD1）结合的分析证实，受精后基因组仍保持甲基化状态。在第一个几个细胞周期中维持这种甲基化状态需要 DNA 甲基转移酶活性的作用。该研究表明，在早期发育过程中发生的任何 5meC 重塑都不能用发育分裂阶段 5meC 染色的广泛主动丢失来解释，并且受精后甲基化的广泛丢失不是小鼠受精卵中表观遗传重编程的主要组成部分。
