Genome-wide promoter methylation patterns are altered by a food component, genistein, during embryonic stem cell differentiation

发育过程中的环境挑战影响胎儿表观基因组，但易受表观遗传失调影响的时期尚不清楚。通过使用大豆植物雌激素染料木黄酮（已知它可以改变胚胎发生期间Avy等位基因的表观遗传状态），我们探索了表观遗传调节的敏感期。植入后时期，当从头DNA甲基化活跃地进行时，适合于使用小鼠胚胎干（ES）细胞分化系统进行体外分析。小鼠ES细胞在存在或不存在染料木黄酮的情况下分化，并通过基于微阵列的启动子甲基化分析结合甲基化敏感性核酸内切酶（HpaII/McrBC）依赖性富集程序比较第10天的DNA甲基化模式。在染料木黄酮处理后的启动子亚组中观察到甲基化水平的中度变化。对Ucp 1和Sytl 1启动子的详细研究进一步揭示，染料木黄酮不影响在第0天和第4天之间发生的从头甲基化，但干扰随后的调控过程，并导致两个启动子的甲基化水平降低。染料木黄酮干扰分化ES细胞从头甲基化后的甲基化模式。我们的观察结果表明，对于一个子集的基因，调控后从头DNA甲基化在早期胚胎可能是敏感的染料木黄酮。

Environmental challenges during development affect the fetal epigenome, but the period(s) vulnerable to epigenetic dysregulation are not clear. By employing a soy phytoestrogen, genistein, that is known to alter the epigenetic states of the Avy allele during embryogenesis, we have explored the sensitive period for epigenetic regulation. The post-implantation period, when de novo DNA methylation actively proceeds, is amenable to in vitro analysis using a mouse embryonic stem(ES) cell differentiation system. Mouse ES cells were differentiated in the presence or absence of genistein, and DNA methylation patterns on day 10 were compared by microarray-based promoter methylation analysis coupled with a methylation-sensitive endonuclease(HpaII/McrBC)-dependent enrichment procedure. Moderate changes in methylation levels were observed in a subset of promoters following genistein treatment. Detailed investigation of the Ucp1 and Sytl1 promoters further revealed that genistein does not affect de novo methylation occurring between day 0 and day 4, but interferes with subsequent regulatory processes and leads to a decrease in methylation level for both promoters. Genistein perturbed the methylation pattern of differentiated ES cells after de novo methylation. Our observations suggest that, for a subset of genes, regulation after de novo DNA methylation in the early embryo may be sensitive to genistein.