Department of Nutritional Science and Food Management, Ewha Womans University, Seoul, Republic of Korea.
PLoS One. 2012;7(3):e33024. doi: 10.1371/journal.pone.0033024. Epub 2012 Mar 5.
Epigenetic marks are fundamental to normal development, but little is known about signals that dictate their placement. Insights have been provided by studies of imprinted loci in mammals, where monoallelic expression is epigenetically controlled. Imprinted expression is regulated by DNA methylation programmed during gametogenesis in a sex-specific manner and maintained after fertilization. At Rasgrf1 in mouse, paternal-specific DNA methylation on a differential methylation domain (DMD) requires downstream tandem repeats. The DMD and repeats constitute a binary switch regulating paternal-specific expression. Here, we define sequences sufficient for imprinted methylation using two transgenic mouse lines: One carries the entire Rasgrf1 cluster (RC); the second carries only the DMD and repeats (DR) from Rasgrf1. The RC transgene recapitulated all aspects of imprinting seen at the endogenous locus. DR underwent proper DNA methylation establishment in sperm and erasure in oocytes, indicating the DMD and repeats are sufficient to program imprinted DNA methylation in germlines. Both transgenes produce a DMD-spanning pit-RNA, previously shown to be necessary for imprinted DNA methylation at the endogenous locus. We show that when pit-RNA expression is controlled by the repeats, it regulates DNA methylation in cis only and not in trans. Interestingly, pedigree history dictated whether established DR methylation patterns were maintained after fertilization. When DR was paternally transmitted followed by maternal transmission, the unmethylated state that was properly established in the female germlines could not be maintained. This provides a model for transgenerational epigenetic inheritance in mice.
表观遗传标记对于正常发育至关重要，但对于决定其位置的信号知之甚少。通过对哺乳动物印迹基因座的研究提供了一些见解，在这些基因座中，单等位基因表达是通过表观遗传控制的。印迹表达受配子发生过程中以性别特异性方式编程的 DNA 甲基化调控，并在受精后得到维持。在小鼠的 Rasgrf1 中，父本特异性 DNA 甲基化在差异甲基化域 (DMD) 上需要下游串联重复序列。DMD 和重复序列构成了调节父本特异性表达的二进制开关。在这里，我们使用两种转基因小鼠品系定义了足以进行印迹甲基化的序列：一种携带整个 Rasgrf1 簇 (RC)；另一种仅携带 Rasgrf1 的 DMD 和重复序列 (DR)。RC 转基因品系重现了内源性基因座上所见印迹的所有方面。DR 在精子中经历了适当的 DNA 甲基化建立，在卵母细胞中被抹去，表明 DMD 和重复序列足以在生殖系中编程印迹 DNA 甲基化。这两种转基因都产生了一个跨越 DMD 的 pit-RNA，先前已证明该 RNA 对于内源性基因座上的印迹 DNA 甲基化是必需的。我们表明，当 pit-RNA 表达受重复序列控制时，它仅在顺式而非反式调节 DNA 甲基化。有趣的是，系谱历史决定了受精后建立的 DR 甲基化模式是否得到维持。当 DR 父本传递后母本传递时，在雌性生殖系中正确建立的非甲基化状态无法维持。这为小鼠中转代表观遗传遗传提供了一个模型。
