Davis T L, Tremblay K D, Bartolomei M S
Howard Hughes Medical Institute, University of Pennsylvania School of Medicine, Philadelphia 19104,
Dev Genet. 1998;23(2):111-8. doi: 10.1002/(SICI)1520-6408(1998)23:2<111::AID-DVG3>3.0.CO;2-9.
The imprinted H19 gene is hypomethylated on the active maternal allele and hypermethylated on the repressed paternal allele in the somatic tissues of mice and humans. We previously demonstrated that the paternal-specific methylation of a 2 kb region located between -2 and -4 kb relative to the start of transcription is maintained throughout murine development, and we thus propose that this region is crucial to determining the imprinted expression of H19. Here, we test the correlation between differential methylation and imprinted expression by analyzing the mouse H19 gene in the undermethylated extraembryonic tissues. During early and midpostimplantation stages, > 95% of the H19 RNA is derived from the maternal allele. Dissection of yolk sac revealed that the paternal allele is expressed at a low level in the viseral endoderm but is completely repressed in visceral mesoderm. Bisulfite methylation analysis of yolk sac DNA showed that the maternal allele was hypomethylated and that 95% of the paternally derived clones were hypermethylated. Thus in extraembryonic lineages, the majority of H19 DNA is differentially methylated. These results lend further support to the hypothesis that DNA methylation confers the imprint on H19.
在小鼠和人类的体细胞组织中，印记基因H19在活跃的母本等位基因上发生低甲基化，而在受抑制的父本等位基因上发生高甲基化。我们之前证明，相对于转录起始点，位于-2至-4 kb之间的2 kb区域的父本特异性甲基化在整个小鼠发育过程中得以维持，因此我们提出该区域对于确定H19的印记表达至关重要。在此，我们通过分析低甲基化的胚外组织中的小鼠H19基因，来检测差异甲基化与印记表达之间的相关性。在植入后早期和中期阶段，超过95%的H19 RNA来源于母本等位基因。对卵黄囊的解剖显示，父本等位基因在内胚层中低水平表达，但在内脏中胚层中完全受抑制。对卵黄囊DNA进行亚硫酸氢盐甲基化分析表明，母本等位基因发生低甲基化，且95%的父本来源克隆发生高甲基化。因此，在胚外谱系中，大多数H19 DNA存在差异甲基化。这些结果进一步支持了DNA甲基化赋予H19印记这一假说。
