Mechanism for Developmental Regulation by Genomic Imprinting in the Female Germ Line.

在哺乳动物中，配子发生过程中基因组的表观遗传修饰导致亲本等位基因之间的印迹基因表达不相等，这是足月发育所必需的。为了进一步了解基因组印记，我们做了几组实验。首先，为了研究母体原代印记是何时以及如何准确建立的，我们制作了单性生殖胚胎，其中一个基因组来自1至20日龄小鼠的未生长或生长阶段的卵母细胞，另一个基因组来自成年小鼠的完全发育的卵母细胞。利用这些胚胎，进行了8个印迹基因Snrpn、Znfl27、Ndn、Peg3、Igf2r、p57^<KIP2>、Peg1和Impact的表达分析。结果表明，每个印迹基因的表观遗传标记并不是在卵母细胞生长的特定时间内全部施加，而是在卵母细胞初生卵泡期到窦卵泡期期间广泛发生。随着核供体卵母细胞的生长，所构建的孤雌生殖的发育能力逐渐降低。因此，本研究清楚地表明，在卵母细胞生长阶段，母体初级印记以一种循序渐进的方式发生在印迹基因组中。其次，我们研究了调节H19单等位基因的表达是否能提高孤雌胚胎的存活率。结果清楚地表明，携带被H19转录单元删除的ng-卵母细胞基因组的ng/fg孤雌胚胎成功发育为活胎，妊娠期为17.5天。对照实验表明，这种独特的现象与遗传背景的影响无关。组织学分析显示，ng^<H19-KO>/fg^<WT>孤雌子胎盘萎缩，伴有严重的黑细胞增生等异常。目前的结果表明，从ng等位基因停止H19基因的表达可导致胎盘发育延长，胎盘的功能缺陷可能对个体发生致命。少

In mammals, epigenetic modification of genomes during gametogenesis leads to an unequivalent expression of imprinted genes between parental alleles and necessary for term development. To understand the further insight into genomic imprinting we have done several lines of experiment. Firstly, to investigate when and how, precisely, maternal primary imprinting is established, we produced parthenogenetic embryos containing one genome from a non-growing or growth stage oocyte from 1- to 20-day old mice, and one from a fully-grown oocyte from adult mice. Using these embryos, expression analysis of eight imprinted genes, Snrpn, Znfl27, Ndn, Peg3, Igf2r, p57^<KIP2>, Peg1 and Impact, was conducted. The results show that the epigenetic markers for each imprinted gene are not imposed all together at a specific time during oocyte growth, but rather occurs within a wide range during the period from primary to antral follicle stage oocytes. The developmental ability of the constructed parthenogenet … More ic embryos is gradually reduced with the growth of nuclear donor oocytes. Thus, the present study clearly demonstrate that maternal primary imprinting occurs in a stepwise fashion for groups of imprinted genes during the oocyte growth stages. Secondly, we examined whether the regulation of H19 monoallelic expression enhances the survival of parthenogenetic embryos. The results clearly show that the ng/fg parthenogenetic embryos carrying the ng-oocyte genome that had been deleted by the H19 transcription unit successfully developed as live fetuses for 17.5 gestation days. Control experiments revealed that this unique phenomenon occurs irrespective of the genetic background effect. Histological analysis showed that the placenta of ng^<H19-KO>/fg^<WT> parthenotes was resulted in atrophia with sever negro-cytosis and other anomalies. The present results suggest that the cessation of H19 gene expression from the ng-allele causes the extended development and that functional defection of the placenta could be fatal for the ontogeny. Less