Analysis of Imprinted Genes Expressed in the Brain.

父系表达基因3(Peg3)和5(Peg5)是我们用一种新的消减杂交方法系统筛选印记基因而鉴定的，它们在成人脑中特异表达。PEG3编码一个大的C2H2型锌指蛋白，可能是一种新的DNA结合蛋白。PEG5被鉴定为Neuronatin，据报道，Neuronatin最早在8.5天的菱形核3和5中表达，后来在中枢神经系统中表达。应用原位杂交技术，结合特异性抗体免疫染色，证明Peg3蛋白定位于成年大鼠脑内神经元和神经胶质细胞的胞核。我们还证明了人PEG3基因通过将完整的PEG3基因导入胶质瘤细胞而抑制了胶质瘤细胞的致瘤性。这些结果表明，Peg3蛋白作为一种DNA结合蛋白，在细胞周期中控制着几个基因的功能，从而在神经胶质细胞中发挥肿瘤抑制作用。对我们分离的印迹基因进行原位杂交的系统分析表明，在早期胚胎中的表达模式分为中胚层特异表达、神经组织特异表达和极低表达。然而，到目前为止分析的所有印记基因都显示在包括卵黄囊在内的胎盘组织中高表达。由于在胎盘组织中的高表达与被质疑基因的印记状态之间存在高度的相关性，我们提出了印记基因在胎盘中表达受调控的新假说，表明基因组印记在哺乳动物胎盘组织的形成中具有生物学意义。

Paternally expressed genes 3 (Peg3) and 5 (Peg5), which were identified by our systematic screening of imprinted genes using a novel subtraction-hybridization method, are expressed in adult brain specifically. Peg3 encodes a large C2H2 type zinc finger protein, presumably functioning a novel DNA binding protein. Peg5 was idenfied with Neuronatin which was reported to be expressed firstly in rhombomere 3 and 5 at day 8.5 embryos and later in central nervus system. Using in situ hybridization technique, immuno-staining with specific antibodies, we demonstrated that Peg3 protein localized in nucleus of neuronal and glial cells in the adult brain. We also demonstrated that human PEG3 gene suppressed the tumorigenecity of glioma cells by introducing a complete PEG3 cDNA into the cells. These results suggested that PEG3 protein fuctions as a DNA binding protein and controls several genes functioning in cell cycles, and as a result acts as tumor suppressor in glial cells.Systematic analysis of in situ hybridization of the imprinted genes we have isolated showed that the expression pattems in early embryos were divided to three groups, mesoderm specific, neural tissue specific and very low expression in the embryos. However, all imprinted genes so far analyzed showed high expression in the placental tissues including yolk sac. Because there existed a high correlation between high expression in the placental tissues and the imprinted status of the gene questioned, we proposed a novel hypothesis that imprinted genes are regulated to be expressed in the placenta, suggesting the biological significance of the genomic imprinting existed in the formation of placental tissues in mammals.