Functional and Imprinting Analysis in Brain of Angelman Syndrome Gene UBE3A

人类UBE3A基因显示出大脑特异性的部分印记，缺乏母体遗传的等位基因会导致Angelman综合征（AS），其特征是神经行为异常。在几种AS模型小鼠中，印迹Ube3a主要在海马、小脑浦肯野细胞和嗅球中表达。因此，小鼠Ube3a的印记被认为是区域特异性的，在不同的大脑区域中，父本Ube3a等位基因的沉默水平不同。为了确定Ube3a印迹表达的细胞类型，我们利用原代皮质细胞培养分析了其在胚胎脑细胞中的印迹状态。RT - PCR和免疫荧光检测该基因的等位基因表达。Ube3a基因在大脑中编码两种RNA转录本：正义转录本和反义转录本。义转录本母系在神经元中表达，但在胚胎脑的神经胶质细胞中双等位表达，而反义转录本仅在神经元中表达，并在父系中表达。我们的数据提供了Ube3a在原代脑细胞培养中脑细胞类型特异性印迹的第一个证据，即神经元特异性印迹，而不是胶质细胞特异性印迹。仅在神经元中存在的正义转录物和反义转录物的相互印迹表明，神经元特异性印迹机制与神经干细胞的谱系决定有关。

The human UBE3A gene shows brain-specific partial imprinting, and lack of a maternally inherited allele causes Angelman syndrome (AS), which is characterized by neurobehavioral anomalies. In several AS model mice, imprinted Ube3a expression is detected predominantly in the hippocampus, cerebellar Purkinje cells, and the olfactory bulb. Therefore, imprinting of mouse Ube3a is thought to be region-specific with different levels of silencing of the paternal Ube3a allele in different brain regions. To determine cell-types of imprinted Ube3a expression, we analyzed its imprinting status in embryonic brain cells by using primary cortical cell cultures. RT PCR and immunofluorescence were performed to determine the allelic expression of the gene The Ube3a gene encodes two RNA transcripts in the brain : sense and antisense transcripts. The sense transcript was expressed maternally in neurons but biallelically expressed in glial cells in the embryonic brain, whereas the antisense transcript was expressed only in neurons and paternally expressed. Our data present the first evidence of brain cell-type specific imprinting, i.e., neuron-specific imprinting but not in glial cells, of Ube3a in primary brain cell cultures. Reciprocal imprinting of sense and antisense transcripts presented only in neurons suggests the neuron-specific imprinting mechanism related to the lineage determination of neural stem cells.