The Role of Genome-Coded ME2 in Epilepsy

描述（由申请人提供）：特发性全面性癫痫（IGE）是最常见的癫痫形式之一，约占所有癫痫的30%；而且，其病因被认为主要是遗传的。连锁和关联研究已经确定了IGE的几个候选基因，即苹果酸酶2 （ME2）和含溴结构域2 （BRD2）。此外，以往的研究也表明IGE易感性是基因-基因相互作用的结果。我们将重点建立ME2和IGE之间明确的生物学联系，我们也将测试ME2和BRD2之间的相互作用。18号染色体上的强连锁和关联结果表明，ME2是IGE的一般易感位点，并遵循隐性遗传模式。对20例IGE病例和18例对照组的序列分析显示，在ME2内含子1中，有一个区域的多态位点在病例中多态位点在对照组中多态。此外，ME2是IGE易感性的一个很好的生物学候选物，因为：1)其在脑神经细胞中的丰度；2)定位于线粒体；最引人注目的是，3)苹果酸脱羧的催化作用，为谷氨酸和GABA合成提供关键的丙酮酸。GABA是中枢神经系统中主要的抑制性神经递质，参与了癫痫样尖峰波活动的调节。抗癫痫药物机制也调节GABA神经递质系统，GABA受体的突变已被证明在某些形式的癫痫易感性中起作用。在这项拨款申请中，我们将根据我们选择的病例样本来确定ME2内含子1的不同单倍型，这些病例的序列数据是可用的。然后，我们将使用报告子结构来检查不同的内含子1单倍型对剪接和转录水平的孤立影响。注意，我们的报告子结构将只包含近端启动子元件和内含子1。接下来，我们将研究内含子1变异在神经元和/或胶质细胞中的作用；不像我们的记者构建，这些实验将涉及整个ME2基因。最后，使用标准的重新采样方法和一种新的统计方法，我们将测试ME2和BRD2在青少年肌阵挛性癫痫（IGE亚型）中的相互作用。总的来说，这项研究应该有助于阐明ME2作为IGE易感基因的一般作用，以及它在其他遗传因素网络中的特殊作用。公共卫生相关性：癫痫仅在美国就影响270万人。特发性全身性癫痫（IGE）占所有癫痫的30%，发病于儿童期或青春期，被认为具有遗传病因。我们的研究已经确定了一个基因（ME2）与几种不同的常见IGE综合征相关，包括青少年肌阵挛性癫痫（JME），青少年癫痫缺失（JAE）和癫痫伴全身性强直-阵挛性发作（EGTCS）。我们建议确定在ME2基因中发现的多态性如何增加对广泛的相关癫痫综合征的易感性。

DESCRIPTION (provided by applicant): Idiopathic Generalized Epilepsy (IGE) is one of the most common forms of epilepsy, representing about 30% of all epilepsies; and, its etiology is considered to be mostly genetic. Both linkage and association studies have identified several candidate genes for IGE, namely: malic enzyme 2 (ME2) and bromodomain containing 2 (BRD2). In addition, previous studies have also suggested that IGE susceptibility results from gene-gene interaction. We will focus on establishing a clear biological link between ME2 and IGE, and we will also test for an interaction between ME2 and BRD2. The strong linkage and association results on chromosome 18 suggest that ME2 is a general IGE susceptibility locus, and that it follows a recessive mode of inheritance. Sequence analysis in 20 IGE cases and 18 controls, revealed a region in intron 1 of ME2 where polymorphic sites in cases are mostly monomorphic in controls. Moreover, ME2 is an excellent biological candidate for susceptibility to IGE due to: 1) its abundance in brain neurons; 2) its localization to the mitochondria; and most compelling, 3) its catalytic function of malate decarboxylation, producing a critical supply of pyruvate for glutamate and GABA synthesis. GABA is the main inhibitory neurotransmitter in the central nervous system, and is involved in the modulation of epileptiform spike-wave activity. Anti-epilepsy drug mechanisms also modulate the GABA neurotransmitter system, and mutations in GABA receptors have been shown to play a role in susceptibility to some forms of epilepsy. In this grant application, we will determine the different haplotypes of ME2 intron 1 based on our selected sample of cases for whom sequence data is available. Then, we will use reporter constructs to examine the isolated effects of different intron 1 haplotypes on splicing and levels of transcription. Note that our reporter constructs will only contain proximal promoter elements and intron 1. Next, we will examine the effects of intron 1 variants in neuronal and/or glial cells; and unlike our reporter constructs, these experiments will involve the entire ME2 gene. Finally, using standard resampling methods and a novel statistical approach, we will test for an interaction between ME2 and BRD2 with respect to juvenile myclonic epilepsy-an IGE subtype. Overall, this propose research should help to elucidate the general role of ME2 as a susceptibility gene for IGE, as well as its particular role in a network of other contributing genetic factors. PUBLIC HEALTH RELEVANCE: Epilepsy affects 2.7 million people in the US alone. Idiopathic Generalized Epilepsy (IGE) accounts for 30% of all epilepsy, starts in childhood or adolescence, and, is thought to have a genetic etiology. Our studies have identified a gene (ME2) as both linked and strongly associated with several different, common IGE syndromes, including Juvenile Myoclonic Epilepsy (JME), Juvenile Absence Epilepsy (JAE), and Epilepsy with Generalized Tonic-Clonic Seizures (EGTCS). We propose to determine how polymorphisms found in the ME2 gene increase susceptibility to a broad class of related epilepsy syndromes.