GENETIC VARIATION IN APOLIPOPROTEIN E EXPRESSION

来自遗传流行病学研究的数据表明，APOE位点的遗传变异与阿尔茨海默病（AD）易感性的关系可能不仅限于已被充分研究的编码多态性，还可能受到侧翼DNA多态性的影响。特别是，我们最近观察到APOE e3/e3基因型人群中apo1 HpaI+等位基因与AD的关联。在本项目中，我们将进行分子分析和实验来验证APOE侧翼序列影响AD易感性的假设，即这种影响是通过这些多态性对等位基因特异性APOE mRNA表达的影响。首先，为了测试AD易感性是否取决于APOE编码和位于染色体顺式的调控多态性的特定组合，我们将开发一种方法来确定APOE启动子、APOE编码区和APOC1启动子中多态性标记的完整单倍型，我们将在项目1中讨论的大型多民族群体样本中生成用于AD关联分析的单倍型数据。其次，使用转染试验筛选，我们将检测APOE和APOC1启动子中是否存在功能序列多态性，我们将确定APOE和APOC1启动子中功能序列多态性的序列，我们将确定APOE和APOC1启动子中这些多态性的序列，我们将确定这些多态性的序列，我们将把它们作为单倍型分析的新标记。第三，我们将通过将个体多态性和单倍型与人淋巴母细胞样细胞系和肝脏、肾脏和脑组织中等位基因特异性APOE mRNA水平的相关性来测试它们在体内的功能意义。第四，使用apoCI启动子“敲除”小鼠，我们将测试APOE和APOC1参与增强竞争的模型，以解释apo1 HpaI+等位基因与AD的关联。通过转染和转基因小鼠，我们还将研究假定的APOE脑增强子的定位，然后寻找该区域的功能多态性。这些研究的结果可能揭示了阿尔茨海默病的基本机制，可以进行药物干预，也可能揭示了在不同种族人群中观察到的阿尔茨海默病明显风险的差异。

Data from genetic epidemiological studies suggest that the involvement of genetic variation at the APOE locus in Alzheimer's disease (AD) susceptibility may not be restricted to the well-studied coding polymorphisms, but may also be influenced by polymorphisms in flanking DNA. In particular, we have recently observed an association of the APOC1 HpaI+ allele with AD in groups of people with the "reference" APOE e3/e3 genotype. In this project we will carry out molecular analyses and experiments to test the hypothesis that APOE flanking sequences influence AD susceptibility is that this effect is through the influence of these polymorphisms on allele-specific APOE mRNA expression. First, to test whether AD susceptibility depends on particular combinations of APOE coding and regulatory polymorphisms located in cis on the chromosome, we will develop a method for determining complete haplotypes of polymorphic markers in the APOE promoter, the APOE coding region and the APOC1 promoter and we will generate haplotype data for AD association analysis in the large multi-ethnic population samples discussed in Project 1. Second, using a transfection assay screen, we will test for the existence of functional sequence polymorphisms in the APOE and APOC1 promoters, we will determine the sequence of functional sequence polymorphisms in the APOE and APOC1 promoters, we will determine the sequence of these polymorphisms in the APOE and APOC1 promoters, we will determine the sequence of these polymorphisms and we will apply them as new markers in the haplotype analyses. Third, we will test for functional significance for the individual polymorphisms and haplotypes in vivo by correlating them with allele-specific APOE mRNA levels in human lymphoblastoid cell lines and liver, kidney and brain tissues. Fourth, using apoCI promoter "knockout" mice, we will test a model in which the APOE and APOC1 engage in enhance-competition as an explanation for the observed association of the APOC1 HpaI+ allele with AD. Using transfections and transgenic mice we will also investigate localization of the putative brain-enhancer of APOE and then search for functional polymorphisms in this region. Results of these studies may reveal a basic mechanism for AD which is accessible to pharmacological intervention and may also shed light on the difference sin apparent AD risk who have been observed in different ethnic groups.