Interpreting the regulatory mechanisms underlying the predisposition to substance use disorders

摘要/摘要 物质使用障碍对人类健康和福祉有着深远的影响。的发展 由于神经回路的复杂性和神经系统的复杂性， 潜在的遗传机制。最近的全基因组关联研究已经开始确定一些位点 与成瘾倾向相关，更多的基因座与较低的信心有关。的 大多数与复杂大脑疾病相关的遗传变异，包括物质使用，可能会 位于非编码调节区，特别是增强子，而不是蛋白质编码基因内。 尽管增强子区域在大脑中很重要，但研究的计算和实验工具 它们的功能仍处于婴儿期。 为了努力破译这一关键的生物机制，物质使用障碍的基础，我们 寻求建立一个框架，研究人类和小鼠大脑增强子区域的体内功能。第一、 我们将分析实验室可用的遗传和表观遗传数据集，以确定人类的遗传变异， 可能影响物质使用倾向的增强子区域。接下来，我们将衡量 该基因变异使用高通量报告基因测定，其具有同时测试 在不同条件下，成千上万的增强子在小鼠大脑中的体内活性。最后我们将 在小鼠中使用CRISPR干扰保守的orthogonal增强子来验证这些预测。中 在最初的测试案例中，我们将应用我们的方法来解释吸烟行为的全基因组关联研究 使用细胞类型特异性人类表观基因组学。从计算分析中得出的关键候选人 将在尼古丁暴露的小鼠模型中进行筛选和验证。 我们研究工作的结果将是一张将高置信度和低置信度物质联系起来的地图 使用相关的遗传变异神经增强功能。此外，灵活的计算和 实验框架具有研究候选增强子和遗传修饰的任何组合的潜力。 在不同的大脑区域，不同的细胞类型和不同的动物模型中的变异。

Summary/Abstract Substance use disorders have a profound impact on human health and wellbeing. The development of new treatments for these disorders has remained difficult due to the complexity of the neural circuits and the underlying genetic mechanisms. Recent genome-wide association studies have begun to identify a few of loci associated with the predisposition to addiction, with many more loci are implicated with lower confidence. The majority of the genetic variants associated with complex brain disorders, including substance use, are likely to be located in non-coding regulatory regions, particularly enhancers, and not within protein-coding genes. Despite the importance of enhancer regions in the brain, the computational and experimental tools to study their function are still in their infancy. To work towards deciphering this critical biological mechanism underlying substance use disorders, we seek to build a framework to study the function of both human and mouse brain enhancer regions in vivo. First, we will analyze publically available genetic and epigenetic data sets to identify human genetic variation at enhancers regions that is likely to influence substance use predisposition. Next, we will measure the impact of that genetic variation using a high-throughput reporter assay, which has the ability to simultaneously test the activity of thousands of enhancers across the mouse brain in vivo under different conditions. Finally, we will validate those predictions using CRISPR interference on conserved orthologous enhancers in the mouse. In an initial test case, we will apply our methods to interpret a genome-wide association study of smoking behavior using cell type-specific human epigenomics. The key candidates that result from the computational analysis will be screened and validated in a mouse model of nicotine exposure. The result of our research effort will be a map that links high-confidence and low-confidence substance use-associated genetic variation to neural enhancer function. Furthermore, the flexible computational and experimental framework has the potential to study any combination of candidate enhancers and genetic variants in the context of different brain regions, different cell types, and different animal models.