Association and Function of Opioid Receptor Gene Variants to Substance Dependence

TFIIS是一个从K99过渡到Roo的应用程序，它为研究四个人的关联寻求进一步的支持 阿片受体基因(0PRM1、0PRD1、0PRK1和0PRL1)和四个阿片肽基因(POMC、 PDYN、PENK和PNOC)基因与非裔美国人(AAs)和 欧洲裔美国人(EA)。 在K99期，我们鉴定了几个opold受体和肽基因变异体，它们显著地 与SD关联。然而，由于只选择了有限数量的变种并对其进行了测试 与SD关联时，因果变量可能被忽略。此外，目前还不清楚 已确定的SD相关变异是功能性的，或者它们只是与原因连锁不平衡 变种。因此，在Roo阶段，我们需要进一步解决这些问题。我们建议(1)分析 通过对阿片受体和多肽重新测序发现常见和罕见变异与SD的关系 在我们的病例和对照中使用下一代测序技术的基因；(2)检查 SD相关阿片受体和肽基因变体使用包括受体在内的几种方法 结合分析、Western blotting、实时定量聚合酶链式反应、等位基因表达 不平衡分析、凝胶迁移率变化分析、甘草素酶报告基因分析； 研究阿片受体和多肽基因调控区的DNA甲基化水平 在SD患者和健康对照组之间有显著差异。 建议的研究将提高我们对阿片受体和多肽的影响的理解。 SD的基因变异，并确定这些基因的表观遗传修饰是否可以增加 SD漏洞。此外，它将为未来的ROI项目生成足够的数据，目标是(1) 建立一套阿片受体和多肽基因标记物作为SD的预测因子；以及(2)研究 这些基因的变异对SD治疗结果的贡献。

Tfiis is a K99 to ROO transition application which seeks further support for studying the association of four opioid receptor genes (0PRM1, 0PRD1, 0PRK1, and 0PRL1) and four opioid peptide genes (POMC, PDYN, PENK, and PNOC) genes and substance dependence (SD) in African Americans (AAs) and European Americans (EAs). In the K99 phase, we identified several opold receptor and peptide gene variants which were significantly associated with SD. However, since only a limited number of variants were selected and tested for their association with SD, the causal variant might have been ignored. In addition, it is unknown whether the identified SD-associated variants are functional or they are just in linkage disequilibrium with the causal variant. Therefore, in the ROO phase, we need to further address these Issues. We propose to (1) analyze the association of both common and rare variants with SD by resequencing the opioid receptor and peptide genes in our cases and controls using the next-generation sequencing technique; (2) examine the function of SD-assoclated opioid receptor and peptide gene variants using several approaches including receptor binding assays, Western blotting, real-time quantitative polymerase chain reaction, allelic expression Imbalance assay, electrophoretic mobility gel shift assay, and liclferase reporter gene assay; and (3) Investigate whether DNA metylatlon levels In the regulatory regions ofthe opioid receptor and peptide genes are significantly different between SD affected cases and healthy controls. The proposed study will improve our understanding about the Influence of opioid receptor and peptide gene variants on SD and determine whether epigenetic modification ofthese genes can Increase vulnerability to SD. In addition, it will generate sufficient data for a future ROI project aimed at (1) establishing a set of opiod receptor and peptide gene markers as predictors of SD; and (2) investigating the contribution of variants In these genes to the outcome of SD treatment.