尼古丁成瘾易感基因甲基化及多态性位点的基因表达调控研究

Tobacco is one of the most widely abused substances, causing 5.4 million deaths worldwide. In China, more than 300 million people use tobacco, almost one third of the total smokers in the world. Genetic and epidemiological studies have demonstrated that nicotine dependence (ND) is a complex disorder influenced by both genetic and environmental factors. During the past several years, significant efforts have been made to identify susceptibility genes for ND using both genome-wide linkage and association analysis. However, the results are inconsistent. Moreover, only limited regulatory or nonsynonymous polymorphisms in the promoter or coding regions have been identified. Thus, a search for factors involved in the regulation of gene expression is the next step toward basic biological knowledge of ND. In this application, we propose to investigate the involvement of methylation and allelic variations in six genes in the etiology of ND: CHRNA4, DDC, COMT, BDNF, NTRK2, and ANKK1, which were chosen because of demonstration that they are significantly associated with ND in at least two independent association studies and are modulated by nicotine and other substances of abuse in animal and in vitro studies. Specifically, this application has three Aims: (1) to determine whether the methylation pattern within the promoter regions of CHRNA4, DDC, COMT, BDNF, NTRK2 and ANKK1 is altered in 450 smokers compared with 450 never-regular smokers of different origins; (2) to determine whether any single nucleotide polymorphisms (SNPs) in the promoter regions of the six genes regulate expression using an in vitro reporter gene assay system; and (3) to investigate whether any coding SNPs of the six genes affect mRNA stability. The proposed study represents an important extension of our ongoing search for vulnerability genes to ND in this laboratory. We expect its completion to greatly advance understanding of the epigenetic determinants of ND and eventually allow targeting of novel prevention and treatment strategies to individuals at risk.

遗传学和流行病学研究表明，尼古丁成瘾作为一种复杂疾病是由遗传因素和环境因素共同决定的。在过去几年中，通过大量的全基因组连锁分析和关联分析已寻找到了一批导致尼古丁成瘾的易感基因。然而，这些研究结果之间不仅缺乏普遍的一致性，而且显著关联的各多态性位点的具体功能也不明确。因此，研究易感基因与尼古丁成瘾之间关联的分子机制是尼古丁成瘾遗传学研究的重要一步。本研究拟对CHRNA4、DDC、COMT、BDNF、NTRK2和ANKK1这六个在尼古丁成瘾病因学方面发挥重要作用的易感基因进行甲基化和多态性位点的基因表达调控研究。以上基因不仅与尼古丁成瘾性状呈显著关联，且其关联结果在多项独立研究中也得到了反复验证。 本研究作为尼古丁成瘾易感基因及多态性位点研究的拓展，从表观遗传学和基因表达调控的角度对尼古丁成瘾的分子机制进行了进一步挖掘，其成果将为尼古丁成瘾的预防和戒断提供必要的理论基础。

尼古丁是烟草中的主要成瘾成份。在过去几年中，通过大量的全基因组连锁分析和关联分析已寻找到了一批导致尼古丁成瘾的易感基因。然而，这些研究结果之间尚缺乏普遍的一致性，且表观遗传学的证据也不充分。本研究针对CHRNA4、DDC、COMT、BDNF、NTRK2、ANKK1这六个在尼古丁成瘾病因学方面发挥重要作用的易感基因进行甲基化和多态性位点的研究。以上基因不仅与尼古丁成瘾性状呈显著关联，且其关联结果在多项独立研究中也得到了反复验证。此外，随着研究的展开，我们还加入了DRD2、尼古丁受体、SLC6A3等与尼古丁成瘾密切相关的基因作为本项目的重要补充。本项目通过遗传学和表观遗传学的手段，寻找确定了这些基因中多个与吸烟成瘾相关表型显著关联的遗传变异和甲基化位点。通过研究发现，NTRK2、ANKK1和COMT基因甲基化水平在吸烟人群和不吸烟人群之间存在显著差异，且这些基因中有多个SNP位点也与吸烟成瘾显著关联。此外，通过meta分析我们还发现，ANKK2/DRD2、SLC6A3和尼古丁受体基因在不同人种中与吸烟相关表型存在显著关联。以上结果作为尼古丁成瘾易感基因及多态性位点研究的拓展，将为尼古丁成瘾的预防和戒断提供必要的理论基础。