Nicotinic receptor genes & substance abuse: Functional studies of associated SNPs

烟碱受体基因

• 批准号: 7701421
• 负责人: MARISSA A EHRINGER
• 金额: $ 44.38万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7701421
• 关键词: Affect; Age; Agonist; Alcohol Phenotype; Alcohol dependence; Alleles; Animals; Behavior; Behavioral; Binding; Biological Assay; Cancer cell line; Cell Culture Techniques; Cell Line; Cells; Chromosomes, Human, Pair 15; Cocaine Dependence; Complex; Conduct Disorder; DNA Sequence; Data; Development; Disease; Disinhibition; Drug Experimentation; Drug usage; Embryo; Environment; Future; Gene Cluster; Gene Expression; Genes; Genetic; Genetic Risk; Genetic Variation; Goals; Haplotypes; Histocompatibility Testing; Human; Human Chromosomes; Human Genetics; In Vitro; Individual; Individual Differences; Intercistronic Region; Knowledge; Laboratories; Lead; Ligand Binding; Luc Gene; Luciferases; Malignant neoplasm of lung; Measures; Mediating; Methods; Molecular; Molecular Genetics; Mus; Neurons; Nicotine; Nicotine Dependence; Nicotinic Receptors; Pattern; Personal Communication; Pharmaceutical Preparations; Pharmacogenomics; Positioning Attribute; Preparation; Prevention; Prevention approach; Protein Subunits; Receptor Gene; Research; Research Personnel; Risk; Risk Factors; Series; Signal Transduction; Single Nucleotide Polymorphism; Smoke; Smoking; Smoking Behavior; Substance abuse problem; System; Tissue-Specific Gene Expression; Transcriptional Regulation; Variant; Work; age related; alcohol response; base; blastomere structure; cancer cell; cell type; cigarette smoking; drug abuse related behavior; embryonic cell culture; follow-up; genetic association; immortalized cell; improved; promoter; public health relevance; receptor; receptor expression; receptor function; research study; response; smoking cessation

DESCRIPTION (provided by applicant): Human genetic studies have identified the cluster of nicotinic receptor genes (CHRNA5/A3/B4) on chromosome 15 as strong candidates for association with drug abuse-related behaviors, including smoking, alcohol phenotypes, and cocaine dependence. These genes have also been associated with lung cancer, but it is remains unclear what aspects of this association may be mediated through smoking behavior. In addition, we have found evidence that the intergenic region between the CHRNA3 and CHRNB4 genes is associated with more general measures of disinhibitory behavior, including conduct disorder, which is a known risk factor for early initiation of drugs and later problems. There is evidence from molecular, pharmacological, and animal studies that transcriptional regulation of these genes is likely to be co-regulated and complex, and that the 13 and 24 subunits might be good targets for development of smoking cessation drugs. We have initiated studies to assess the putative functional differences of alleles for SNPs that have been implicated in our human genetic studies. Our results provide evidence that at least two SNPs in this region may lead to differences in gene expression, using luciferase-gene assays in immortalized cell culture lines and in primary mouse neuronal embryonic cell cultures. The goals of this application are to extend these findings in several ways. First, we will use these assays to evaluate how the DNA sequence surrounding these SNPs may affect their ability to differentially drive gene expression ("promoter bashing"). Second, we will examine the effects of these SNPs in different cell types, including a variety of immortalized neuronal cells, lung cancer cells, and primary mouse neuronal embryonic cells (gene x cell type interaction). Third, we will challenge the cells with nicotine to assess whether this drug modifies gene expression patterns (gene x environment interaction). Fourth, we will determine whether allele-specific changes in gene expression using the luciferase assays lead to corresponding differences in protein subunit and receptor expression. Finally, we will assess function of nicotinic receptors whose subunit constituents were generated using our in vitro system containing different alleles for individual SNPs and haplotypes of SNPs. We expect the methods developed and applied to these genes to be easily applied to future studies of other nicotinic receptor genes that may also be associated with drug abuse-related behaviors. PUBLIC HEALTH RELEVANCE: Results from this project will facilitate a better understanding of how naturally occurring variations in the CHRNA3 and CHRNB4 genes might contribute to the underlying molecular mechanisms responsible for individual differences in behaviors relevant to drug use and abuse. Such knowledge should lead to the development of improved prevention and treatment of individuals who suffer from these disorders.

描述（由申请人提供）：人类遗传学研究已确定 15 号染色体上的烟碱受体基因簇 (CHRNA5/A3/B4) 是与药物滥用相关行为（包括吸烟、酒精表型和可卡因依赖）相关的有力候选基因。这些基因也与肺癌有关，但目前尚不清楚这种关联的哪些方面可能是通过吸烟行为介导的。此外，我们发现证据表明 CHRNA3 和 CHRNB4 基因之间的基因间区域与更一般的去抑制行为测量相关，包括品行障碍，这是早期开始用药和随后出现问题的已知危险因素。分子、药理学和动物研究的证据表明，这些基因的转录调控可能是共同调控且复杂的，并且 13 和 24 亚基可能是开发戒烟药物的良好靶点。我们已经启动了研究来评估人类遗传学研究中涉及的 SNP 等位基因的假定功能差异。我们的结果提供证据表明，在永生化细胞培养系和原代小鼠神经元胚胎细胞培养物中使用荧光素酶基因测定，该区域中至少有两个 SNP 可能导致基因表达差异。该应用程序的目标是以多种方式扩展这些发现。首先，我们将使用这些测定来评估这些 SNP 周围的 DNA 序列如何影响它们差异驱动基因表达的能力（“启动子打击”）。其次，我们将检查这些 SNP 在不同细胞类型中的影响，包括各种永生化神经元细胞、肺癌细胞和原代小鼠神经元胚胎细胞（基因 x 细胞类型相互作用）。第三，我们将用尼古丁挑战细胞，以评估该药物是否改变基因表达模式（基因 x 环境相互作用）。第四，我们将使用荧光素酶测定确定基因表达的等位基因特异性变化是否导致蛋白质亚基和受体表达的相应差异。最后，我们将评估烟碱受体的功能，其亚基成分是使用我们的体外系统生成的，该系统包含个体 SNP 和 SNP 单倍型的不同等位基因。我们期望开发并应用于这些基因的方法能够轻松应用于其他烟碱受体基因的未来研究，这些基因也可能与药物滥用相关行为有关。 公共卫生相关性：该项目的结果将有助于更好地了解 CHRNA3 和 CHRNB4 基因中自然发生的变异如何促成导致药物使用和滥用相关行为个体差异的潜在分子机制。这些知识应该有助于改进对患有这些疾病的个体的预防和治疗。