Regulatory Role of CACNA1C Intronic DNA Variation Relevant to Psychiatric Disease

CACNA1C 内含子 DNA 变异与精神疾病相关的调节作用

基本信息

批准号：
8684348
负责人：
YEN PEI CHRISTY CHANG
金额：
$ 23.03万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-04-01 至 2016-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8684348
关键词：
Adult Affect Alleles Animal Model Autopsy Behavior Binding Binding Sites Bioinformatics Biological Biological Assay Biology Bipolar Disorder Brain Cardiovascular system Cells Code Complex Computer Simulation DNA Data Development Diagnosis Disease Electrophoretic Mobility Shift Assay Electroporation Elements Functional disorder Future Gene Expression Gene-Modified Genes Genetic Genetic Risk Genetic Transcription Genetic Variation Genomics Goals Hereditary Disease Human Human Cell Line In Vitro Inbred Mouse Inherited Introns Knowledge L-Type Calcium Channels Laboratories Lead Linkage Disequilibrium Location Maps Mental Depression Mental disorders Messenger RNA Methodology Methods Modeling Molecular Genetics Mood Disorders Mus Mutation National Institute of Mental Health Nucleic Acid Regulatory Sequences Pathogenesis Predisposition Prevention Proteins Publishing Regulation Regulatory Element Reporter Research Research Domain Criteria Resources Risk Risk Factors Role Schizophrenia Staging System Testing Transfection Transgenic Mice Validation Variant base brain tissue chromatin immunoprecipitation disorder risk epigenetic marker experience genetic association genetic variant genome wide association study improved in vitro activity in vivo mouse model neurobehavioral neuropsychiatry novel novel strategies programs public health relevance research study risk variant skills transcription factor vector

项目摘要

DESCRIPTION (provided by applicant): One of the most consistent findings to have emerged from psychiatric disorder genome wide association studies (GWAS) is with CACNA1C, a gene that codes for the alpha1 subunit of a voltage-dependent L-type calcium channel. Consistent with the NIMH Research Domain Criteria (RDOC) initiative, the biological implications of CACNA1C function are relevant to a diagnosis of bipolar disorder, depression, and schizophrenia. However, in spite of strong genetic data implicating sequence variations in CACNA1C as a risk factor, it is not known how genetic variants located within the gene modify risk. All GWAS-identified SNPs in CACNA1C are located within a single large intron 3 and do not lead directly to changes in the sequence of the coded protein. The central hypothesis guiding the present research effort, supported by our preliminary data, is that specific genetic variation in CACNA1C intron 3 modifies regulatory functions that can be bioinformatically predicted and experimentally validated. Our hypothesis is based on our bioinformatic analyses of the regions surrounding associated human SNPs and preliminary data from our in vitro functional validation of a subset of the in silico predictions. This proposal proposes a multiple methodology strategy, consistent with studies already underway in the laboratory. We will, in Specific Aim #1, use a bioinformatics approach to define sequences in human CACNA1C that are likely to harbor regulatory elements. We predict that the human CACNA1C gene harbors putative regulatory elements containing alleles in linkage disequilibrium (LD) with GWAS identified SNPs. Specific Aim #2 proposes to test human candidate regions in reporter vector systems to assess regulatory activity. We will clone putative regulatory elements into reporter vectors to assess their function as modifiers of transcription in vitro, fine map the location of these regulatory elements, and evaluate these elements for putative TF binding using co-transfection and TF-specific gel-shift assays. We predict that psychiatric condition-associated SNPs (or genetic variations inherited with them) will result in allele-specific changes in CACNA1C gene expression and/or altered function through cis-acting regulatory elements and that we will identify proteins that interact with such regions. As these in vitro and cell-based assays incompletely predict endogenous activity, we will map the presence of human regulatory domains in the mouse (Specific Aim #3). We will locate the corresponding regions in the mouse Cacna1c gene, validate their activity in vitro, and assess in vivo TF binding during different developmental stages. Our intent is to plan future in vivo studies with inbred or transgenic mice harboring similar genetic variations. Overall, our studies will improve basic knowledge of CACNA1C regulation, and significantly progress understanding of the mechanism by which CACNA1C gene intronic variation may modify risk for developing psychiatric disorders.

描述（由申请人提供）：从精神障碍基因组广泛关联研究（GWAS）中出现的最一致的发现之一是与Cacna1c（Cacna1c），该基因编码为电压依赖性L型钙通道的alpha1亚基。与NIMH研究领域标准（RDOC）倡议一致，CACNA1C功能的生物学意义与对躁郁症，抑郁症和精神分裂症的诊断有关。然而，尽管有强大的遗传数据将Cacna1c的序列变化视为危险因素，但尚不知道位于基因内的遗传变异如何修饰风险。 CACNA1C中的所有GWAS识别的SNP都位于单个大内含子3中，不会直接导致编码蛋白的序列变化。指导当前研究工作的中心假设在我们的初步数据的支持下，Cacna1c内含子3的特定遗传变异修改了可以在生物信息上预测和实验验证的调节功能。我们的假设基于我们对相关人类SNP的区域的生物信息学分析，以及我们从我们的体外功能验证中的一部分中的一部分硅硅预测中的验证。该建议提出了一种多种方法策略，与实验室中已经正在进行的研究一致。在特定的目标＃1中，我们将使用生物信息学方法来定义可能具有调节元素的人类Cacna1c中的序列。我们预测，与GWAS鉴定的SNP相关的链接不平衡（LD）中的等位基因（LD）中的等位基因的人体CACNA1C基因港口港。具体目的＃2提议测试记者向量系统中的人类候选区域以评估监管活动。我们将推定的定义调节元素插入报告基因载体，以评估其作为体外转录修饰符的功能，详细绘制这些调节元件的位置，并评估这些元素使用共转染和TF特异性凝胶递移测定法对推定的TF结合进行评估。我们预测，与精神病疾病相关的SNP（或与之遗传的遗传变异）将导致CACNA1C基因表达的等位基因特异性变化和/或通过顺式作用调节元素改变功能，并且我们将识别与此类区域相互作用的蛋白质。由于这些在体外和基于细胞的测定中无法完全预测内源性活性，因此我们将绘制小鼠中人类调节结构域的存在（特定目标＃3）。我们将在小鼠CACNA1C基因中找到相应的区域，在体外验证其活性，并在不同的发育阶段评估体内TF结合。我们的目的是用具有相似遗传变异的近交或转基因小鼠计划未来的体内研究。总体而言，我们的研究将提高CACNA1C调节的基础知识，并显着发展对Cacna1c基因内含子差异可能会改变心理疾病风险的机制的理解。