Genetic and functional analyses of a novel locus associated with LDL-C and MI

项目总结 这项建议将有助于首席调查员的职业目标，即促进对 血脂性状和心肌梗死(MI)的遗传基础并将这一理解应用于预防 病人中的疾病。基于他丰富的研究经验，该提案将培训他使用 互补方法-人类复杂心血管特征的遗传学，以及小鼠模型的使用 和胚胎干细胞对与这些特征相关的基因进行功能验证--以实现生物学上的 对全基因组关联研究(GWAS)结果的理解。这也将为他提供说教 统计遗传学、遗传学和基因组学方法、实验生物学和研究伦理方面的培训。 最后，它将促进他向独立研究职位的过渡，并为他提供强有力的科学 申请R01级资助的基金会。 首席调查员将能够利用绝对独特的环境 心血管疾病动物模型的优势(马萨诸塞州总医院心血管疾病 心血管遗传学和基因组学(麻省理工学院博德研究所和马萨诸塞州哈佛大学 总医院人类遗传研究中心)和干细胞生物学(马萨诸塞州总医院 医院心血管研究中心、哈佛干细胞研究所)完成拟议的工作。 提出这项研究计划的理由是，对人类有效的新疗法的发现 心血管疾病需要在人类身上识别和验证新的疾病机制。 最近，对基因组变异的研究进入了一个新的阶段，在这个阶段，无偏见的GWAS研究可以识别 常见疾病的新基因贡献者。在过去的两年里，超过50个新的和可复制的 发现了影响血脂水平、心肌梗死和2型糖尿病的基因座。 尽管这些观察结果可能是有希望的，但要将新的联想转化为 治疗。关键的一步是确定作图基因座中的因果DNA变异。多个基因座包含多个 基因；其他基因座不包含已知的基因，这表明它们可能作用于基因座外的反式基因。 因此，有必要明确识别每个基因座调控的因果基因(S)。最后，还有一个 需要解决基因座上的DNA变异影响因果基因的机制。简而言之，一条道路必须 从基因组定位到新的机械洞察力。 这项建议探索了一条为低密度脂蛋白胆固醇(LDL-C)确定新的遗传基因座的途径。 C)和MI。在初步研究中，GWA检测低密度脂蛋白和心肌梗死，并发现了单核苷酸 染色体1p13.3上与低密度脂蛋白密切相关的基因座的单核苷酸多态性(SNPs) 比基因组中的任何其他基因座都强-极小的低密度脂蛋白(VsLDL)颗粒浓度和MI。 携带这些SNPs主要等位基因的两个副本的个体的低密度脂蛋白/分升高16毫克/分升，高出40% VsLDL水平，与次要等位基因纯合子相比，MI的风险增加18%。 1p13.3上的相关基因组区间跨度约100kb，包含4个基因-CELSR2， PSRC1、MYBPHL、SORT1-与低密度脂蛋白-C调节没有建立联系。 该提案的核心问题是：(1)哪些1p13.3 DNA变异会导致低密度脂蛋白-C、vsLDL和心肌梗死，(2) 哪些1p13.3基因对低密度脂蛋白、低密度脂蛋白和心肌梗死有贡献，以及(3)DNA变异通过什么机制 会影响基因功能吗？小鼠的遗传精细定位、基因重测序、基因敲除和过度表达， 而人类胚胎干细胞的实验将被用来回答这些问题。成功应该是结果 在精确定义一个新的致病基因(S)及其调控机制，从而提供了一个潜在的新的 治疗血脂异常和预防心肌梗塞的目标。

PROJECT SUMMARY This proposal will facilitate the principal investigator's career goals of advancing the understanding of the inherited basis for lipid traits and myocardial infarction (MI) and applying this understanding to the prevention of disease in patients. Building on his extensive research experience, the proposal will train him in the use of complementary approaches-human genetics of complex cardiovascular traits, and the use of mouse models and embryonic stem cells for functional validation of genes associated with these traits-to achieve a biological understanding of the results of genome-wide association studies (GWAS). It will also provide him with didactic training in statistical genetics, genetic and genomic methods, experimental biology, and research ethics. Finally, it will promote his transition to an independent research position and provide him a robust scientific foundation from which to apply for R01-level funding. The principal investigator will be able to take advantage of an absolutely unique environment with diverse strengths in animal models of cardiovascular disease (Massachusetts General Hospital Cardiovascular Research Center), cardiovascular genetics and genomics (Broad Institute of MIT and Harvard, Massachusetts General Hospital Center for Human Genetic Research), and stem cell biology (Massachusetts General Hospital Cardiovascular Research Center, Harvard Stem Cell Institute) to complete the proposed work. The rationale for the proposed research plan is that the discovery of new and effective treatments for human cardiovascular diseases requires the identification and validation in humans of novel disease mechanisms. Recently, studies of genomic variation entered a new phase, in which unbiased GWAS studies can identify novel genetic contributors to common diseases. In the last two years, more than fifty new and reproducible genomic loci were discovered contributing to lipid levels, MI, and type 2 diabetes. As promising as these observations may be, much work will be needed to convert novel associations into therapies. A key step is to identify the causal DNA variants in the mapped loci. Many loci contain multiple genes; other loci contain no known genes, suggesting that they may act in trans on genes outside of the loci. Thus, there is a need to unambiguously identify the causal gene(s) regulated by each locus. Finally, there is a need to address the mechanisms by which DNA variants in loci affect the causal genes. In short, a path must be blazed from genomic localization to new mechanistic insights. This proposal explores such a path for new genetic loci identified for low-density lipoprotein cholesterol (LDL- C) and MI. In preliminary studies, GWAS for LDL-C and MI and have identified single nucleotide polymorphisms (SNPs) in a locus on chromosome 1p13.3 that is robustly associated with LDL-C-more strongly than any other locus in the genome-very small LDL (vsLDL) particle concentration, and MI. Individuals who carry two copies of the major alleles of these SNPs have 16 mg/dl higher LDL-C, 40% higher vsLDL levels, and 18% greater risk of MI when compared with homozygotes for the minor alleles. The associated genomic interval on 1p13.3 spans ~100 kilobases (kb) and harbors four genes-CELSR2, PSRC1, MYBPHL, SORT1-with no established links to LDL-C regulation. The proposal's core questions are: (1) which 1p13.3 DNA variants contribute to LDL-C, vsLDL, and MI, (2) which 1p13.3 genes contribute to LDL-C, vsLDL, and MI, and (3) by what mechanisms do the DNA variants affect gene function? Genetic fine-mapping, gene resequencing, gene knockdown and overexpression in mice, and experiments in human embryonic stem cells will be used to answer these questions. Success should result in precise definition of a novel causal gene(s) and regulatory mechanism and thereby provide a potential new target for the treatment of dyslipidemia and prevention of MI.