Determining the Molecular Chain of Causality Through Which Genetic Variants Affect Physiology

项目摘要 生物医学研究的一个中心目标是破译复杂特征的遗传基础。尽管全基因组 关联研究已经成功地检测到数千种与复合体相关的变异 心血管疾病、自身免疫疾病和神经系统疾病，其分子机制仅为人们所知 这些风险相关变种的有限子集。对风险相关变种的机械性理解是 很难确定，因为GWAS确定的绝大多数变种都位于监管区域， 这表明基因表达的变异是复合体遗传风险的重要组成部分。 人类疾病。然而，将基因表达变异映射到遗传变异的统计能力往往是有限的 在这类地图研究中使用的样本量很小。我们刻画分子链特征的能力 将遗传变异与复杂的生理特征联系起来的因果关系进一步受到限制，因为越来越多的证据 调控变异体通常在特定的细胞和组织类型中表现出与疾病相关的效应。因为 在这些限制中，我将利用线虫线虫的遗传多样性来实现 精确量化遗传变异具有的细胞和组织特异性效应所需的统计能力 在基因表达和生理学上。 我们最近开发了一种技术来识别影响细胞和组织特异性的遗传变异 实验线虫杂交中的基因表达(表达数量性状基因座或eQTL)。这种方法 利用线虫短暂的生命周期，很容易产生数十万 重组个体，以及成熟的细胞准备方法，用于单细胞RNA测序 将单细胞转录物丰度与实验杂交中的遗传变异分离联系起来。通过 将这种单细胞eQTL作图方法与实验进化相结合，我已经识别了几个基因组 影响生物体适应性和组织特异性基因表达变化的区域有数十到数百个基因。 在目标1中，我将扩大这些初步实验的范围，以调查各种线虫的影响 生物体适合性以及细胞和组织特异性基因表达的自然遗传变异。在《目标2》中，我将使用 线虫中可用的大量分子和遗传工具箱，用于确定相同的潜在变异是否会影响 既有组织特异性基因表达，又有适合性。这些目标的完成将1)描述细胞的特征--以及 数十万个遗传变异的组织特异性表型效应；2)决定是否组织特异性 表达差异可以影响生物体生理；以及3)提供了一种机制上的理解 遗传变异是其对生物体生理影响的中介。总之，这些见解将有助于 解释监管变异如何影响人类健康。

Project Summary A central goal of biomedical research is to decipher the genetic basis of complex traits. Though genome-wide association studies (GWAS) have successfully detected thousands of variants that are associated with complex cardiovascular, autoimmune, and neurological diseases, the molecular mechanisms are only known for a very limited subset of these risk-associated variants. A mechanistic understanding of risk-associated variants is difficult to ascertain because a vast majority of variants identified by GWAS are located in regulatory regions, which suggests that gene expression variation contributes a substantial portion of the genetic risk for complex human diseases. However, statistical power to map gene expression variation to genetic variants is often limited by the small sample sizes used in such mapping studies. Our ability to characterize the molecular chain of causality that links genetic variants to complex physiological traits is further limited because evidence is mounting that regulatory variants often manifest their disease-associated effects in specific cell and tissue types. Because of these limitations, I will leverage genetic diversity in the nematode Caenorhabditis elegans to achieve the statistical power necessary to precisely quantify the cell- and tissue-specific effects that genetic variants have on gene expression and physiology. We have recently developed a technique to identify genetic variants that affect cell- and tissue- specific gene expression (expression quantitative trait loci or eQTL) in experimental C. elegans crosses. This approach takes advantage of the short life cycle of C. elegans, the ability to easily generate hundreds of thousands of recombinant individuals, and well-established methods to prepare cells for single-cell RNA sequencing to associate single-cell transcript abundance with genetic variation segregating in experimental crosses. By combining this single-cell eQTL mapping approach with experimental evolution, I have identified several genomic regions that affect organismal fitness and tissue-specific gene expression variation of tens to hundreds of genes. In Aim 1, I will extend the scope of these initial experiments to survey the effects of a wide-range of C. elegans natural genetic variation on organismal fitness and cell- and tissue- specific gene expression. In Aim 2, I will use the vast molecular and genetic toolkit available in C. elegans to determine if the same underlying variants affect both tissue-specific gene expression and fitness. The completion of these aims will 1) characterize the cell- and tissue-specific phenotypic effects of hundreds of thousands of genetic variants; 2) determine if tissue-specific expression differences can affect organismal physiology; and 3) provide a mechanistic understanding of how genetic variation mediates its effect on organismal physiology. Together, these insights will facilitate the interpretation of how regulatory variation affects human health.