Mitochondrial DNA, Nuclear DNA Methylation, and Cardiometabolic Disease Traits

项目摘要 能量代谢在人类疾病中起着至关重要的作用。线粒体，细胞的能量来源， 每个细胞都有自己的基因组（mtDNA），每个细胞有数千个拷贝。线粒体DNA编码基因， 能量代谢的蛋白质。我们（由本申请的PI Liu领导）最近发现， 全血中的数量是心血管疾病（CMD）风险因素水平较高的独立预测因子 来自多个祖先的约60，000名参与者。例如，线粒体DNA减少的一个标准差 拷贝数与肥胖（OR=1.15，p= 8 e-31）和代谢综合征的几率增加相关 (OR=1.14，p= 1 e-32），以及增加水平的几个数量性状定义这些疾病。 尽管有这些发现，线粒体DNA与CMD相关的分子基础尚不清楚， 核基因组（nDNA）也编码许多参与线粒体能量产生的蛋白质， 生物合成，因此，线粒体功能的维持需要线粒体DNA的广泛协调， nDNA建立了小鼠nDNA-mtDNA杂交系统。利用这个模型，研究人员发现， 在相同nDNA的杂交小鼠中的差异nDNA甲基化、基因表达和细胞适应性反应， 但线粒体DNA背景不同。此外，我们（由Arking领导，该应用程序的Co-I）确定了几个 甲基化位点与mtDNA拷贝数相关。此外，实验性修改 通过CRISPR-Cas9敲除TFAM的mtDNA拷贝数，TFAM是mtDNA复制的调节因子， 研究表明，mtDNA拷贝数的调节直接驱动特定DNA的nDNA甲基化的变化。 附近转录物的CpG和基因表达。基于这些先前在小鼠模型和我们自己的研究 研究中，我们假设nDNA的甲基化和基因表达介导了mtDNA对 心脏代谢疾病特征。在这项建议中，我们将利用现有资源，包括 基因组测序和多组学在六个大的队列多个祖先;我们将严格测试我们的 通过追求四个具体目标的假设。在目标1和目标2中，我们将进行关联分析，以确定 mtDNA相关的nDNA甲基化位点和基因表达水平。mtDNA特征包括 mtDNA同质和异质突变，以及mtDNA拷贝数。在目标3中，我们将研究 nDNA甲基化和/或基因表达是否介导mtDNA拷贝数的影响， 连续性心脏代谢疾病性状的异质性。在目标4中，我们将进行综合分析， 确定线粒体DNA和心脏代谢疾病特征的基因调控网络。我们还将在功能上 通过编辑的细胞系测试mtDNA对这些基因网络的影响（例如，通过CRISPR-Cas9系统）。的 这项研究项目产生的知识体系将加深我们对分子机制的理解 潜在的线粒体DNA和心脏代谢疾病，这将最终促进新的 心脏代谢疾病的诊断、预防和治疗模式。

PROJECT SUMMARY Energy metabolism plays a critical role in human disease. Mitochondria, the energy powerhouses of the cell, have their own genome (mtDNA) which is present up to thousands of copies per cell. mtDNA encodes genes for proteins of energy metabolism. We (led by Liu, PI of this application) recently discovered that lower mtDNA copy number in whole blood is an independent predictor for higher levels of cardiovascular disease (CMD) risk factors in ~60,000 participants from multiple ancestries. For example, one standard deviation of decrease in mtDNA copy number was associated with increased odds of obesity (OR=1.15, p=8e-31) and metabolic syndrome (OR=1.14, p=1e-32), as well as with increased levels of several quantitative traits defining these diseases. Despite these findings, the molecular basis underlying the association of mtDNA with CMD is unclear because the nuclear genome (nDNA) also encodes many of the proteins engaged in mitochondrial energy production and biosynthesis, and thus, maintenance of mitochondrial function requires extensive coordination of mtDNA and nDNA. A mouse hybrid nDNA-mtDNA system was developed. Using this model, the researchers found differential nDNA methylation, gene expression, and cellular adaptive response in hybrid mice of identical nDNA, but with different mtDNA background. Additionally, we (led by Arking, Co-I of this application) identified several significant DNA methylation sites associated with mtDNA copy number. In addition, experimental modification of mtDNA copy number through knockout via CRISPR-Cas9 of TFAM, a regulator of mtDNA replication, demonstrated that modulation of mtDNA copy number directly drives changes in nDNA methylation of specific CpGs and gene expression of nearby transcripts. Based on these previous studies in mouse model and our own research, we hypothesize that methylation and gene expression of nDNA mediate the effects of mtDNA on cardiometabolic disease traits. In this proposed proposal, we will leverage existing resources, including whole genome sequencing and multi-omics in six large cohorts of multiple ancestries; we will rigorously test our hypothesis by pursuing four specific aims. In Aim 1 and Aim 2, we will perform association analyses to identify mtDNA-associated nDNA methylation sites and gene expression levels, respectively. mtDNA features include mtDNA homoplasmic and heteroplasmic mutations, and mtDNA copy number. In Aim 3, we will investigate whether nDNA methylation and/or gene expression mediates the effects of mtDNA copy number and heteroplasmy on continuous cardiometabolic disease traits. In Aim 4, we will perform integrative analyses to identify gene regulation networks underlying mtDNA and cardiometabolic disease traits. We will also functionally test the impact of mtDNA on these gene networks via edited cell lines (e.g., via CRISPR-Cas9 system). The body of knowledge generated by this research project will deepen our understanding of molecular mechanisms underlying mtDNA and cardiometabolic diseases, which will ultimately facilitate the development of new modalities for the diagnosis, prevention, and treatment of cardiometabolic diseases.