A Twin Approach for Genome-Wide Differential DNA Methylation in Orofacial Clefting

摘要 非综合征性口面部裂(OFCs)是人类最常见的头面部出生缺陷，影响 全球约每700人中就有1人感染。全基因组链接和关联研究已经确定 OFC的几个风险等位基因；然而，它们只占其估计遗传力的一小部分。因此， 虽然正在进行的遗传和表达研究可能会导致在理解 OFC的生物学基础，一个更全面的图景只会随着易感变异的相互作用而浮现 与其他因素，如表观遗传变化，是确定的。表观遗传修饰可能是一种机制 环境因素和遗传变异可能通过这种方式改变基因表达。DNaM是一种共价键 在胞嘧啶核苷酸中添加甲基(CH3)基团，可导致转录水平的变化。 靶基因的活性。我们假设甲基化的变化和由此产生的差异基因 表达是OFC的表观遗传风险因素，dNaM介导了OFC风险的遗传影响。我们的 主要目标是表征可能影响离岸金融中心风险的全基因组dNaM变异；并探索 遗传-表观遗传互作(MeQTL)参与OFC的病因学。为了做到这一点，这个项目提出了一个 强大的策略使用(到目前为止)最大的单卵双胞胎和兄弟姐妹对样本集 用于研究表观遗传风险因素的非综合征性OFCS。这是关键的一步，也是一种创新的方法 将允许发现常规GWAs和DNA测序看不到的表观遗传学风险因素 方法：研究方法。这项建议的中心假设将以以下具体目标进行检验：(1) 确定与OFCS不一致的MZ双胞胎的全基因组DNA甲基化模式；(2)探索 遗传-表观遗传相互作用(MeQTL)参与了OFCS的病因学。在Aim 2中，我们将尝试 复制在初步分析中确定的遗传-表观遗传相互作用(从第一次获得 提交本提案)，以及目标1产生的最高点击率，使用裂隙不协调的独立队列 兄弟姐妹。杰出的导师团队(Jeff Murray博士、Robert Philbert博士、Mary Marazita博士和 莫雷诺-乌里韦博士)、顾问(谢博士和德雷克博士)、合作者(李博士)和顾问(阿莫特博士、安德鲁博士)。 Wehby和Butali博士)涵盖了急需的额外培训的所有领域 完成拟议的研究和培训目标所必需的。他们将提供指导，并促进 在向独立过渡的过程中，PI的成长。虽然一开始训练很好，但这五年的间隔 在PI的职业生涯中证明了在快速发展的表观遗传学和大数据领域需要额外培训的合理性 申请。拟议的培训和研究目标是根据PI以往的经验量身定做的 并提供表观遗传学和分析方法方面的额外培训，以推动PI 发展为能够整合表观遗传学和遗传学数据以研究复合体的独立研究人员 头面部先天缺陷。

Abstract Nonsyndromic orofacial clefts (OFCs) are the most common craniofacial birth defects in humans, affecting approximately 1 in 700 individuals worldwide. Genome-wide linkage and association studies have identified several risk alleles for OFCs; however, they account for a minority of their estimated heritability. Therefore, while the ongoing genetic and expression studies may lead to important advances in understanding the biological basis underlying OFCs, a fuller picture will only emerge as the interaction of susceptibility variants with other factors, such as epigenetic changes, are established. Epigenetic modification is a likely mechanism through which environmental factors and genetic variation may alter gene expression. DNAm is a covalent addition of a methyl (CH3) group to the nucleotide cytosine, which can lead to changes in transcriptional activity of the targeted gene. We hypothesize that changes in methylation and the resulting differential gene expression is an epigenetic risk factor for OFCs, and that DNAm mediates genetic influences in OFCs risk. Our main goals are to characterize genome-wide DNAm variation that can impact the risk for OFCs; and explore the genetic-epigenetic interactions (meQTLs) involved in the etiology of OFC. To do so, this project proposes a powerful strategy using the largest sample set (to date) of monozygotic twins and sibling pairs discordant for nonsyndromic OFCs used to study epigenetic risk factors. This is a key step and an innovative approach that will allow uncovering epigenetics risk factors that are invisible to conventional GWAS and DNA sequencing methods. The central hypothesis of this proposal will be tested with the following specific aims: (1) To determine genome-wide DNA methylation patterns in MZ twins discordant for OFCs; (2) To explore the genetic-epigenetic interactions (meQTLs) involved in the etiology of OFCs. Within aim 2 we will attempt to replicate the genetic-epigenetic interactions identified in the preliminary analysis (obtained since the first submission of this proposal), and top hits resulting from Aim 1, using an independent cohort of cleft discordant sibling pairs. The outstanding team of mentors (Dr. Jeff Murray, Dr. Robert Philibert, Dr. Mary Marazita, and Dr. Moreno-Uribe), consultants (Dr. Xie, and Dr. Drake), collaborator (Dr. Lie), and advisors (Dr. Amendt, Dr. Wehby and Dr. Butali) assembled in this proposal covers all areas of the much needed additional training necessary to accomplish the proposed research and training aims. They will provide guidance and facilitate the growth of the PI during the transition to independence. Although very well trained initially, the five years hiatus in the PI's career justify the need for additional training in the rapidly moving fields of epigenetics and big data applications. The proposed training and research aims are tailored to build upon the PI's previous experience and to provide the additional training in epigenetics and analytical methods necessary to propel the PI's development as an independent researcher able to integrate epigenetic and genetic data to study complex craniofacial birth defects.