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

摘要 非综合征性口面裂（OFC）是人类最常见的颅面出生缺陷， 全世界每700人中就有1人。全基因组连锁和关联研究已经确定 OFC的几个风险等位基因;然而，它们占估计遗传力的少数。因此，我们认为， 虽然正在进行的遗传和表达研究可能会导致理解的重要进展， OFC的生物学基础，只有在易感性变体的相互作用下， 与其他因素，如表观遗传变化，建立。表观遗传修饰是一种可能的机制 环境因素和遗传变异可通过该途径改变基因表达。DNAm是共价键 在核苷酸胞嘧啶上添加甲基（CH 3），这可能导致转录水平的变化。 目标基因的活性。我们假设甲基化的变化和由此产生的差异基因 DNAm表达是OFCs的表观遗传风险因素，DNAm介导OFCs风险的遗传影响。我们 主要目标是表征可能影响OFC风险的全基因组DNAm变异;并探索 遗传-表观遗传相互作用（meQTL）参与OFC的病因。为此，该项目提出了一个 强大的战略，使用最大的样本集（迄今为止）的单卵双胞胎和兄弟姐妹对不一致， 用于研究表观遗传风险因素的非综合征OFC。这是一个关键步骤，也是一种创新方法， 将允许发现常规GWAS和DNA测序不可见的表观遗传学风险因素 方法.本提案的中心假设将通过以下具体目标进行检验：（1）为了 确定与OFC不一致的MZ双胞胎的全基因组DNA甲基化模式;（2）探讨OFC与MZ双胞胎基因组DNA甲基化的关系。 遗传-表观遗传相互作用（meQTL）参与OFC的病因。在目标2中，我们尝试 为了复制在初步分析中鉴定的遗传-表观遗传相互作用（自第一次分析以来获得）， 提交本提案），以及目标1产生的最高命中率，使用一个独立的裂缝不一致队列， 兄弟姐妹杰出的导师团队（Jeff Murray博士、Robert Philibert博士、玛丽Marazita博士和 博士莫雷诺乌里韦），顾问（谢博士和德雷克博士），合作者（李博士），和顾问（阿门特博士，博士。 Wehby和Butali博士）在这份建议书中汇集的内容涵盖了急需的额外培训的所有领域 为实现拟议的研究和培训目标所必需的。他们将提供指导和便利， 在向独立过渡期间，PI的增长。虽然一开始训练得很好， 在PI的职业生涯中，需要在表观遗传学和大数据等快速发展的领域进行额外的培训， 应用.拟议的培训和研究目标是根据PI以前的经验量身定制的 并提供必要的表观遗传学和分析方法的额外培训，以推动PI的 发展成为一个独立的研究人员，能够整合表观遗传和遗传数据，研究复杂的 颅面先天缺陷

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.