Identification of human orofacial enhancers and their role in orofacial clefts

描述（由申请人提供）：口面裂是最常见的人类出生缺陷之一，但其根本原因仍在很大程度上未知。全基因组关联研究表明这种缺陷的遗传性，然而绝大多数关联都在基因之外，这表明基因调控缺陷是一个主要因素。拟议的工作旨在确定非综合征性唇腭裂（NSCLP）的遗传原因，通过识别和表征基因调控序列，这些序列在早期人类口面发育中是活跃的，并在受影响的个体中被破坏。利用功能基因组学方法直接在人类胚胎组织中进行的初步研究已经确定了数千个潜在的口面调控序列，其中包括几个与 单核苷酸多态性与NSCLP密切相关。这个“独立之路”奖的申请包括一个指导的职业发展计划的候选人，博士过渡到一个独立的研究员，以及一个附带的研究计划，描述了拟议的实验上的识别和表征疾病相关的变异基因调控序列。候选人Cotney博士是耶鲁大学医学院的博士后研究员，在遗传学系James Noonan博士的实验室工作。他在埃默里大学获得遗传学和分子生物学研究生学位的工作是在埃默里大学生物化学系和耶鲁大学遗传学系的Gerald Shadel博士的实验室进行的。在那里，他专注于了解一类新型转录因子的两个成员对人类线粒体基因表达和逆行信号的贡献。在努南实验室，Cotney博士应用生物化学和计算方法直接在胚胎肢体组织中识别基因调控序列，这些序列有助于人类特有的手和脚的进化。解决胚胎发育中这些进化问题的技术和计算方法可以适用于了解任何发育组织中的基因调控，这使Cotney博士处于研究NSLP原因的独特位置。内详述的指导和职业发展计划将补充他在基础分子生物学和功能基因组学方面的背景，并提供统计遗传学和人类口面发育方面的额外培训和指导。Cotney博士的目标是成为跨学科生物科学，发育生物学或学术机构类似部门的教员，在那里他可以研究发育基因调控功能障碍在常见人类疾病中的作用。该研究计划将利用Cotney博士在胚胎组织功能分析方面的专业知识，以确定在咽弓形成到腭和面部结构融合期间活跃的基因调控序列。该项目进一步提出确定这些激活的调控元件中的哪些在NSCLP患者中具有变体和结构变化，并确定发育基因调控中断的后果。对NSCLP患者中常见干扰的基因调控网络的鉴定将提供可以预测这些缺陷的发生的遗传标记，并为未来的预防或治疗措施提供靶点。减轻口面裂的影响并防止其发生将增加全球公共卫生，并解除受影响个人及其家庭目前面临的巨大经济负担。

DESCRIPTION (provided by applicant): Orofacial clefts are one of the most commonly occurring human birth defects, yet the underlying causes remain largely unknown. Genome wide association studies indicate heritability for such defects, however the vast majority of associations fall outside of genes suggesting defective gene regulation is a major contributor. The proposed work seeks to identify the genetic causes of nonsyndromic cleft lip and palate (NSCLP) by identifying and characterizing gene regulatory sequences that are active in early human orofacial development and disrupted in affected individuals. Preliminary research using functional genomics methods directly in human embryonic tissue has identified thousands of potential orofacial regulatory sequences, including several that are in linkage disequilibrium with single nucleotide polymorphisms strongly associated with NSCLP. This "Pathway to Independence" award application includes a mentored career development plan for transition of the candidate, Dr. Justin Cotney, into an independent investigator, as well an accompanying research plan describing the proposed experiments on identification and characterization of disease-associated variants in gene regulatory sequences. The candidate, Dr. Cotney, is a postdoctoral fellow at Yale University School of Medicine, in the lab of Dr. James Noonan in the Department of Genetics. The work leading to his graduate degree in Genetics and Molecular Biology at Emory University was conducted in the lab of Dr. Gerald Shadel in the Department of Biochemistry at Emory and Department of Genetics at Yale. There he focused on understanding the contributions of two members of a novel class of transcription factors to human mitochondrial gene expression and retrograde signaling. In the Noonan lab, Dr. Cotney applied biochemical and computational methods to identify gene regulatory sequences directly in embryonic limb tissue that have contributed to human-specific evolution of the hand and foot. The techniques and computational methods to address these evolutionary questions in embryonic development can be adapted to understand gene regulation in any developing tissue, placing Dr. Cotney in a unique position to investigate the causes of NSCLP. The mentoring and career development plan detailed within will supplement his background in basic molecular biology and functional genomics with additional training and instruction in statistical genetics and human orofacial development. Dr. Cotney's goal is to become a faculty member in an interdisciplinary bioscience, developmental biology, or similar department at an academic institution, in which he can research the role of dysfunction of developmental gene regulation in common human diseases. The research plan will leverage Dr. Cotney's expertise in functionally profiling embryonic tissue to identify gene regulatory sequences that are active during the formation of pharyngeal arches to fusion of palates and facial structures. The project further proposes to determine which of these activated regulatory elements are burdened with variants and structural changes in NSCLP patients and identify the consequences of disrupted developmental gene regulation. The identification of gene regulatory networks that are commonly perturbed in NSCLP patients will provide genetic markers that can predict the occurrence of these defects and provide targets for future preventative or therapeutic measures. Alleviating the impact of orofacial clefts and preventing their occurrence will increase global public health and lift a large financial burden currently faced by affected individuals and their families.