The role of regulatory variants in FZD6 and interacting genes in nonsyndromic cleft lip and palate

项目摘要 非综合征性唇裂伴或不伴腭裂(NSCLP)是最常见的头面部出生缺陷 由于胚胎面部隆起的不完全融合，在嘴唇、主腭部和/或 次要味觉。每700名新生儿中有1名的普遍出生流行率导致135,000名非小细胞肺癌新生儿 每年在全球范围内。虽然多学科治疗现在取得了令人满意的结果，但也有许多 长期存在的健康问题，造成重大的经济和社会心理负担。家族性 聚合和分离分析表明是多基因遗传，但尽管经过几十年的研究，只有一小部分 NSCLP遗传责任的一部分已被确定，留下了很大的知识缺口。这一差距可能是 大多数基因组分析旨在识别外显子/编码变异和罕见突变的结果。最近， 调节基因表达的非编码功能变异已被证明在出生缺陷中发挥作用；我们的 研究和其他人的研究表明，在颅面过程中基因和基因网络的失调 发展有助于NSCLP。这项提议利用了FZD6、LRP5、 WNT途径受体水平的LRP6和Dkk1基因是NSCLP候选基因。在一起，他们 作用于β-连环蛋白介导的信号转导：FZD6与LRP5或LRP6形成共受体复合体，而 Dkk1是这种共受体复合体的抑制物。这些基因在头面部发育中起着关键作用。 对它们表达的严格调控对上唇和上唇的正常形成和融合至关重要。 味觉。该项目的目标是识别FZD6、LRP5、LRP6和FZD6的功能性非编码遗传变异。 Dkk1及其对NSCLP的贡献。为了实现这一目标，我们最初假定优先考虑21个 在FZD6、LRP5、LRP6和Dkk1中发现了10个与转录因子结合的变体 等位基因特有的方式。在这项持续的工作中，荧光素酶报告在胚胎细胞系和活体中进行了分析 在斑马鱼中使用转基因记者的分析将被用来确定这些变异对基因的作用 表情。对于在发育过程中具有细胞特异性和时空效应的变体，关联分析 将在我们庞大的、特点良好的NSCLP家庭中开展。总之，这项工作的结果将是 确定非编码调控变异体在基因控制中的个体和组合贡献 β-连环蛋白Wnt信号在口腔面部发育中的作用。这些结果将使我们更好地了解 NSCLP中的非编码变体，并有可能被翻译到临床环境中，以帮助 NSCLP风险评估。

Project Summary Nonsyndromic cleft lip with or without cleft palate (NSCLP) is the most common craniofacial birth defect resulting from incomplete fusion of the embryonic facial prominences which leaves a gap in the lip, primary palate and/or the secondary palate. The common birth prevalence of 1 per 700 newborns results in 135,000 NSCLP newborns worldwide each year. While multidisciplinary treatments now achieve satisfactory outcomes, there are many long-term health issues that persist, imposing significant economic and psychosocial burdens. Familial aggregation and segregation analyses suggest polygenic inheritance, but despite decades of study, only a small portion of the NSCLP genetic liability has been identified leaving a large knowledge gap. This gap could be the result of most genomic analyses aimed at identification of exonic/coding variants and rare mutations. Recently, noncoding functional variants that regulate gene expression have been shown to play a role in birth defects; our work and that of others demonstrated that dysregulation of genes and gene networks during craniofacial development contributes to NSCLP. This proposal takes advantage of the strong evidence that FZD6, LRP5, LRP6 and DKK1, genes at the receptor level of the WNT pathway are NSCLP candidate genes. Together, they act on the β-catenin mediated signaling: FZD6 forms a co-receptor complex with either LRP5 or LRP6 while DKK1 is an inhibitor of this co-receptor complex. These genes play a critical role in craniofacial development and the tight regulation of their expression is crucial for the normal formation and fusion of the upper lip and palate. The goal of this project is to identify functional noncoding genetic variants in FZD6, LRP5, LRP6 and DKK1 and their contribution to NSCLP. Towards accomplishing this goal, we initially prioritized 21 putatively functional variants in FZD6, LRP5, LRP6 and DKK1 and found 10 variants that bind transcription factors in an allele-specific manner. In this continuing work, luciferase reporter assays in embryonic cell lines and in vivo analysis with transgenic reporters in zebrafish will be used to define the role of these variants on gene expression. For variants with cell-specific and spatiotemporal effects during development, association analyses will be carried out in our large and well-characterized NSCLP families. Together, results from this work will determine both individual and combinatorial contributions of noncoding regulatory variants in genes controlling β-catenin Wnt signaling during orofacial development. These results will enhance our knowledge of the role of noncoding variants in NSCLP and have the potential to be translated into the clinical setting to aid in the assessment of NSCLP risk.