Discovering Human Birth Defect Genes from Chromosomal Rearrangements

描述（由申请人提供）：虽然复杂疾病对人类遗传学来说是一个严峻的挑战，但出生缺陷代表了一类基本上尚未解决的复杂疾病。由于表型和遗传异质性、环境因素的作用以及扩展谱系和分层良好群体的稀有性，出生缺陷很难通过传统的遗传学方法进行研究。尽管如此，出生缺陷造成的社会负担是巨大的。该应用程序将利用有效的、已建立的出生缺陷基因发现管道，该管道专注于与平衡染色体重排相关的重要出生缺陷。在这些情况下，我们将识别被重排断点破坏的基因，并且我们将检验被断点破坏的基因与受试者的表型有关的假设。为了测试所确定的候选基因的有效性，将采用两种进一步的策略：（1）对不涉及染色体重排的表型相似病例进行突变分析，以寻找候选基因中的基因内突变，以及（2）通过候选基因的靶向突变或敲低来重演先证者表型的关键方面。这些分析的终点是获得确凿的证据，证明在任何特定情况下因染色体重排而破坏的基因是导致相关出生缺陷的原因，并对基因的发育功能进行初步分析。为了实现这些目标，我们将采用强大的权责发生制从国家外展和主要商业细胞遗传学提供商处招募感兴趣的病例。为了最大限度地利用我们实验室现有的专业知识，我们将把研究范围限制在每​​年两个病例（总共十个），并且我们将特别关注影响眼睛、肾脏和颅面复合体器官发生的病例。在目标 1 中，我们将确定涉及与颅面、眼部或肾脏发育障碍相关的平衡染色体重排（即易位和倒位）的病例，并且我们将绘制重排断点。在目标 2 中，我们将识别被断点破坏的候选基因并确定相关的突变机制。在目标 3 中，为了确定这些候选基因的因果关系，我们将对表型相关的人类病例进行突变分析，在目标 4 中，我们将进一步寻求通过使用遗传手段重现小鼠表型来建立因果关系，并且我们还将确定基因的发育功能。这种强大的基因发现方法将识别与医学上重要的出生缺陷新近相关的人类基因，并帮助确定其发育功能。公众健康启示：这项资助旨在识别因与先天性出生缺陷相关的自然发生的人类染色体异常而被破坏的基因。我们将重点关注涉及颅面部区域（例如唇裂和腭裂）、眼睛（例如先天性白内障）和肾脏（例如膀胱输尿管反流或 VUR）的出生缺陷。为了证明损坏的基因与伴随的出生缺陷存在因果关系，我们将在其他患有相同出生缺陷的患者中寻找相同基因的突变，并且我们将研究小鼠中基因突变的后果。

DESCRIPTION (provided by applicant): Although complex diseases represent a critical challenge for human genetics, birth defects represent one class of complex diseases that remain largely unresolved. Because of phenotypic and genetic heterogeneity, the role of environmental factors, and the rarity of extended pedigrees and well-stratified cohorts, birth defects are difficult to study by conventional genetic methods. Nonetheless, the societal burden imposed by birth defects is enormous. This application will take advantage of an efficient, established birth defect gene discovery pipeline that focuses on important birth defects associated with balanced chromosomal rearrangements. In these cases, we will identify genes that are disrupted by the rearrangement breakpoints, and we will test the hypothesis that the genes disrupted by breakpoints are causal for the subject's phenotype. To test the validity of the candidate genes identified, two further strategies will be employed: (1) mutational analysis of phenotypically similar cases that do not involve chromosomal rearrangements, to search for intragenic mutations in the candidate gene, and (2) recapitulation of key aspects of the proband's phenotype in the mouse by targeted mutation or knockdown of the candidate gene. The endpoint for these analyses is to obtain conclusive evidence that the gene disrupted by chromosomal rearrangement in any given case is responsible for the associated birth defects, and to conduct an initial analysis of the gene's developmental function. To accomplish these goals, we will employ a robust accrual system to recruit cases of interest from both national outreach and from major commercial cytogenetic providers. To take maximal advantage of the expertise that already exists within our laboratory, we will restrict our scope to two cases per year (ten overall) and we will focus specifically upon cases that affect organogenesis of the eye, kidney and craniofacial complex. In Aim 1, we will ascertain cases involving balanced chromosomal rearrangements (i.e., translocations and inversions) that are associated with disorders of craniofacial, ocular or renal development, and we will map the rearrangement breakpoints. In Aim 2, we will identify candidate genes disrupted by the breakpoints and determine the relevant mutational mechanisms. In Aim 3, to establish causality for these candidate genes, we will perform mutational analyses of phenotypically related human cases, and in Aim 4 we will further seek to establish causality by using genetic means to recapitulate the phenotype in mouse, and we will also determine the gene's developmental function. This powerful gene discovery approach will identify human genes that are newly linked to medically important birth defects, and help to define their developmental functions. PUBLIC HEALTH REVELANCE: This grant proposes to identify genes that are broken by naturally occurring human chromosome abnormalities associated with congenital birth defects. We will focus on birth defects involving the craniofacial region (e.g., cleft lip and palate), the eye (e.g., congenital cataracts), and the kidney (e.g., vesicoureteral reflux, or VUR). To prove that the broken genes are causally linked to the accompanying birth defect, we will search for mutations in the same gene in other patients with the same birth defect, and we will investigate the consequences of mutating the gene in mice.