Molecular mechanisms of anterior segment disorders

项目总结 眼前段发育不全(ASD)表型包括一组以结构性疾病为特征的疾病 涉及虹膜、角膜、虹膜角膜角和晶状体的异常，并与高视力风险相关 青光眼、角膜混浊和其他并发症造成的损害。这组条件包括 阿森菲尔德-里格畸形(ARA)和综合征(ARS)，无虹膜，彼得斯畸形/角膜混浊，以及 儿童青光眼(即使在没有可见的ASD的情况下，据信与发育异常有关 流出道)。我们确定PITX2的突变是ARS的原因，然后扩展了相关 其他ASD的表型；第二个ARS基因FOXC1随后也被证明解释了不同的 ASD的表型和最近的一项研究表明，它与多个人群的POAG有关。PITX2和FOXC1扮演着 在ASD中起主要作用，可解释40%-60%的ARA/ARS，而大多数阴性病例仍未解释。 对于更广泛的自闭症谱系，识别基因诊断的成功率因人群而异，但 同样，仍然不完整(不到50%)，需要确定新的机制。这个 该项目的总体目标是通过以下方式揭示ASD和相关的青光眼/视力损害的机制 研究这种疾病的不同方面：人类表型，与PITX2/FOXC1相关的机制， 以及通过外显子组/基因组人类研究和斑马鱼CRISPR-Cas9编辑发现新的基因。 具体地说，我们的目标是：1)揭示‘眼前段发育不全’的各种眼部异常 精神错乱。ASD患者的高分辨率成像将调查后节段的可能性 缺陷及其对视力丧失的贡献。这一目标将为人们提供对人类疾病的新见解。 通过我们的初始数据，在两个(两个中的)ARS患者中识别出新的特征；2)定义发育 FOXC1和PITX2的作用和途径，并探讨它们在人类疾病中的作用。这一目标将 利用最新开发的品系识别斑马鱼foxc1的靶标并探索其调控元件 吉恩。我们的一个foxc1调节缺失突变体的初步结果显示青光眼很严重。 表型，不同于功能丧失系，很可能成为斑马鱼的第一个胚胎 青光眼突变体。所获得的结果将与先前生成的PITX2数据一起分析以 揭示共同的路径。因为我们早期的数据显示WNT通路的失调是一个主要的 Pitx2缺乏的后果，它在眼睛表型中的作用将被进一步检测；最后，3)发现 利用家系外显子组/基因组数据研究与前段发育不全相关的新因子 斑马鱼受自闭症和基因组编辑影响的建模。将对已确定的因素进行评估 利用我们之前开发的资源，它们在已知路径中的位置。我们应用程序的优势在于 在其创新的方法中，我们产生了独特的资源，强大的方法，以及杰出的 我们团队在脊椎动物遗传学、眼睛发育和自闭症表型方面的专业知识。

PROJECT SUMMARY Anterior segment dysgenesis (ASD) phenotypes comprise a group of disorders characterized by structural abnormalities involving the iris, cornea, iridocorneal angle, and lens, and associated with a high risk of visual impairment through glaucoma, corneal opacification and other complications. This group of conditions includes Axenfeld-Rieger anomaly (ARA) and syndrome (ARS), aniridia, Peters anomaly/corneal opacities, and pediatric glaucoma (even in the absence of visible ASD, believed to be associated with abnormal development of the outflow tract). We identified mutations in PITX2 as the cause of ARS and then expanded associated phenotypes to other ASDs; the second ARS gene, FOXC1, was also subsequently shown to explain diverse ASD phenotypes and a recent study implicates it in POAG in multiple populations. PITX2 and FOXC1 play a major role in ASD, explaining 40-60% of ARA/ARS with the majority of negative cases remaining unexplained. For the broader ASD spectrum, the success rate for identifying a genetic diagnosis varies in populations but, likewise, continues to be incomplete (less than 50%) necessitating the identification of novel mechanisms. The overall goal of this project is to uncover mechanisms of ASD and associated glaucoma/visual impairment by investigating diverse aspects of this disease: human phenotype, mechanisms associated with PITX2/FOXC1, and novel gene discovery through exome/genome human studies and CRISPR-Cas9 editing in zebrafish. Specifically, we aim: 1) To reveal the full range of ocular anomalies in ‘anterior segment dysgenesis’ disorders. High-resolution imaging in individuals with ASD will investigate the possibility of posterior segment defects and their contribution to vision loss. This aim will provide new insight into human disease as suggested by our initial data which identified novel features in two (out of two) ARS patients; 2) To define developmental roles and pathways of FOXC1 and PITX2 and explore their contribution to human disease. This aim will identify targets of foxc1 in zebrafish using recently developed lines and explore regulatory elements of this gene. Our preliminary results from one of the foxc1 regulatory deletion mutants show a strong glaucoma phenotype, different from the loss-of-function line, and is likely to become the first zebrafish embryonic glaucoma mutant. The obtained results will be analyzed together with the previously generated pitx2 data to reveal common pathways. Since our earlier data indicate dysregulation of the WNT pathway as a major outcome of pitx2 deficiency, its role in the eye phenotype will be further examined; and finally, 3) To discover novel factors with a role in anterior segment dysgenesis by utilizing exome/genome data from families affected with ASD and genome editing-based modeling in zebrafish. The identified factors will be evaluated for their position in known pathways using our previously developed resources. The strength of our application is in its innovative approaches, the unique resources that we generated, robust approach, and the outstanding expertise of our team in vertebrate genetics, eye development and ASD phenotypes.