Genetically testing mechanisms of ocular development and disease

描述(由申请人提供)：IV型胶原α1基因(Col4a1)突变会导致小鼠高度穿透性眼前段发育不全(ASD)和眼压升高。最近，在一个患有ASD、高眼压和青少年青光眼的家系中发现了COL4A1突变，但COL4A1并未被广泛认为是ASD的候选基因。在这项建议中，我们确定了COL4A1和COL4A2对人类ASD的贡献，并将利用新的遗传资源来确定Col4a 1诱导的ASD的细胞和分子机制。COL4A1是一种细胞外基质分子，存在于所有眼基底膜中。为了了解致病侮辱发生的地点和时间，我们开发了COL4a1的有条件表达的突变等位基因。在目标1中，我们将以空间或时间受限的方式表达突变等位基因，以定义这些参数并确定发病的主要部位。为了了解Col4a1突变导致眼发育不全的机制，并开始组装发育和致病途径，我们将识别ASD的遗传修饰物。我们已经成功地定位了一个能够有力拯救ASD的基因座。在目标2中，我们将对两个不同的遗传背景进行系统的大规模筛选，以确定额外的显性或隐性修饰基因座和基因。了解基因修饰物如何拯救疾病可以为有针对性的治疗干预如何起到同样的作用提供有价值的见解。重要的是，在一个患有自闭症、高眼压和青少年青光眼的家庭中发现了COL4A1突变。我们假设COL4A1及其结合伙伴COL4A2的突变是尚未发现突变的ASD患者的基础。在目标3中，我们将通过对已知没有其他ASD致病基因突变的ASD患者进行突变分析，直接检验我们的假设。据我们所知，目前还没有其他小组评估COL4A1或COL4A2在眼睛发育和疾病中的作用。这项建议中概述的实验利用了宝贵和独特的资源，并将为正常眼睛发育和致病途径的机制提供重要的新见解。

DESCRIPTION (provided by applicant): Mutations of the type IV collagen alpha 1 gene (Col4a1) cause highly penetrant ocular anterior segment dysgenesis (ASD) and elevated intraocular pressure in mice. Recently, a COL4A1 mutation was identified in a family with ASD, ocular hypertension and juvenile glaucoma, however, COL4A1, has not widely been considered a candidate gene for ASD. In this proposal we identify the contribution of COL4A1 and COL4A2 to human ASD and will use novel genetic resources to determine the cellular and molecular mechanisms of Col4a 1-induced ASD. COL4A1 is an extracellular matrix molecule that is present in all ocular basement membranes. To understand where and when the pathogenic insult occurs we have developed a conditionally expressed mutant allele of Col4a1. In Aim 1 we will express the mutant allele in a spatially or temporally restricted manner to define these parameters and determine the primary site of pathogenesis. To understand the mechanism by which Col4a1 mutations lead to ocular dysgenesis and to begin to assemble developmental and pathogenic pathways, we will identify genetic modifiers of ASD. We have already successfully mapped a locus that is able to strongly rescue ASD. In Aim 2, we will perform systematic large-scale screens of two distinct genetic backgrounds to identify additional dominant or recessive modifier loci and genes. Understanding how genetic modifiers rescue disease could provide valuable insight for how targeted therapeutic interventions might do the same. Importantly, a COL4A1 mutation was identified in a family with ASD, ocular hypertension and juvenile glaucoma. We hypothesize that mutations of COL4A1 and its binding partner COL4A2 underlie ASD in patients for whom mutations have not yet been identified. In Aim 3, we will test our hypothesis directly by performing mutational analysis on ASD patients that are known not to have mutations in other ASD-causing genes. To our knowledge, no other group is currently evaluating the role of COL4A1 or COL4A2 in ocular development and disease. The experiments outlined in this proposal take advantage of valuable and unique resources and will provide important new insights into the mechanisms of normal ocular development and pathogenic pathways.