Genetic Modulation of Glaucoma

描述（由申请人提供）：在本提案中，我们使用我们的BXD重组自交系扩大集来确定参与调节青光眼严重程度的基因位点。我们的方法结合了彻底的临床和实验室检查，并对过去10年产生的81个BXD系进行了微阵列分析，目的是研究眼病和青光眼的遗传学。BXD亲本菌株之一DBA/2J可发展为年龄相关性青光眼，其发病前伴有虹膜萎缩和色素分散。虽然两种基因的突变等位基因Tryp1和Gpnmb可引起D2患者的虹膜病变，但文献强烈提示这些突变不足以引起青光眼。具体来说，这两个突变等位基因渗入到BXD的另一个亲本菌株C57BL/6J上，导致了对视神经损伤的显著抵抗。这表明除了引起色素分散的基因外，其他基因也影响青光眼的表型。目前的提议描述了一个独特的机会来定义修饰位点。在Aim 1中，我们验证了Tryp1和Gpnmb的联合突变不足以引起青光眼表型的所有方面的假设。如果我们的假设是正确的，我们期望看到疾病的严重程度不仅仅取决于这两个突变的等位基因。我们已经鉴定了多个BXD菌株，其中IOP和遗传二倍型不相关。当我们系统地检查所有BXD菌株并建立表型和二倍型之间的关系时，我们完全期望找到其他菌株，这些菌株违背了简单的双位点疾病模型的期望。在目标2中，我们定义了调节青光眼严重程度的位点和基因。为此，我们将鉴定和评估调节青光眼表型严重程度的基因座内的候选基因。我们将利用我们新的D2全基因组鸟枪序列（2009年在UTHSC生成的短读覆盖率约为50倍）以及我们也作为这项工作的前奏生成的大量全眼和全视网膜表达数据集。我们期望有效地提名和评估候选青光眼基因使用最先进的生物信息学方法和传统的分子分析。在Aim 3中，我们使用双向翻译。我们使用人类青光眼患者队列来测试Aim 2候选小鼠的翻译有效性。J. Wiggs博士及其同事将使用Aim 2中提名的候选青光眼基因进行重点基因关联研究。具体地说，我们使用关联分析的标记包括人类染色体的共染色体区域。在反向翻译中，我们（MMJ和LL）将评估来自临床队列的已知、新的和候选青光眼基因，并确定这些变异是否以及如何与bxd中青光眼相关性状相关。结合小鼠和人类青光眼研究中最优先的候选基因，我们将建立易感性候选基因的分子和统计模型，相关表型和相关机制。

DESCRIPTION (provided by applicant): In this proposal, we use our enlarged set of BXD recombinant inbred strains to identify gene loci that are involved in modulating the severity of glaucoma. Our approach combines a thorough clinical and laboratory examination, and microarray analysis of the entire set of 81 BXD lines generated over the last 10 years with the express purpose of studying the genetics of eye disease and glaucoma. One of the parental strains of BXD, DBA/2J, develop an age-related glaucoma that is preceded by iris atrophy and pigment dispersion. While mutant alleles of two genes, Tryp1 and Gpnmb, cause the iris disease in D2, the literature strongly suggests that these mutations are not sufficient to cause glaucoma. Specifically, introgression of both mutant alleles onto C57BL/6J, the other parent strain of BXD, results in a marked resistance to optic nerve damage. This indicates that genes other than those that cause pigment dispersion influence the glaucomatous phenotype. The current proposal describes a unique opportunity to define the modifying loci. In Aim 1, we test the hypothesis that the combined mutations in Tryp1 and Gpnmb are not sufficient to cause all aspects of the glaucoma phenotype. If our hypothesis is true, we expect to see that the severity of disease is not solely dependent on the two mutant alleles. We have already identified multiple BXD strains in which IOP and genetic diplotype are not correlated. As we systematically examine all BXD strains and establish relations between phenotype and diplotype, we fully expect to find additional strains that defy expectations of a simple two-locus disease model. In Aim 2, we define loci and genes that modulate glaucoma severity. To do so, we will identify and evaluate candidate genes within loci that modulate the severity of the glaucoma phenotype. We will exploit our new whole genome shotgun sequence for D2 (about >50x short read coverage generated at UTHSC in 2009) along with massive whole eye and whole retina expression datasets that we have also generated as a prelude to this work. We expect to efficiently nominate and evaluate candidate glaucoma genes using state-of-the-art bioinformatic methods and conventional molecular assays. In Aim 3, we use bidirectional translation. We test the translational validity of mouse candidates from Aim 2 using cohorts of human glaucoma patients. Dr. J. Wiggs and colleagues will perform focused gene association studies using candidate glaucoma genes nominated in Aim 2. Specifically, we use association analyses of markers encompassing syntenic regions of human chromosomes. In reciprocal reverse translation, we (MMJ and LL) will evaluate known, new, and candidate glaucoma genes from clinical cohorts and determine if and how these variants are associated with glaucoma- associated traits in BXDs. Combining the top priority gene candidates from both mouse and human glaucoma studies, we will generate molecular and statistical models of susceptibility candidate genes, linked phenotypes, and associated mechanisms. PUBLIC HEALTH RELEVANCE: Glaucoma is a highly prevalent group of diseases that, if uncontrolled, causes irreversible loss of vision. The underlying cause of the disease is not known in the majority of cases, therefore treatment options are limited to lowering the intraocular pressure. Outcomes of our proposed investigations will identify gene loci that modulate glaucoma severity to expand this sparse list of genes that are known to contribute to this disease.