Characterization of RGC death susceptibility alleles

描述（由申请人提供）：青光眼是一种慢性致盲性神经退行性疾病，其特征为视网膜神经节细胞的进行性丧失和视神经变性。虽然在这种疾病的遗传学领域已经取得了重大进展，但迄今为止，大多数这些研究都检查了表现出孟德尔遗传模式的罕见形式的疾病。然而，大多数恶性肿瘤是复杂的遗传疾病，具有影响个体易感性的多个相互作用的基因座。一个可能的易感领域是由死亡的神经节细胞执行的遗传控制的细胞死亡程序。我们在小鼠中使用实验遗传学来帮助识别可能影响这一过程的潜在易感等位基因。对15个近交系小鼠进行视神经挤压后神经节细胞损失量的筛选，发现2个品系对该程序具有不同的抗性。抗性表型被发现作为显性性状遗传，并且全基因组作图已经鉴定出负责基因位于染色体5的25 cM间隔内（Chr5.loc34-59）。该基因座被命名为视网膜神经节细胞易感1（Rgcs 1）。我们建议通过继续精细绘制Rgcs 1基因座来缩小感兴趣的区域来扩展这些观察结果。精细作图将开始，使用SNP分析将区间细化为预测的10 cM区域，然后生成区间特异性同源系（ISCL），将该区域分割为1 cM重叠区间。此外，我们将使用计算机绘图来帮助定义区间内的候选基因，因为它变得更小。迄今为止，我们已经使用这种方法从已知存在于该区域的578个基因中鉴定出7个潜在候选基因。候选基因将通过两个亲本近交系之间的定量和定性分析来表征。在产生ISCL的过程中，我们还将建立DBA/2 J小鼠的亚系，以检查Rgcs 1基因座在青光眼易感性中所起的作用。野生型DBA/2 J小鼠携带抗性等位基因。有趣的是，这些小鼠也患有慢性遗传性青光眼，这似乎是一个悖论，但也可能与真正的易感性等位基因一致。我们对Rgcs 1在青光眼中的作用的测试是将来自BALB/cByJ动物的易感Rgcs 1基因座与DBA/2 J背景杂交以产生DBA/2J.BALBRgcs 1亚株。我们预计这些小鼠会出现眼内压升高，并且由于携带易感的Rgcs 1等位基因，会出现更严重的青光眼。最后，当我们获得更多关于小鼠Rgcs 1基因座的信息时，我们将开始通过使用杜克大学人类遗传学中心的人类青光眼患者数据集检查关键候选基因中的SNP差异来询问人类基因组的同线区域。本提案描述了旨在鉴定和表征位于小鼠5号染色体区域的基因的研究，该基因影响视神经挤压后视网膜神经节细胞的存活。我们还将测试该染色体区域影响青光眼小鼠模型中神经节细胞存活的能力，然后扩展这些观察结果以确定相应的基因座/基因是否影响人类的青光眼。这项工作的意义在于，它将有助于阐明与青光眼复杂性状相关的一些易感等位基因。发现和表征这些等位基因将导致更好的诊断和治疗青光眼的个人制度。

DESCRIPTION (provided by applicant): Glaucoma is a chronic blinding neurodegenerative disease characterized by the progressive loss of retinal ganglion cells and degeneration of the optic nerve. Although significant progress has been made in the field of the genetics of this disease, the majority of these studies have so far examined rarer forms of the disease that exhibit Mendelian inheritance patterns. The majority of glaucomas, however, are complex genetic diseases that have multiple interacting loci that affect an individual's susceptibility. One possible area of susceptibility is the genetically controlled cell death program that is executed by dying ganglion cells. We have used experimental genetics in mice to help identify potential susceptibility alleles that could affect this process. A screen of 15 inbred mouse lines for the amount of ganglion cell loss after optic nerve crush revealed 2 lines with varying resistance to this procedure. The resistant phenotype was found to be inherited as a dominant trait, and genome wide mapping has identified the responsible gene to be located within a 25 cM interval of chromosome 5 (Chr5.loc34-59). This locus has been named Retinal ganglion cell susceptible 1 (Rgcs1). We propose to extend these observations by continuing to fine map the Rgcs1 locus to narrow the region of interest. Fine mapping will begin using SNP analysis to refine the interval to a predicted 10 cM region, followed by the generation of Interval Specific Congenic Lines (ISCLs) to dissect the region into 1 cM overlapping intervals. In addition, we will use in silico mapping to help define candidate genes of interest in the interval as it becomes smaller. To date, we have used this approach to identify 7 potential candidates from among the 578 genes known to exist in this region. Candidate genes will be characterized by quantitative and qualitative analyses between the two parental inbred strains. During the course of generating ISCLs, we will also create a substrain of DBA/2J mice to examine the role that the Rgcs1 locus plays in susceptibility to glaucoma. Wild type DBA/2J mice carry the resistant allele. Interestingly, these mice also develop chronic inherited glaucoma, which may seem like a paradox, but could also be consistent with a true susceptibility allele. Our test of the role of Rgcs1 in glaucoma, will be to cross the susceptible Rgcs1 locus from BALB/cByJ animals onto the DBA/2J background to generate the substrain DBA/2J.BALBRgcs1. We expect these mice to develop elevated intraocular pressure, and by virtue of carrying a susceptible Rgcs1 allele, a more severe form of glaucoma. Lastly, as we acquire more information on the mouse Rgcs1 locus, we will begin to interrogate the syntenic region of the human genome by examining for SNP differences in key candidate genes using a data set of human glaucoma patients housed at the Center for Human Genetics at Duke University. PUBLIC HEALTH RELEVANCE This proposal describes studies designed to identify and characterize a gene located in a region of mouse chromosome 5 that affects retinal ganglion cell survival after optic nerve crush. We will also test the ability of this chromosomal region to affect ganglion cell survival in a mouse model of glaucoma, and then extend these observations to determine if the corresponding locus/gene affects glaucoma in humans. The significance of this work is that it will help elucidate some of the susceptibility alleles associated with the complex trait of glaucoma. Finding and characterizing these alleles will lead to better diagnosis and treatment regimes for individuals with glaucoma.