Modifiers of Retinal Phenotypes in Ciliopathies

描述(申请人提供)：影响纤毛结构和功能的遗传损伤会引起广泛的遗传异质性和临床重叠疾病，统称为纤毛病变，其特征是表型重叠和外显率和表现力不同。在视网膜中，修饰的纤毛在光感受器的蛋白质运输中起着不可或缺的作用，对视网膜的结构和功能至关重要，这一事实证明了进行性的光感受器退化是许多纤毛疾病的标志。越来越多的证据表明，在一些纤毛疾病中突变的基因可以在整个纤毛病谱中贡献原因等位基因和修改等位基因，从而产生这样的想法，即顺式和反式等位基因都可以促进纤毛疾病患者的突变负荷，并提供了这样一种可能性，即了解纤毛疾病的遗传结构可能会为人类遗传疾病表型变异的机制提供信息。为了探索这一概念，我们之前已经对已知/预计对纤毛生物发生和功能重要的基因进行了无偏见的医学重新测序，这些基因在一大批临床不同的患者中跨越了严重程度的光谱。在RPGRIP1L中，我们发现了一个高度保守的A229T突变，该突变存在于中等人群频率，在视网膜变性患者中显著丰富。RPGRIP1L是一种已知导致新生儿致命的Meckel-Gruber综合征(MKS)和中度重度Joubert综合征(JBTS)的基因。使用跨学科的方法，我们进一步证明Thr229等位基因是一种非中性的改变，它扰乱了RPGRIP1L和RPGR之间的直接相互作用，RPGR是X-连锁视网膜色素变性(XLRP)最常见的遗传原因。这些数据为我们提供了探索纤毛疾病视网膜表型第二位点修饰的遗传机制(S)的机会，并开发了可用于进一步探索此类现象的模型。我们提出了两个目标。首先，由于有机会开发一个强大的模型来研究上位性，我们将通过将A229T引入小鼠模型来模拟A229T的变化，然后将Thr229等位基因杂交到具有敏化睫毛功能的系中，以确定该等位基因是否会诱导或加剧视网膜表型。其次，由于我们的初步数据表明RPGRIP1L也可能导致非综合征视网膜变性的上位性等位基因，我们将扩大RPGRIP1L的突变分析，以扩大非综合征患者和匹配对照的队列。使用我们之前建立的体内互补策略，然后我们将测试新发现的等位基因的致病潜力，并根据功能数据，我们将确定RPGRIP1L等位基因在视网膜变性中的总体浓缩。我们研究的完成将确定视网膜变性患者的候选修饰等位基因，产生研究此类现象的新模型，并有可能揭示表型变异的遗传基础，这反过来将有助于更好地诊断和长期管理患者。 公共卫生相关性：睫状蛋白功能障碍引起的视网膜变性是早发性和成年性失明的常见原因，在受影响的个体中观察到的临床变异性对预后和治疗构成了重大挑战。然而，最近对纤毛蛋白质组的表征和体内工具的发展为识别和模拟这些疾病中外显性和表达能力的修饰物提供了独特的机会，反过来，这又具有增强基因预测能力的潜力。

DESCRIPTION (provided by applicant): Genetic lesions affecting ciliary structure and function give rise to a broad collection of genetically heterogeneous and clinically overlapping disorders, known collectively as the ciliopathies, which are characterized by both phenotypic overlap and variable penetrance and expressivity. In the retina, a modified cilium plays an integral role in protein transport across the photoreceptor and is critical for retinal architecture and function, as evidenced by the fact that progressive photoreceptor degeneration is a hallmark of numerous ciliopathies. Accumulating evidence suggests that genes mutated in some ciliopathies can contribute both causal and modifying alleles across the ciliopathy spectrum, giving rise to the idea that both cis and trans acting alleles can contribute to the mutational load of ciliopathy patients and offer the possibility that understanding the genetic architecture of ciliopathies might inform the mechanisms that underlie phenotypic variability in human genetic disorders. To explore this notion, we have previously conducted unbiased medical resequencing of genes known/expected to be important to ciliary biogenesis and function in a large, clinically diverse cohort of patients that span the spectrum of severity. In RPGRIP1L, a gene known to cause neonatal lethal Meckel-Gruber Syndrome (MKS) and moderately severe Joubert Syndrome (JBTS), we identified a highly-conserved A229T change which was present at intermediate population frequency, and was significantly enriched in patients with retinal degeneration. Using an interdisciplinary approach, we went on to show that the Thr229 allele is a non-neutral change that disrupts the direct interaction between RPGRIP1L and RPGR, the most frequent genetic cause of X-linked Retinitis Pigmentosa (XLRP). These data offer us the opportunity explore the genetic mechanism(s) of second-site modification in retinal phenotypes in ciliopathies, and to develop models that can be used to probe such phenomena further. We propose two aims. First, motivated by the opportunity to develop a robust model to study epistasis, we will model the A229T change by introducing it into a mouse model and subsequently crossing the Thr229 allele into lines with sensitized ciliary function to determine if this allele will either induce or exacerbate retinal phenotypes. Second, because our preliminary data suggest that RPGRIP1L might also contribute epistatic alleles to non-syndromic retinal degeneration, we will expand the mutational analysis of RPGRIP1L to an extended cohort of non-syndromic patients and matched controls. Using our previously established in vivo complementation strategy, we will then test the pathogenic potential of newly discovered alleles, and, empowered with functional data, we will determine the overall enrichment of RPGRIP1L alleles in retinal degeneration. The completion of our studies will identify candidate modifier alleles in patients with retinal degeneration, generate new models to study such phenomena and has the potential to inform the genetic basis of phenotypic variability, which in turn will contribute to the better diagnosis and long-term management of patients. PUBLIC HEALTH RELEVANCE: Retinal degeneration caused by dysfunction of ciliary proteins represents a frequent cause of both early onset and adult blindness, and the observed clinical variability among affected individuals poses a significant challenge in terms of prognosis and treatment. However, recent characterization of the ciliary proteome and the development of in vivo tools pose a unique opportunity to identify and model modifiers of both penetrance and expressivity in these disorders which in turn, has the potential of enhancing the predictive power of the genotype.