Characterization of DHDDS RP59 Knockin Models of Retinitis Pigmentosa

项目概要/摘要 视网膜色素变性（RP）是一组异质性疾病，导致视网膜变性。到目前为止， 超过58个基因的突变与经典的视网膜色素变性有关，并且共同地与所有形式的视网膜色素变性有关。 遗传性视网膜营养不良涉及至少300个基因。我的研究重点是视网膜色素变性-59 （RP 59），一种非综合征性常染色体隐性形式的RP引起的突变，在脱氢dolichyl 二磷酸合酶（DHDDS）基因，定位于染色体1p36.11上的外显子3。最普遍的 突变为K42 E，T206 A和R98 W也有报道。我们在小鼠体内构建了一个Dhdds K42 E敲除基因， 在小鼠模型中使用CRISPR/Cas9基因编辑技术。初步数据显示， ERG b波振幅，我们已经报道了显著的GFAP上调，没有视网膜变性或 降低蛋白质N-糖基化。这表明缺陷性DHDDS依赖性视网膜变性可能是 比简单的由于蛋白质糖基化改变而导致的功能丧失机制更复杂。更好地 为了了解与RP 59突变相关的选择性视网膜病理学的基础，我们计划表征 DhddsK 42 E/K42 E基因敲入小鼠的解剖学和生理学。此外，我们将创建和分析两个新的 DHDDS相关的遗传性视网膜疾病模型，Dhdds T206 A/T206 A和Dhdds T206 A/K42 E。研究视网膜在 这些小鼠模型将使我们深入了解这种遗传性疾病的病理生理学，并可能导致新的 这种疾病的治疗选择。

Project Summary/Abstract Retinitis pigmentosa (RP) is a heterogeneous group of disorders that cause retinal degeneration. To date, mutations in more than 58 genes have been linked to classic retinitis pigmentosa, and collectively all forms of inherited retinal dystrophies involves at least 300 genes. My studies are focused on retinitis pigmentosa-59 (RP59), a non-syndromic autosomal recessive form of RP caused by mutations in the dehydrodolichyl diphosphate synthase (DHDDS) gene, localized to exon 3 on chromosome 1p36.11. The most prevalent mutation is K42E, while T206A and R98W also have been reported. We generated a murine Dhdds K42E knock- in mouse model using CRISPR/Cas9 gene editing technology. Preliminary data shows significant reduction of ERG b-wave amplitudes and we have reported marked GFAP up-regulation, without retinal degeneration or decreased protein N-glycosylation. This suggests that defective DHDDS-dependent retinal degeneration may be more complicated than simply a loss-of-function mechanism due to altered protein glycosylation. To better understand the basis for selective retinal pathology associated with RP59 mutations, we plan to characterize the DhddsK42E/K42E knock-in mouse anatomically and physiologically. Additionally, we will create and analyze two new models of DHDDS-related inherited retinal disease, Dhdds T206A/T206A and DhddsT206A/K42E. Studying the retina in these mouse models will give us insight into the pathophysiology of this inherited disorder, and may lead to novel treatment options for this disorder.