UCD Conway Institute and UCD School of Biomolecular and Biomedical Sciences, University College Dublin, Dublin 4, Ireland.
Exp Eye Res. 2013 Mar;108:120-8. doi: 10.1016/j.exer.2013.01.003. Epub 2013 Jan 15.
Cone-rod dystrophy 6 (CORD6) is an inherited blindness that presents with defective cone photoreceptor function in childhood, followed by loss of rod function. CORD6 results from mutations in GUCY2D, the human gene encoding retinal guanylate cyclase 1 (RETGC-1). RETGC-1 functions in phototransduction, synthesising cGMP to open ion channels in photoreceptor outer segments. As there is limited histopathological data on the CORD6 retina, our goal was to generate a CORD6 model by expressing mutant human RETGC-1 in zebrafish cone photoreceptors and to investigate effects on retinal morphology and function. cDNAs encoding wildtype and mutant (E837D R838S) RETGC-1 were cloned under the control of the cone-specific gnat2 promoter and microinjected into zebrafish embryos to generate transgenic lines. RETGC-1 mRNA expression in zebrafish eyes was confirmed by RT-PCR. Fluorescent microscopy analysed retinal morphology and visual behaviour was quantified by the optokinetic response (OKR). Stable transgenic lines expressing mutant or wildtype human RETGC-1 in zebrafish eyes were generated. OKR assays of 5-day-old larvae did not uncover any deficits in visual behaviour. However, transgenic (E837D R838S) RETGC-1 expression results in aberrant cone morphology and a reduced cone density. A reduction in the number of photoreceptor nuclei, the thickness of the outer nuclear layer and the labelling of rod outer segments, particularly in the central retina, was evident. Expression of mutant human RETGC-1 leads to a retinal phenotype that includes aberrant photoreceptor morphology and a reduced number of photoreceptors. This phenotype likely explains the compromised visual function, characteristic of CORD6.
Cone-rod 营养不良 6 型（Cone-rod dystrophy 6，CORD6）是一种遗传性失明，其特征为儿童时期出现视锥细胞功能缺陷，随后视杆细胞功能丧失。CORD6 由 GUCY2D 基因突变引起，GUCY2D 是编码视网膜鸟苷酸环化酶 1（retinal guanylate cyclase 1，RETGC-1）的人类基因。RETGC-1 在光转导中发挥作用，合成 cGMP 以打开视锥细胞外段中的离子通道。由于 CORD6 视网膜的组织病理学数据有限，我们的目标是通过在斑马鱼视锥细胞中表达突变型人类 RETGC-1 来生成 CORD6 模型，并研究其对视网膜形态和功能的影响。野生型和突变型（E837D R838S）RETGC-1 的 cDNA 在 cone-specific gnat2 启动子的控制下克隆，并显微注射到斑马鱼胚胎中以生成转基因系。通过 RT-PCR 证实了斑马鱼眼睛中 RETGC-1 的 mRNA 表达。荧光显微镜分析了视网膜形态，通过视动反应（optokinetic response，OKR）量化了视觉行为。在斑马鱼眼睛中生成了稳定表达突变或野生型人类 RETGC-1 的转基因系。对 5 天大的幼虫进行 OKR 测定并未发现任何视觉行为缺陷。然而，转基因（E837D R838S）RETGC-1 的表达导致视锥细胞形态异常和视锥细胞密度降低。明显的是，视锥细胞核数量减少、外核层厚度减少以及视杆细胞外段的标记减少，尤其是在中央视网膜。表达突变型人类 RETGC-1 导致包括视锥细胞形态异常和视锥细胞数量减少在内的视网膜表型。这种表型可能解释了 CORD6 特征性的视觉功能受损。
