Department of Anatomy and Neuroscience, The University of Melbourne, Parkville, Victoria, Australia.
Invest Ophthalmol Vis Sci. 2013 May 9;54(5):3350-9. doi: 10.1167/iovs.13-11831.
This work investigates a novel, naturally occurring mouse model of achromatopsia. The specific missense mutation within the Gnat2 gene was identified and the subsequent retinal phenotype characterized.
The Gnat2 sequence was amplified using PCR from BALB/c and Gnat2(c.518A>G) retinae and the product sequenced. Retinal function was assessed at 3, 6, 9, and 12 months using the electroretinogram. Transducin and opsin expression were assessed at 3 and 12 months using immunohistochemistry and quantitative PCR. Retinal remodeling and Müller cell gliosis were investigated using immunocytochemistry.
An A to G missense mutation at position 518 of the Gnat2 gene was identified that resulted in an aspartic acid to glycine substitution. Gnat2(c.518A>G) animals showed no cone response, while the rod response was normal except for a decrease in the photoreceptor response at 12 months (a-wave, -14%). Gnat2(c.518A>G) retinal sections showed no transducin immunolabeling; however, protein was detected via Western blot. Gnat2 gene expression was only decreased at 12 months of age (-27%). There was reduced cone number at 12 months (-27%) and M-opsin showed evidence of mislocalization. Displaced photoreceptor terminals and altered horizontal cell, cone/rod bipolar cell morphology were evident at 3 months, becoming more extensive at 12 months with the emergence of Müller cell gliosis.
The Gnat2(c.518A>G) mouse contains a missense mutation that results in no cone function due to a misfolding of transducin. Cone photoreceptors also show signs of opsin mislocalization, retinal remodeling and degeneration. This naturally occurring model shows all the hallmark signs of achromatopsia.
本研究旨在探索一种新型、天然发生的色盲小鼠模型。鉴定了 Gnat2 基因中的特定错义突变，并对随后的视网膜表型进行了特征描述。
使用 PCR 从 BALB/c 和 Gnat2(c.518A>G) 视网膜中扩增 Gnat2 序列，并对产物进行测序。使用视网膜电图在 3、6、9 和 12 个月时评估视网膜功能。使用免疫组织化学和定量 PCR 在 3 和 12 个月时评估转导蛋白和视蛋白表达。使用免疫细胞化学研究视网膜重塑和 Müller 细胞胶质增生。
鉴定出 Gnat2 基因第 518 位的 A 到 G 错义突变，导致天冬氨酸突变为甘氨酸。Gnat2(c.518A>G)动物没有视锥反应，而视杆反应正常，除了在 12 个月时（a 波，-14%）感光器反应下降。Gnat2(c.518A>G)视网膜切片没有转导蛋白免疫标记；然而，通过 Western blot 检测到了蛋白。Gnat2 基因表达仅在 12 个月时下降（-27%）。12 个月时视锥数量减少（-27%），M-opsin 显示出定位错误的迹象。3 个月时已可见感光器末端移位和水平细胞、视锥/视杆双极细胞形态改变，12 个月时更明显，出现 Müller 细胞胶质增生。
Gnat2(c.518A>G)小鼠携带一个错义突变，导致转导蛋白错误折叠，视锥功能丧失。视锥感光器也表现出视蛋白定位错误、视网膜重塑和变性的迹象。这种天然发生的模型显示出色盲的所有标志性特征。
