Targeted knockdown of Cerkl, a retinal dystrophy gene, causes mild affectation of the retinal ganglion cell layer
Targeted knockdown of Cerkl, a retinal dystrophy gene, causes mild affectation of the retinal ganglion cell layer
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DOI：
10.1016/j.bbadis.2012.04.004
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发表时间：
2012-08-01
影响因子：
6.2
通讯作者：
Marfany, Gemma
中科院分区：
文献类型：
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作者：
Garanto, Alejandro;Vicente-Tejedor, Javier;Marfany, Gemma
为了探讨视网膜营养不良症CERKL基因的功能，我们通过cre介导靶向缺失Cerk1第一外显子和近端启动子的方法建立了一种新的基因敲除小鼠模型。切除的基因组区域(2.3kb)包含第一个Cerk1外显子，上游序列包括近端启动子和第一内含子的起始片段。Cerkl-/-小鼠是活的和有生育能力的。考虑到替代启动子(当时未报道)指导Cerk1的基础表达(35%)，靶向Cerk1的缺失导致了敲除比敲除模型更多的基因敲除。原位杂交和免疫组织化学结果显示，该残存蛋白在感光细胞中呈中度表达，在神经节和内细胞层表达较弱。即使在12个月龄时，Cerkl-/-视网膜的形态特征也没有显示出任何明显的结构变化。TUNEL检测到神经节细胞持续的非进行性扰动(从出生到12月龄)，Brn3a标记物表达的减少和视网膜电图记录的振荡电位的降低支持了神经节细胞持续的非进行性扰动。总之，Cerkl-/-基因敲除表现为轻微的视网膜表型，细胞应激和细胞凋亡指标水平增加，神经节细胞层有明显的功能改变迹象，但没有可检测到的形态变化。(C)2012爱思唯尔B.V.保留所有权利。
In order to approach the function of the retinal dystrophy CERKL gene we generated a novel knockout mouse model by cre-mediated targeted deletion of the Cerkl first exon and proximal promoter. The excised genomic region (2.3 kb) encompassed the first Cerkl exon, upstream sequences including the proximal promoter and the initial segment of the first intron. The Cerkl -/- mice were viable and fertile. The targeted Cerkl deletion resulted in a knockdown more than a knockout model, given that alternative promoters (unreported at that time) directed basal expression of Cerkl (35%). In situ hybridizations and immunohistochemistry showed that this remnant expression was moderate in the photoreceptors and weak in the ganglion and inner cell layers. Morphological characterization of the Cerkl -/- retinas did not show any gross structural changes, even at 12 months of age. However, some clear and consistent signals of gliosis and retinal stress were detected by the statistically significant increase of i) the glial fibrillary antigen protein (GFAP) expression, and ii) apoptosis, as detected by TUNEL Remarkably, consistent non-progressive perturbation (from birth up to 12 months of age) of ganglion cells was supported by the decrease of the Brn3a marker expression as well as the reduced oscillatory potentials in the electroretinographic recordings. In conclusion, the Cerkl -/- knockdown shows a mild retinal phenotype, with increased levels of cellular stress and apoptosis indicators, and clear signs of functional alteration at the ganglion cell layer, but no detectable morphological changes. (C) 2012 Elsevier B.V. All rights reserved.