Molecular Genetics Laboratory, Institute for Ophthalmic Research, University of Tübingen, Tübingen, Germany; Graduate School of Cellular and Molecular Neuroscience, University of Tübingen, Tübingen, Germany.
Molecular Genetics Laboratory, Institute for Ophthalmic Research, University of Tübingen, Tübingen, Germany.
Exp Eye Res. 2018 Jun;171:48-53. doi: 10.1016/j.exer.2018.02.027. Epub 2018 Feb 27.
The cyclic nucleotide-gated (CNG) channel - composed of CNGA3 and CNGB3 subunits - mediates the influx of cations in cone photoreceptors after light stimulation and thus is a key element in cone phototransduction. Mutations in CNGA3 and CNGB3 are associated with achromatopsia, a rare autosomal recessive retinal disorder. Here, we demonstrate that the presence of an early nonsense mutation in CNGA3 induces the usage of a downstream alternative translation initiation site giving rise to a short CNGA3 isoform. The expression of this short isoform was verified by Western blot analysis and DAB staining of HEK293 cells and cone photoreceptor-like 661W cells expressing CNGA3-GST fusion constructs. Functionality of the short isoform was confirmed by a cellular calcium influx assay. Furthermore, patients carrying an early nonsense mutation were analyzed for residual cone photoreceptor function in order to identify a potential role of the short isoform to modify the clinical outcome in achromatopsia patients. Yet the results suggest that the short isoform is not able to compensate for the loss of the long isoform leaving the biological role of this variant unclear.
环核苷酸门控（CNG）通道由 CNGA3 和 CNGB3 亚基组成，介导光刺激后圆锥光感受器内阳离子的内流，因此是圆锥光感受器转导的关键元件。CNGA3 和 CNGB3 的突变与色盲症有关，这是一种罕见的常染色体隐性视网膜疾病。在这里，我们证明了 CNGA3 中早期无意义突变的存在诱导了下游替代翻译起始位点的使用，从而产生了短的 CNGA3 同工型。通过 Western blot 分析和表达 CNGA3-GST 融合构建体的 HEK293 细胞和圆锥光感受器样 661W 细胞的 DAB 染色证实了这种短同工型的表达。通过细胞钙内流测定证实了短同工型的功能。此外，还分析了携带早期无意义突变的患者的残余圆锥光感受器功能，以确定短同工型是否有可能改变色盲症患者的临床结果。然而，结果表明短同工型不能补偿长同工型的缺失，这使得该变体的生物学作用仍不清楚。
