Cyp27c1 red-shifts the spectral sensitivity of photoreceptors by converting vitamin A1 into A2. Cyp27c1 red-shifts the spectral sensitivity of photoreceptors by converting vitamin A1 into A2

The goal of this project was to identify the dehydrogenase enzyme that converts vitamin A1 into A2, resulting in red-shifted spectral sensitivity of the visual system. We conducted next-generation sequencing of mRNA derived from the retinal pigment epithelium (RPE) of two animal models that use vitamin A2 as a chromophore to identify this enzyme. Zebrafish switch from vitamin A1 to A2 when treated with thyroid hormone, so we profiled the RPE from TH- and vehicle-treated zebrafish (n=3). The adult American bullfrog sequesters vitamin A2 within the dorsal part of the retina, but lacks vitamin A2 in the ventral retina. We therefore also profiled dorsal and ventral American bullfrog RPE (n=3). Using this approach, we identified cytochrome p450 family member cyp27c1 as strongly enriched in both datasets, and determined through additional experiments that cyp27c1 is both necessary and sufficient for vitamin A2 production. Overall design: Differential gene expression analysis in TH-treated vs. control zebrafish RPE and dorsal vs. ventral bullfrog RPE, conducted in triplicate

本项目旨在鉴定可将维生素A1转化为维生素A2的脱氢酶（dehydrogenase enzyme），该酶可赋予视觉系统光谱红移的敏感性。我们对两种以维生素A2作为生色团（chromophore）的动物模型的视网膜色素上皮（retinal pigment epithelium, RPE）来源的信使RNA（messenger RNA, mRNA）开展下一代测序（next-generation sequencing），以鉴定该目标酶。斑马鱼经甲状腺激素（thyroid hormone, TH）处理后，会从利用维生素A1转换为利用维生素A2；因此我们对经TH处理与载体处理的斑马鱼RPE进行了基因表达谱分析，每组设置3次生物学重复。成年美洲牛蛙会将维生素A2储留于视网膜背侧区域，但腹侧视网膜中缺乏维生素A2；因此我们同样对美洲牛蛙的背侧与腹侧RPE进行了基因表达谱分析，每组设置3次生物学重复。通过该分析策略，我们在两个数据集中共发现细胞色素P450家族成员cyp27c1存在显著富集；后续通过额外实验证实，cyp27c1对于维生素A2的生成既是必需的，也是充分的。整体实验设计如下：对经TH处理与对照组的斑马鱼RPE、以及背侧与腹侧的美洲牛蛙RPE开展差异基因表达分析，所有实验均设置3次生物学重复。