Analyses of visual pigments and G proteins coupled with them in light-sensitive chromatophores of teleost fishes

1.在尼罗罗非鱼红细胞中，色素聚集是由400和600 nm的光引起的，而分散是响应于500 nm的光而发生的。我们发现红色、绿色和蓝色视锥蛋白的mRNA在色素细胞中表达。此外，存在于侧条纹皮肤中的氖四虹膜细胞也是光敏的，并且500 nm附近的光是最有效的。RT-PCR结果显示，在虹彩细胞存在的部位，视紫红质和两种视锥细胞视蛋白（Pi-绿色1和Pi绿色2）的mRNA均有表达。这些mRNA也在侧眼中表达。锥视蛋白基因Pi-green 1和Pi-green 2与墨西哥盲洞鱼特有的绿色锥视蛋白（属于MWS/LWS组）的g101和g102基因具有很高的相似性，氨基酸序列的同源性在90%以上，红视蛋白基因在河豚皮肤中不表达.利用抗视蛋白（MWS/LWS）和抗视紫红质抗体，免疫印迹分析也证实了视蛋白在虹膜细胞存在的皮肤中的存在，并通过免疫组织化学分析证实了视蛋白在虹膜细胞中的定位.使用参与转导的酶的各种抑制剂和激活剂表明环AMP参与光转导。也就是说，光刺激伴随着光敏色素细胞中cAMP水平的降低。用细菌毒素处理尼罗罗非鱼红细胞和虹彩霓虹细胞也表明Gi在光转导中的重要作用.我们还分别用RT-PCR和Western blotting证实了Gi mRNA和Gi蛋白在四虹彩载体中的表达。虽然Gt mRNA的部分表达被证实，Gt蛋白没有检测到氖虹彩载体的四。

1. In Nile tilapia erythrophores, pigment aggregation is caused by light at 400 and 600 nm and dispersion occurred in response to light at 500 nm. We found the expression of mRNAs of cone opsins, red, green and blue, in the chromatophores. Further neon tetra iridophores present in the lateral stripe skin are also light-sensitive, and light near 500 nm is most effective. RT-PCR showed the expression of mRNAs encoding rhodopsin and two kinds of cone opsins (Pi-green 1 and Pi green 2) in neon tetra skin where the iridophores exist. These mRNAs are also expressed in the lateral eyes. The cone opsin genes, Pi-green 1 and Pi-green 2, show high similarity with the g101 and g102 genes of unique green cone opsins (belonging to the MWS/LWS group) of the blind Mexican cavefish The identity of amino acid sequences between Pi-green 1 and Pi-green 2 is considered to be more than 90% The red opsin gene was not expressed in neon tetra skin.2. Using anti-opsin (MWS/LWS) and anti-rhodopsin antibodies, western blotting analysis also demonstrated the presence of opsins in the skin where iridophores exist, and the localization of the opsins in iridophores was confirmed by immunohistochemical analysis.3. The involvement of cyclic AMP in the phototransduction was indicated using various inhibitors and activators of enzymes participating in the transduction. That is, light stimulation was accompanied by the decrease in cAMP level in the light-sensitive chromatophores. The treatment of Nile tilapia etythrophores and neon tetra iridophores with bacteria toxins also suggested the important role of Gi in the phototransduction.4. We also confirmed the expression of Gi mRNA and Gi protein in neon tetra iridophores by RT-PCR and western blotting respectively. Although partial expression of Gt mRNA was confirmed, Gt protein was not detected in the iridophores of neon tetra.