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Gene duplication of green (RH2) opsin genes and their spectral differentiation are well documented in many teleost fish. However, their evolutionary divergence or conservation patterns among phylogenetically close but ecologically diverse species is not well explored. Medaka fish (genus Oryzias) are broadly distributed in fresh and brackish waters of Asia, with many species being laboratory-housed and feasible for genetic studies. We previously showed that a Japan strain (HNI) of medaka (Oryzias latipes) possessed three RH2 opsin genes (RH2-A, RH2-B, and RH2-C) encoding spectrally divergent photopigments. Here, we examined the three RH2 opsin genes from six Oryzias species representing three species groups: the latipes, the celebensis, and the javanicus. Photopigment reconstitution revealed that the peak absorption spectra (λmax) of RH2-A were divergent among the species (447–469 nm), whereas those of RH2-B and RH2-C were conservative (516–519 and 486–493 nm, respectively). For the RH2-A opsins, the largest spectral shift was detected in the phylogenetic branch leading to the latipes group. A single amino acid replacement T94C explained most of the spectral shift. For RH2-B and -C opsins, we detected tracts of gene conversion between the two genes homogenizing them. Nevertheless, several amino acid differences were maintained. We showed that the spectral difference between the two opsins was attributed to largely the E/Q amino acid difference at the site 122 and to several sites with individually small spectral effects. These results depict dynamism of spectral divergence of orthologous and paralogous green opsin genes in phylogenetically close but ecologically diverse species exemplified by medaka.

Orthologous Divergence and Paralogous Anticonvergence in Molecular Evolution of Triplicated Green Opsin Genes in Medaka Fish, Genus Oryzias