Multiple ancestral duplications of the red-sensitive opsin gene (LWS) in teleost fishes and convergent spectral shifts to green vision in gobies

Photopigments, formed by an opsin protein bound to a light-sensitive chromophore, underlie vertebrate vision. Long-wavelength-sensitive (LWS) opsins mediate red-light detection, and most teleosts retain a single functional LWS1. A shorter-shifted green-sensitive paralog (LWS2) is found only in a few lineages. By mining teleost genomes and sequencing retinal transcriptomes, we identify elopomorphs as an additional lineage retaining LWS2 (alongside characins and osteoglossomorphs), and we reveal a previously overlooked shorter-shifted paralog, LWS3, restricted to gobies (Percomorpha) and arising from an ancient duplication. Structural modeling of twelve LWS opsins reveals convergent evolution at four key amino acid sites (214, 259, 261, 269) in the retinal-binding pocket, indicating convergent substitutions in human MWS, teleost LWS2, and goby LWS3 relative to red-sensitive counterparts, consistent with repeated spectral shifts toward green wavelengths. In several lineages--including characins, mormyrids, gobies, and primates--these shorter-shifted LWS opsins have functionally replaced the canonical green-sensitive RH2 opsin. Retinal transcriptomes and in situ hybridization demonstrate variable lws3 expression across gobies, with localization to a distinct double-cone member in Amblygobius phalaena, analogous to rh2 expression in other fish species. Together, these results show that repeated convergent evolution toward green sensitivity over 500 million years involves coordinated changes at the molecular, regulatory, and functional levels, providing a striking example of multilevel sensory adaptation. Significance statementVertebrate vision depends on opsins, which detect specific wavelengths of light. Most teleosts retain a single long-wavelength-sensitive opsin (LWS1), while a green-sensitive paralog (LWS2) occurs only in a few lineages. We identify a previously overlooked green-shifted opsin, LWS3, which is restricted to gobies, and show that LWS2 and LWS3 have repeatedly evolved green sensitivity through convergent amino acid changes. In parallel to the primate MWS/LWS evolution, these shorter-shifted opsins have functionally replaced the canonical green-sensitive RH2 opsin in multiple teleost lineages. By integrating genomic, structural, and expression data, we reveal how multilevel convergent evolution--from molecular tuning to photoreceptor specialization--has repeatedly shaped green-light vision over the past 500 million years of evolution, illustrating the remarkable flexibility of vertebrate visual systems.