The vertebrate ancestor possessed ultraviolet (UV) vision and many species have retained it during evolution. Many other species switched to violet vision and, then again, some avian species switched back to UV vision. These UV and violet vision are mediated by short wavelength-sensitive (SWS1) pigments that absorb light maximally ( However, many species, including humans, have switched from UV vision to violet vision even when they receive abundant UV light in their environments. Two major reasons are given for these changes (4): (i) the eye structures of many species have been modified so that UV light does not reach the retina, protecting their retinal tissues, and somehow violet vision evolved; and (ii) without UV vision, organisms can improve visual resolution and subtle contrast detection. However, a certain avian ancestor replaced violet vision by UV vision (5), and it is not clear why the avian ancestor abandoned such evolutionary achievements associated with violet vision.To understand possible selective advantages of the evolutionary switches between UV and violet vision (5), we must identify critical amino acid changes that cause such functional changes and relate UV and violet vision of organisms to their ecological environments and life styles (6, 7). Living in diverse and reasonably well-defined light environments, fish offer the best opportunity to elucidate the molecular bases for the spectral tuning and adaptive evolution of SWS1 pigments at the same time (8). However, to achieve these goals, we are faced with two major problems. First, certain amino acid changes at 13 sites are known to shift the max s of SWS1 pigments to date (5, 9-14), but it is a daunting task to identify other critical amino acid changes because their effects on the max -shift are often not detectible when they are studied individually (9,15,16). Second, and perhaps more disturbingly, no violet-sensitive SWS1 pigment has been isolated from fish (17). Having no violet-sensitive SWS1 pigment in fish, we cannot relate UV and violet vision to organisms' ecological environments. Hence, to elucidate the evolutionary mechanisms of UV and violet vision, it is of utmost importance to isolate violet-sensitive SWS1 pigments from fish.
ResultsScabbardfish and Lampfish SWS1 Opsins and Visual Pigments. Juvenile scabbardfish (Lepidopus fitchi) live at depths of 20-50 m, but adult fish move to the depth of 100-500 m (www.fishbase.org). In contrast, lampfish (Stenobrachius leucopsarus) can be active even at a depth of 3,000 m but rapidly ascend to near the surface after sunset and feed on zooplankton (www.fishbase.org). We isolated the complete SWS1 opsin genes from these species by using PCR amplification and inverse PCR methods. The cloning results show that the scabbardfish and lampfish genes consist of 336 and 338 codons, respectively (Fig. 1). Compared with the SWS1 pigment in the vertebrate ancestor (pigment a) (5), these genes are missing several codons at the 5Ј-and 3Ј-ends (Fig. 1). By using the in vitro assay (18), we found that ...