UC Irvine

Opsins are ancient molecules that enable animal vision by coupling to a vitamin-derived chromophore to form light-sensitive photopigments. The primary drivers of evolutionary diversification in opsins are thought to be visual tasks related to spectral sensitivity and color vision. Typically, only a few opsin amino acid sites affect photopigment spectral sensitivity. We show that opsin genes of the North American butterfly Limenitis arthemis have diversified along a latitudinal cline, consistent with natural selection due to environmental factors. We sequenced single nucleotide (SNP) polymorphisms in the coding regions of the ultraviolet (UVRh), blue (BRh), and long-wavelength (LWRh) opsin genes from ten butterfly populations along the eastern United States and found that a majority of opsin SNPs showed significant clinal variation. Outlier detection and analysis of molecular variance indicated that many SNPs are under balancing selection and show significant population structure. This contrasts with what we found by analysing SNPs in the wingless and EF-1 alpha loci, and from neutral amplified fragment length polymorphisms, which show no evidence of significant locus-specific or genome-wide structure among populations. Using a combination of functional genetic and physiological approaches, including expression in cell culture, transgenic Drosophila, UV-visible spectroscopy, and optophysiology, we show that key BRh opsin SNPs that vary clinally have almost no effect on spectral sensitivity. Our results suggest that opsin diversification in this butterfly is more consistent with natural selection unrelated to spectral tuning. Some of the clinally varying SNPs may instead play a role in regulating opsin gene expression levels or the thermostability of the opsin protein. Lastly, we discuss the possibility that insect opsins might have important, yet-to-be elucidated, adaptive functions in mediating animal responses to abiotic factors, such as temperature or photoperiod.