Using fish, and FISH, to interrogate the vertebrate opsin repertoire

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The longterm aim of my research program is to advance our understanding of opsin gene function and of vertebrate light sensitivity.  I am especially interested in non-visual opsins and extra-ocular light sensitivity. We have known for sometime that opsin expression is not limited to eyes and that other tissues are light sensitive, but studies connecting these two observations are rare. In addition, no studies to date adequately explain opsin repertoire size or expression domain diversity. The ancestor of all vertebrates had at least 23 opsins, five visual opsins and 18 non-visual opsins.  These repertoires are larger than would seem necessary for colour vision and circadian rhythm. In addition, opsin expression patterns are puzzling; opsins are expressed in human skin, brain, lung, liver, heart, and kidney and zebrafish express opsins in gills, heart, brain and testes. Visual and non-visual opsins are also expressed in zebrafish embryos well before eyes develop. The research proposed here includes the study of repertoire and expression variation in three fish species adapted to peculiar light exposures. Genome sequences, in-situ hybridization and digital PCR data will be collected and analyzed.  I will work on three distinct projects. 1) Genome sequencing of Phreatobius cisternarum, a troglomorphic, subterranean catfish from the Amazonian region of Brazil.  The goal of this project is to characterize the opsin repertoire and thereby identify opsins retained in a species adapted to life in perpetual darkness. Genome sequences will also be used to study the molecular basis of troglomorphism in general, and will be used to quantify population structure providing insight into the connectedness of subterranean aquatic habits in this region of Brazil, 2) We will quantify the levels of opsin expression during Anoplopoma fimbria (sablefish) development. This species has at least 27 opsins, but lives below the photic zone (i.e., at a depth of 200 m to 2,000 m). We predict that this large opsin repertoire is utilized primarily during development. We will use digital PCR to quantify expression in i) adults caught at depth and ii) a diversity of developmental stages (starting with fertilized eggs) in individuals from Golden Eagle Sable Fish, the only sable fish aquaculture facility in Canada. 3) We will use in-situ hybridization to characterize opsin expression in the brain of Platichthys stellatus, the starry flounder. This is a local and abundant species and, unlike most other flatfish, occurs in left- and right-sided morphs. Opsin expression in zebrafish brain is perfectly symmetrical (Davies et al., 2015). If these opsins are light sensors, we predict that expression domains will be asymmetric in flatfish, whose brains are essentially lying on their side. We also predict that left- and right-sided starry flounder will differ from one another. The research described engages a post-doc, 2 PhDs and 10 undergraduates.