The source of organismal diversity and the ways in which genotypes result in phenotypes are key questions for understanding evolution. The genomic mechanisms shaping diversity can vary across organisms, with species having small population sizes (e.g. humans) shaped by different evolutionary forces from species having large population sizes (e.g. drosophila). In this proposal, we focus on African cichlid fishes as they are an ideal model to identify genomic mechanisms shaping phenotypic diversity and speciation in relatively small populations. We will examine the genetic basis of opsin gene expression, which determines visual sensitivities. African cichlids are colorful, visual animals that show significant differences in spectral sensitivity, even between closely related species. This is a result of their having seven spectrally distinct cone opsin genes, with subsets of the genes expressed in three combinations or visual palettes: short, medium and long. Some species progress from the short to medium to long palettes as they develop from larvae to juveniles to adults. However, adults of sister taxa can differ in which palette they express by altering this developmental progression. We have previously made genetic crosses between species with different palettes and identified four candidate genes with corresponding regulatory mutations. In this proposal, we seek to confirm the role of these regulatory mutations in shaping the opsin gene regulatory network. We will use in vitro methods (luciferase assays; electron mobility shift assays) as well as assessments of CRISPR generated mutants (qPCR, RNAseq, in situ hybridization) to discern how the regulatory mutations alter gene expression. Our preliminary findings suggest that the opsin gene regulatory network is set up as a series of developmental switches. Within Lake Malawi cichlids, regulatory mutations arose for three transcription factors and one opsin. These large (0.4-1kb) indels alter the regulatory domains of these candidate genes, removing the network links and so switching the developmental state. We hypothesize that these regulatory indels occurred as the result of mobile transposable elements, with fixation of these indels in small populations leading to rapid phenotypic divergence. Our goal is to detail how the regulatory network of the opsin genes has evolved across different species. Evolution of opsin gene expression could potentially serve as a model for how other cichlid phenotypes diverge.
Understanding the link between genotypes and phenotypes is critical to unraveling the genetic basis of human health traits, such as disease susceptibility. This proposal seeks to determine the kinds of genetic changes found in organisms with small populations, like humans.
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