Genomic and genetic insights into speciation and pigment pattern diversification in Danio fishes

The Danioninae subfamily of teleost fishes boasts up to four hundred distinct species that have evolved to display a stunning diversity of morphological forms. Here we use newly assembled genome sequences of four laboratory and wild zebrafish strains as well as eleven species of the Danio and Danionella genera to explore their phylogenetic history and the genetic basis of pigment pattern diversification. Phylogenomic analyses uncover extensive introgression and incomplete lineage sorting that have obscured phylogenetic relationships within Danio and corroborate an ancient hybrid origin of zebrafish. Whereas D. rerio inherited ancestral horizontal stripes, relatives repeatedly evolved spots and vertical bars. Interspecific complementation tests reveal functional divergence of the adhesion molecule gene igsf11 and the gap junction gene gja5b between the striped zebrafish and Danio species with divergent patterns. Comparative genomic and transcriptomic analyses suggest that protein and regulatory evolution have accompanied pigment pattern diversification. Our analyses elucidate complex genetic changes underlying the phylogenetic history and morphological diversification in the Danio genus. Resolved phylogenetic relationships, available genome assemblies, transcriptomes, and genetic tractability establish Danio fish species as excellent models for biomedical research in vertebrates. Overall design: Bulk RNA-seq of FACS-sorted pigment cells and whole skin from adult interspecies Danio hybrids carrying cell-type specific fluorescent reporters for melanophores (tyrp1b:mem-mCherry), xanthophores (aox5:mem-EGFP)49 or iridophores (pnp4a:mem-mCherry). D. aesculapii, D. kyathit, D. albolineatus, and D. nigrofasciatus to female D. rerio. We dissociated and FACS sorted adult hybrid skin to enrich for iridophores, melanophores, or xanthophores based on reporter expression, yielding 36 cell-enriched biological samples (3 cell types x 3 replicates x 4 paternal species) and 12 whole skin control samples (3 replicates x 4 paternal species).