Abstract:
The genetic basis of morphological variation provides a major topic in evolutionary developmental biology. Fish of the genus Danio, containing the model species zebrafish, Danio rerio, represent a system to study pigment pattern diversification as they display amazingly different patterns ranging from horizontal stripes, to vertical bars or spots. Stripe formation in D. rerio is a self-organizing process based on cell-contact mediated interactions between melanophores, xanthophores and iridophores. Little is known about the genetic and cellular basis of pigment pattern formation and evolution in other Danio species. Genes known to be involved in stripe formation in D. rerio might have functionally diverged to produce a pattern of vertical bars in its sibling species, Danio aesculapii. In collaboration with my colleagues, I showed by mutant analysis that the same three pigment cell types are required for bar formation. Reciprocal hemizygosity tests with genes, which are known to be involved in interactions between the pigment cells in D. rerio, identified the potassium channel gene kcnj13, but not the gap junction genes, gja4 and gja5b, or the adhesion molecule gene igsf11 as diverged between the two species. Further complementation tests with eight additional Danio species suggested evolutionary change in pigment patterns through repeated and independent functional divergences in kcnj13, gja5b and igsf11 across the genus. Focusing on kcnj13, we used in vivo imaging of transgenic reporters, transplantation experiments and lineage tracing of pigment cell population in chimeras and found that the shapes of all three types of pigment cells are affected in the D. rerio mutants, although the gene function is only required in melanophores. These differences, similar to the ones in D. rerio mutants, were also partly observed between D. rerio and D. aesculapii, might therefore underlie the evolutionary change of the divergent patterns. Using molecular, biochemical and bioinformatic analyses we confirmed the homo-tetrameric structure of the channel, which explains the dominant phenotype of most known mutations. A transcriptome-wide allele-specific expression analysis indicated higher expression of the D. rerio allele in hybrids between the two species. Together with our findings that the protein from both species are able to rescue the stripe phenotype in transgenic rescue lines, this confirmed cis-regulatory evolution of kcnj13. Species-specific pigment cell interactions could be important factors contributing to the variation in pigment patterns. This work highlights the genetic complexity underlying the diversification of pigment patterning and shows that the evolutionary history of biodiversity can be reconstructed in Danio fish.
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