Abstract
Butterfly eyespot colour patterns are a key example of how a novel trait can appear in association with the co-option of developmental patterning genes. Little is known, however, about how, or even whether, co-opted genes function in eyespot development. Here we use CRISPR/Cas9 genome editing to determine the roles of two co-opted transcription factors that are expressed during early eyespot determination. We found that deletions in a single gene, spalt, are sufficient to reduce or completely delete eyespot colour patterns, thus demonstrating a positive regulatory role for this gene in eyespot determination. Conversely, and contrary to previous predictions, deletions in Distal-less (Dll) result in an increase in the size and number of eyespots, illustrating a repressive role for this gene in eyespot development. Altogether our results show that the presence, absence and shape of butterfly eyespots can be controlled by the activity of two co-opted transcription factors.
Highlights
Eyespot colour patterns are a key example of how a novel trait can appear in association with the co-option of developmental patterning genes
To test the utility of CRISPR/Cas[9] for making G0 deletion mosaic butterflies, we optimized a protocol in the painted lady butterfly Vanessa cardui by targeting the melanin pigmentation gene Dopa decarboxylase (Ddc), reasoning that pigmentation defects should allow for easy visualization of potential mosaicism in wings (Fig. 1)
In this study we used CRISPR/Cas[9] somatic mosaics to show that the transcription factor genes spalt and Dll play key roles in butterfly wing pattern development, and by extension, that the co-option of these genes has played a role in colour pattern evolution
Summary
Eyespot colour patterns are a key example of how a novel trait can appear in association with the co-option of developmental patterning genes. Comparative expression studies show that the transcription factor spalt is one of the earliest factors expressed in eyespot foci, and phylogenetic work suggests that this early spalt expression is a conserved ancestral trait across nymphalid butterflies[6,8] Because of these observations it has been proposed that spalt may be a key upstream regulator of eyespot determination[1]. We use CRISPR/Cas[9] genome editing to assess the developmental functions of spalt and Dll in eyespot colour pattern development, and, more broadly, to show that the co-option of these transcription factors has played a role in the evolution of butterfly wing patterns
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