A Single Origin for Nymphalid Butterfly Eyespots Followed by Widespread Loss of Associated Gene Expression

Abstract

Understanding how novel complex traits originate involves investigating the time of origin of the trait, as well as the origin of its underlying gene regulatory network in a broad comparative phylogenetic framework. The eyespot of nymphalid butterflies has served as an example of a novel complex trait, as multiple genes are expressed during eyespot development. Yet the origins of eyespots remain unknown. Using a dataset of more than 400 images of butterflies with a known phylogeny and gene expression data for five eyespot-associated genes from over twenty species, we tested origin hypotheses for both eyespots and eyespot-associated genes. We show that eyespots evolved once within the family Nymphalidae, approximately 90 million years ago, concurrent with expression of at least three genes associated with early eyespot development. We also show multiple losses of expression of most genes from this early three-gene cluster, without corresponding losses of eyespots. We propose that complex traits, such as eyespots, may have originated via co-option of a large pre-existing complex gene regulatory network that was subsequently streamlined of genes not required to fulfill its novel developmental function.

Introgression and repeated co-option facilitated the recurrent emergence of C4photosynthesis among close relatives

“…Decomposing the phenotype into its components can solve this problem, especially when the underlying genetic determinism is considered (Oliver et al. 2012; Niemiller et al. 2013; Glover et al.…”

Nymphalid eyespot serial homologues originate as a few individualized modules

“…This assumption is not unwarranted because both spots and eyespots occur in the centre of wing sectors, and both appear to differentiate from central signalling cells [25,26]. Note that the origin of eyespots has previously been associated with the origin of expression of a series of developmental genes that are not associated with simple monochromatic spots found in more basal butterfly lineages [13]. The 'eyespot' character state is thus assumed to be different from but homologous to the character state 'spot'.…”

“…A clear example of multiple potential points of network disruption, coupled with preservation of a functional eyespot network in other locations, is evident in the expression dynamics of eyespot-associated genes in B. anynana and J. coenia ( figure 3). In B. anynana, at least five proteins associated with adult eyespots are expressed in the future eyespot centres of larval wing discs [13], including the transcription factors Spalt (Sal) and distal-less (Dll). In the M 2 and M 3 sectors of the anterior wing, which lack adult eyespots, Sal and Dll are initially expressed, then subsequently the proteins disappear, presumably through downregulation of the respective genes.…”

“…Eyespots, which are characterized by a series of concentric rings of colour, likely originated early in nymphalid evolution, approximately 90 Ma [13]. These conspicuous bull's eye traits serve crucial roles in mate signalling and predator avoidance [14 -17].…”

Camouflage Variations on a Theme of the Nymphalid Ground Plan

“…To further understand the evolutionary trajectories of the NGP, it is necessary to investigate the molecular mechanisms underlying NGP. Previous studies revealed the molecular mechanisms underlying eyespots (ocelli), one of the NGP elements in butterfly wings (Carroll et al 1994;Brakefield et al 1996;Keys et al 1999;Brunetti et al 2001;Beldade and Brakefield 2002;Monteiro et al 2006;Oliver et al 2012;Monteiro et al 2013;Monteiro 2015;Zhang and Reed 2016;Beldade and Peralta 2017). Molecular studies have also uncovered several morphogens (e.g., Wnt1/wingless, WntA) and transcription factors (e.g., aristaless2, engrailed) associated with other elements of NGP (Brunetti et al 2001;Monteiro et al 2006;Reed 2010, 2014).…”