Comparative Functional Genomics of Swallowtail Butterfly Mimicry Polymorphism

Many animals have evolved sexual dimorphism and even polymorphism, where males and females of the same species share nearly identical genomes but develop one or more different sex-limited morphs. In one striking example, multiple species of Papilio swallowtail butterflies exhibit female-limited polymorphism, in which one or more of the female morphs evolved wing color mimicry of a separate toxic model butterfly. The development of these differing wing color phenotypes is controlled by a single transcription factor, doublesex (dsx), that evolved a novel allele which switches on the mimetic pattern developmental pathway. Many studies have been conducted to understand the dsx mimicry switch in Papilio alphenor, but the overarching evolutionary history of the novel dsx allele, and whether the mimicry patterning mechanism differs across these multiple other species of Papilio, is unknown. To better understand the mechanism by which dsx controls mimicry, I characterized the genetic and developmental mechanisms that control this polymorphism in the previously uncharacterized Papilio rumanzovia. The mimicry switch is most likely controlled by gene expression differences between different dsx alleles, which are regulated by cis-regulatory elements (CREs). Therefore, I analyzed dsx spatial expression, the CREs controlling dsx expression, and genes that operate downstream of dsx, to form a distinct understanding of the mechanisms behind dsx-controlled mimicry in P. rumanzovia. I found multiple CREs in dsx that are differentially accessible between mimetic and non-mimetic P. rumanzovia, one which is notably specific to the mimetic allele and orthologous to the mimetic P. alphenor dsx allele. I also identified strong candidate genes with roles in dsx-controlled wing color patterning through a genome-wide analysis of RNA-seq data, in which sirtuin-2 is a gene linked to dsx-controlled mimicry through past work in P. alphenor. These results suggest the emergence of the mimetic allele in a common ancestor of Papilio rather than independent evolution across multiple species, a finding which contributes to our existing knowledge of sexual dimorphism, supergene evolution and development.