Optix and cortex/ivory/mir-193 again: the repeated use of two mimicry hotspot loci.

The extent to which evolution is repeatable has been a debated topic among evolutionary biologists. Although rewinding the tape of life perhaps would not lead to the same outcome every time, repeated evolution of analogous genes for similar functions has been extensively reported. Wing phenotypes of butterflies and moths have provided a wealth of examples of gene re-use, with certain 'hotspot loci' controlling wing patterns across diverse taxa. Here, we present an example of convergent evolution in the molecular genetic basis of Batesian wing mimicry in two Hypolimnas butterfly species. We show that mimicry is controlled by variation near cortex/ivory/mir-193, a known butterfly hotspot locus. By dissecting the genetic architecture of mimicry in Hypolimnas misippus and Hypolimnas bolina, we present evidence that distinct non-coding regions control the development of white pattern elements in the forewing and hindwing of the two species, suggesting independent evolution, and that no structural variation is found at the locus. Finally, we also show that orange coloration in H. bolina is associated with optix, a well-known patterning gene. Overall, our study once again implicates variation near the hotspot loci cortex/ivory/mir-193 and optix in butterfly wing mimicry and thereby highlights the repeatability of adaptive evolution.

Transposable Element Insertions Are Associated with Batesian Mimicry in the Pantropical Butterfly Hypolimnas misippus.

Hypolimnas misippus is a Batesian mimic of the toxic African Queen butterfly (Danaus chrysippus). Female H. misippus butterflies use two major wing patterning loci (M and A) to imitate three color morphs of D. chrysippus found in different regions of Africa. In this study, we examine the evolution of the M locus and identify it as an example of adaptive atavism. This phenomenon involves a morphological reversion to an ancestral character that results in an adaptive phenotype. We show that H. misippus has re-evolved an ancestral wing pattern present in other Hypolimnas species, repurposing it for Batesian mimicry of a D. chrysippus morph. Using haplotagging, a linked-read sequencing technology, and our new analytical tool, Wrath, we discover two large transposable element insertions located at the M locus and establish that these insertions are present in the dominant allele responsible for producing mimetic phenotype. By conducting a comparative analysis involving additional Hypolimnas species, we demonstrate that the dominant allele is derived. This suggests that, in the derived allele, the transposable elements disrupt a cis-regulatory element, leading to the reversion to an ancestral phenotype that is then utilized for Batesian mimicry of a distinct model, a different morph of D. chrysippus. Our findings present a compelling instance of convergent evolution and adaptive atavism, in which the same pattern element has independently evolved multiple times in Hypolimnas butterflies, repeatedly playing a role in Batesian mimicry of diverse model species.

Colour polymorphism associated with a gene duplication in male wood tiger moths.

Colour is often used as an aposematic warning signal, with predator learning expected to lead to a single colour pattern within a population. However, there are many puzzling cases where aposematic signals are also polymorphic. The wood tiger moth, Arctia plantaginis, displays bright hindwing colours associated with unpalatability, and males have discrete colour morphs which vary in frequency between localities. In Finland, both white and yellow morphs can be found, and these colour morphs also differ in behavioural and life-history traits. Here, we show that male colour is linked to an extra copy of a yellow family gene that is only present in the white morphs. This white-specific duplication, which we name valkea, is highly upregulated during wing development. CRISPR targeting valkea resulted in editing of both valkea and its paralog, yellow-e, and led to the production of yellow wings. We also characterise the pigments responsible for yellow, white, and black colouration, showing that yellow is partly produced by pheomelanins, while black is dopamine-derived eumelanin. Our results add to a growing number of studies on the genetic architecture of complex and seemingly paradoxical polymorphisms, and the role of gene duplications and structural variation in adaptive evolution.

A novel terpene synthase controls differences in anti-aphrodisiac pheromone production between closely related Heliconius butterflies.

Plants and insects often use the same compounds for chemical communication, but not much is known about the genetics of convergent evolution of chemical signals. The terpene (E)-ß-ocimene is a common component of floral scent and is also used by the butterfly Heliconius melpomene as an anti-aphrodisiac pheromone. While the biosynthesis of terpenes has been described in plants and microorganisms, few terpene synthases (TPSs) have been identified in insects. Here, we study the recent divergence of 2 species, H. melpomene and Heliconius cydno, which differ in the presence of (E)-ß-ocimene; combining linkage mapping, gene expression, and functional analyses, we identify 2 novel TPSs. Furthermore, we demonstrate that one, HmelOS, is able to synthesise (E)-ß-ocimene in vitro. We find no evidence for TPS activity in HcydOS (HmelOS ortholog of H. cydno), suggesting that the loss of (E)-ß-ocimene in this species is the result of coding, not regulatory, differences. The TPS enzymes we discovered are unrelated to previously described plant and insect TPSs, demonstrating that chemical convergence has independent evolutionary origins.

The genomics of coloration provides insights into adaptive evolution.

Coloration is an easily quantifiable visual trait that has proven to be a highly tractable system for genetic analysis and for studying adaptive evolution. The application of genomic approaches to evolutionary studies of coloration is providing new insight into the genetic architectures underlying colour traits, including the importance of large-effect mutations and supergenes, the role of development in shaping genetic variation and the origins of adaptive variation, which often involves adaptive introgression. Improved knowledge of the genetic basis of traits can facilitate field studies of natural selection and sexual selection, making it possible for strong selection and its influence on the genome to be demonstrated in wild populations.

Publisher Correction: The genomics of coloration provides insights into adaptive evolution.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The dynamics of cyanide defences in the life cycle of an aposematic butterfly: Biosynthesis versus sequestration.

Heliconius butterflies are highly specialized in Passiflora plants, laying eggs and feeding as larvae only on them. Interestingly, both Heliconius butterflies and Passiflora plants contain cyanogenic glucosides (CNglcs). While feeding on specific Passiflora species, Heliconius melpomene larvae are able to sequester simple cyclopentenyl CNglcs, the most common CNglcs in this plant genus. Yet, aromatic, aliphatic, and modified CNglcs have been reported in Passiflora species and they were never tested for sequestration by heliconiine larvae. As other cyanogenic lepidopterans, H. melpomene also biosynthesize the aliphatic CNglcs linamarin and lotaustralin, and their toxicity does not rely exclusively on sequestration. Although the genes encoding the enzymes in the CNglc biosynthesis have not yet been biochemically characterized in butterflies, the cytochromes P450 CYP405A4, CYP405A5, CYP405A6 and CYP332A1 have been hypothesized to be involved in this pathway in H. melpomene. In this study, we determine how the CNglc composition and expression of the putative P450s involved in the biosynthesis of these compounds vary at different developmental stages of Heliconius butterflies. We also establish which kind of CNglcs H. melpomene larvae can sequester from Passiflora. By analysing the chemical composition of the haemolymph from larvae fed with different Passiflora diets, we show that H. melpomene is able to sequestered prunasin, an aromatic CNglcs, from P. platyloba. They are also able to sequester amygdalin, gynocardin, [C13/C14]linamarin and [C13/C14]lotaustralin painted on the plant leaves. The CNglc tetraphyllin B-sulphate from P. caerulea is not detected in the larval haemolymph, suggesting that such modified CNglcs cannot be sequestered by Heliconius. Although pupae and virgin adults contain dihydrogynocardin resulting from larval sequestration, this compound was metabolized during adulthood, and not used as nuptial gift or transferred to the offspring. Thus, we speculate that dihydrogynocardin is catabolized to recycle nitrogen and glucose, and/or to produce fitness signals during courtship. Mature adults have a higher concentration of CNglcs than any other developmental stages due to increased de novo biosynthesis of linamarin and lotaustralin. Accordingly, all CYP405As are expressed in adults, whereas larvae mostly express CYP405A4. Our results shed light on the importance of CNglcs for Heliconius biology and their coevolution with Passiflora.