Richard H. ffrench-Constant

Butterfly genome reveals promiscuous exchange of mimicry adaptations among species

The evolutionary importance of hybridization and introgression has long been debated. Hybrids are usually rare and unfit, but even infrequent hybridization can aid adaptation by transferring beneficial traits between species. Here we use genomic tools to investigate introgression in Heliconius, a rapidly radiating genus of neotropical butterflies widely used in studies of ecology, behaviour, mimicry and speciation. We sequenced the genome of Heliconius melpomene and compared it with other taxa to investigate chromosomal evolution in Lepidoptera and gene flow among multiple Heliconius species and races. Among 12,669 predicted genes, biologically important expansions of families of chemosensory and Hox genes are particularly noteworthy. Chromosomal organization has remained broadly conserved since the Cretaceous period, when butterflies split from the Bombyx (silkmoth) lineage. Using genomic resequencing, we show hybrid exchange of genes between three co-mimics, Heliconius melpomene, Heliconius timareta and Heliconius elevatus, especially at two genomic regions that control mimicry pattern. We infer that closely related Heliconius species exchange protective colour-pattern genes promiscuously, implying that hybridization has an important role in adaptive radiation.

Chromosomal rearrangements maintain a polymorphic supergene controlling butterfly mimicry

Supergenes are tight clusters of loci that facilitate the co-segregation of adaptive variation, providing integrated control of complex adaptive phenotypes. Polymorphic supergenes, in which specific combinations of traits are maintained within a single population, were first described for ‘pin’ and ‘thrum’ floral types in Primula and Fagopyrum, but classic examples are also found in insect mimicry and snail morphology. Understanding the evolutionary mechanisms that generate these co-adapted gene sets, as well as the mode of limiting the production of unfit recombinant forms, remains a substantial challenge. Here we show that individual wing-pattern morphs in the polymorphic mimetic butterfly Heliconius numata are associated with different genomic rearrangements at the supergene locus P. These rearrangements tighten the genetic linkage between at least two colour-pattern loci that are known to recombine in closely related species, with complete suppression of recombination being observed in experimental crosses across a 400-kilobase interval containing at least 18 genes. In natural populations, notable patterns of linkage disequilibrium (LD) are observed across the entire P region. The resulting divergent haplotype clades and inversion breakpoints are found in complete association with wing-pattern morphs. Our results indicate that allelic combinations at known wing-patterning loci have become locked together in a polymorphic rearrangement at the P locus, forming a supergene that acts as a simple switch between complex adaptive phenotypes found in sympatry. These findings highlight how genomic rearrangements can have a central role in the coexistence of adaptive phenotypes involving several genes acting in concert, by locally limiting recombination and gene flow.

Genomic islands of divergence in hybridizing Heliconius butterflies identified by large-scale targeted sequencing

Heliconius butterflies represent a recent radiation of species, in which wing pattern divergence has been implicated in speciation. Several loci that control wing pattern phenotypes have been mapped and two were identified through sequencing. These same gene regions play a role in adaptation across the whole Heliconius radiation. Previous studies of population genetic patterns at these regions have sequenced small amplicons. Here, we use targeted next-generation sequence capture to survey patterns of divergence across these entire regions in divergent geographical races and species of Heliconius. This technique was successful both within and between species for obtaining high coverage of almost all coding regions and sufficient coverage of non-coding regions to perform population genetic analyses. We find major peaks of elevated population differentiation between races across hybrid zones, which indicate regions under strong divergent selection. These ‘islands’ of divergence appear to be more extensive between closely related species, but there is less clear evidence for such islands between more distantly related species at two further points along the ‘speciation continuum’. We also sequence fosmid clones across these regions in different Heliconius melpomene races. We find no major structural rearrangements but many relatively large (greater than 1 kb) insertion/deletion events (including gain/loss of transposable elements) that are variable between races.

DDT resistance in Drosophila correlates with Cyp6g1 over-expression and confers cross-resistance to the neonicotinoid imidacloprid

Abstract. Mutagenesis can be used as a means of predicting likely mechanisms of resistance to novel classes of insecticides. We used chemical mutagenesis in Drosophila to screen for mutants that had become resistant to imidacloprid, a neonicotinoid insecticide. Here we report the isolation of two new dominant imidacloprid-resistant mutants. By recombinational mapping we show that these map to the same location as Rst(2)DDT. Furthermore, we show that pre-existing Rst(2)DDT alleles in turn confer cross-resistance to imidacloprid. In order to localize the Rst(2)DDT gene more precisely, we mapped resistance to both DDT and imidacloprid with respect to P-element markers whose genomic location is known. By screening for recombinants between these P-elements and resistance we localized the gene between 48D5–6 and 48F3–6 on the polytene chromosome map. The genomic sequence in this interval shows a cluster of cytochrome P450 genes, one of which, Cyp6g1, is over-expressed in all resistant strains examined. We are now testing the hypothesis that resistance to both compounds is associated with over-expression of this P450 gene.

A single Photorhabdus gene, makes caterpillars floppy (mcf), allows Escherichia coli to persist within and kill insects

Photorhabdus luminescens, a bacterium with alternate pathogenic and symbiotic phases of its lifestyle, represents a source of novel genes associated with both virulence and symbiosis. This entomopathogen lives in a “symbiosis of pathogens” with nematodes that invade insects. Thus the bacteria are symbiotic with entomopathogenic nematodes but become pathogenic on release from the nematode into the insect blood system. Within the insect, the bacteria need to both avoid the peptide- and cellular- (hemocyte) mediated immune response and also to kill the host, which then acts as a reservoir for bacterial and nematode reproduction. However, the mechanisms whereby Photorhabdus evades the insect immune system and kills the host are unclear. Here we show that a single large Photorhabdus gene, makes caterpillars floppy (mcf), is sufficient to allow Esherichia coli both to persist within and kill an insect. The predicted high molecular weight Mcf toxin has little similarity to other known protein sequences but carries a BH3 domain and triggers apoptosis in both insect hemocytes and the midgut epithelium.

A Conserved Supergene Locus Controls Colour Pattern Diversity in Heliconius Butterflies

We studied whether similar developmental genetic mechanisms are involved in both convergent and divergent evolution. Mimetic insects are known for their diversity of patterns as well as their remarkable evolutionary convergence, and they have played an important role in controversies over the respective roles of selection and constraints in adaptive evolution. Here we contrast three butterfly species, all classic examples of Müllerian mimicry. We used a genetic linkage map to show that a locus, Yb, which controls the presence of a yellow band in geographic races of Heliconius melpomene, maps precisely to the same location as the locus Cr, which has very similar phenotypic effects in its co-mimic H. erato. Furthermore, the same genomic location acts as a “supergene”, determining multiple sympatric morphs in a third species, H. numata. H. numata is a species with a very different phenotypic appearance, whose many forms mimic different unrelated ithomiine butterflies in the genus Melinaea. Other unlinked colour pattern loci map to a homologous linkage group in the co-mimics H. melpomene and H. erato, but they are not involved in mimetic polymorphism in H. numata. Hence, a single region from the multilocus colour pattern architecture of H. melpomene and H. erato appears to have gained control of the entire wing-pattern variability in H. numata, presumably as a result of selection for mimetic “supergene” polymorphism without intermediates. Although we cannot at this stage confirm the homology of the loci segregating in the three species, our results imply that a conserved yet relatively unconstrained mechanism underlying pattern switching can affect mimicry in radically different ways. We also show that adaptive evolution, both convergent and diversifying, can occur by the repeated involvement of the same genomic regions.

Molecular biology of insect neuronal GABA receptors

Ionotropic gamma-aminobutyric acid (GABA) receptors are distributed throughout the nervous systems of many insect species. As with their vertebrate counterparts, GABAA receptors and GABAC receptors, the binding of GABA to ionotropic insect receptors elicits a rapid, transient opening of anion-selective ion channels which is generally inhibitory. Although insect and vertebrate GABA receptors share a number of structural and functional similarities, their pharmacology differs in several aspects. Recent studies of cloned Drosophila melanogaster GABA receptors have clarified the contribution of particular subunits to these differences. Insect ionotropic GABA receptors are also the target of numerous insecticides and an insecticide-resistant form of a Drosophila GABA-receptor subunit has enhanced our understanding of the structure-function relationship of one aspect of pharmacology common to both insect and vertebrate GABA receptors, namely antagonism by the plant-derived toxin picrotoxinin.

The tc genes of Photorhabdus: a growing family.

The toxin complex (tc) genes of Photorhabdus encode insecticidal, high molecular weight Tc toxins. These toxins have been suggested as useful alternatives to those derived from Bacillus thuringiensis for expression in insect-resistant transgenic plants. Although Photorhabdus luminescens is symbiotic with nematodes that kill insects, tc genes have recently been described from other insect-associated bacteria such as Serratia entomophila, an insect pathogen, and Yersinia pestis, the causative agent of bubonic plague, which has a flea vector. Here, recent advances in our understanding of the tc gene family are reviewed in view of their potential development as insect-control agents.

An antibiotic produced by an insect-pathogenic bacterium suppresses host defenses through phenoloxidase inhibition

Photorhabdus is a virulent pathogen that kills its insect host by overcoming immune responses. The bacterium also secretes a range of antibiotics to suppress the growth of other invading microorganisms. Here we show that Photorhabdus produces a small-molecule antibiotic (E)-1,3-dihydroxy-2-(isopropyl)-5-(2-phenylethenyl)benzene (ST) that also acts as an inhibitor of phenoloxidase (PO) in the insect host Manduca sexta. The Photorhabdus gene stlA encodes an enzyme that produces cinnamic acid, a key precursor for production of ST, and a mutation in stlA results in loss of ST production and PO inhibitory activity, which are both restored by genetic complementation of the mutant and also by supplying cinnamic acid. ST is produced both in vitro and in vivo in sufficient quantities to account for PO inhibition and is the only detectable solvent-extractable inhibitor. A Photorhabdus stlA− mutant is significantly less virulent, proliferates slower within the host, and provokes the formation of significantly more melanotic nodules than wild-type bacteria. Virulence of the stlA− mutant is also rescued by supplying cinnamic acid. The proximate cause of the virulence effect, however, is the inhibition of PO, because the effect of the stlA− mutation on virulence is abolished in insects in which PO has been knocked down by RNA interference (RNAi). Thus, ST has a dual function both as a PO inhibitor to counter host immune reactions and also as an antibiotic to exclude microbial competitors from the insect cadaver.

Cis-Regulatory Elements in the Accord Retrotransposon Result in Tissue-Specific Expression of the Drosophila melanogaster Insecticide Resistance Gene Cyp6g1

Transposable elements are a major mutation source and powerful agents of adaptive change. Some transposable element insertions in genomes increase to a high frequency because of the selective advantage the mutant phenotype provides. Cyp6g1-mediated insecticide resistance in Drosophila melanogaster is due to the upregulation of the cytochrome P450 gene Cyp6g1, leading to the resistance to a variety of insecticide classes. The upregulation of Cyp6g1 is correlated with the presence of the long terminal repeat (LTR) of an Accord retrotransposon inserted 291bp upstream of the Cyp6g1 transcription start site. This resistant allele (DDT-R) is currently at a high frequency in D. melanogaster populations around the world. Here, we characterize the spatial expression of Cyp6g1 in insecticide-resistant and -susceptible strains. We show that the Accord LTR insertion is indeed the resistance-associated mutation and demonstrate that the Accord LTR carries regulatory sequences that increase the expression of Cyp6g1 in tissues important for detoxification, the midgut, Malpighian tubules, and the fat body. This study provides a significant example of how changes in tissue-specific gene expression caused by transposable-element insertions can contribute to adaptation.