Distinct developmental mechanisms influence sexual dimorphisms in the milkweed bug Oncopeltus fasciatus

Sexual dimorphism is a common feature of animals. Sex determination mechanisms vary widely among species and evolve rapidly. Until recently studies have found consistent mechanisms across the body of each individual determine female or male dimorphic body structures. In sexually dimorphic cells throughout the body of Drosophila, the relative dosage of autosomes and X chromosomes leads indirectly to alternatively spliced transcripts from the gene doublesex. The female Dsx isoform interacts with the mediator complex protein encoded by intersex to activate female development in flies. In males the transcription factor encoded by fruitless promotes male-specific behavior. In the milkweed bug Oncopeltus fasciatus, we find a requirement for different combinations of these genes during development of distinct dimorphic structures, within the same sex, suggesting a previously unappreciated level of diversity in sex determination. While intersex and fruitless are structurally conserved, doublesex has a history of duplication and divergence among Paraneoptera. Three doublesex paralogs in O. fasciatus produce multiple transcripts with sex- and tissue-specific expression. intersex and fruitless are expressed across the body, in females and males. RNA interference reveals only one doublesex paralog functions in somatic sex determination. Knockdown of doublesex and fruitless produces intersex phenotypic conditions in two sexually dimorphic structures: genitalia and abdominal sternites. In contrast, intersex is required for dimorphic development of female and male genitalia, but not for sternite dimorphism. These results reveal sex determination roles for intersex and fruitless distinct from their orthologs in other insects. Our results illuminate a novel form of developmental diversity in insect sex determination.

Dietary cardenolides enhance growth and change the direction of the fecundity-longevity trade-off in milkweed bugs (Heteroptera: Lygaeinae)

1. Sequestration, i.e., the accumulation of plant toxins into body tissues for defence, is primarily observed in specialised insects. Sequestration was frequently predicted to incur a physiological cost mediated by increased exposure to plant toxins and may require resistance traits different from those of non-sequestering insects. Alternatively, sequestering species could experience a cost in the absence of toxins due to selection on physiological homeostasis under permanent exposure of sequestered toxins in body tissues. 2. Milkweed bugs (Heteroptera: Lygaeinae) sequester high amounts of plant-derived cardenolides. Although being potent inhibitors of the ubiquitous animal enzyme Na+/K+-ATPase, milkweed bugs can tolerate cardenolides by means of resistant Na+/K+-ATPases. Both adaptations, resistance and sequestration, are ancestral traits shared by most species of the Lygaeinae. 3. Using four milkweed bug species and the related European firebug (Pyrrhocoris apterus) showing different combinations of the traits ′cardenolide resistance′ and ′cardenolide sequestration′, we set out to test how the two traits affect larval growth upon exposure to dietary cardenolides in an artificial diet system. While cardenolides impaired the growth of P. apterus nymphs neither possessing a resistant Na+/K+-ATPase nor sequestering cardenolides, growth was not affected in the non-sequestering milkweed bug Arocatus longiceps, which possesses a resistant Na+/K+-ATPase. Remarkably, cardenolides increased growth in the sequestering dietary specialists Caenocoris nerii and Oncopeltus fasciatus but not in the sequestering dietary generalist Spilostethus pandurus, which all possess a resistant Na+/K+-ATPase. 4. We then assessed the effect of dietary cardenolides on additional life history parameters, including developmental speed, the longevity of adults, and reproductive success in O. fasciatus. Remarkably, nymphs under cardenolide exposure developed substantially faster and lived longer as adults. However, fecundity of adults was reduced when maintained on cardenolide-containing diet for their entire life-time but not when adults were transferred to non-toxic sunflower seeds. 5. We speculate that the resistant Na+/K+-ATPase of milkweed bugs is selected for working optimally in a ′toxic environment′, i.e. when sequestered cardenolides are stored in the body tissues. Contrary to the assumption that toxins sequestered for defence produce a physiological burden, our data suggest that they can even increase fitness in specialised insects.

Shifting roles of Drosophila pair-rule gene orthologs: segmental expression and function in the milkweed bug Oncopeltus fasciatus

The discovery of pair-rule genes (PRGs) in Drosophila revealed the existence of an underlying two-segment-wide prepattern directing embryogenesis. The milkweed bug Oncopeltus, a hemimetabolous insect, is a more representative arthropod: most of its segments form sequentially after gastrulation. Here we report the expression and function of orthologs of the complete set of nine Drosophila PRGs in Oncopeltus. Seven Of-PRG-orthologs are expressed in stripes in the primordia of every segment, rather than every-other segment, Of-runt is PR-like, and several are also expressed in the segment addition zone. RNAi-mediated knockdown of Of-odd-skipped, paired and sloppy-paired impacted all segments, with no indication of PR-like register. We confirm that Of-E75A is expressed in PR-like stripes, although it is not PR in Drosophila, demonstrating the existence of an underlying PR-like prepattern in Oncopeltus. These findings reveal that a switch occurred in regulatory circuits leading to segment formation: while several holometabolous insects are “Drosophila-like,” utilizing PRG-orthologs for PR-patterning, most Of-PRGs are expressed segmentally in Oncopeltus, a more basally-branching insect. Thus, an evolutionarily stable phenotype – segment formation – is directed by alternate regulatory pathways in diverse species.Summary StatementDespite the broad of conservation of segmentation in insects, the regulatory genes underlying this process in Drosophila have different roles in the hemipteran, Oncopeltus fasciatus.