Browsing by Person "Betz, Anja"

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Kinetics and dynamics of plant toxins in sequestering insects
(2024) Betz, Anja; Petschenka, Georg
Many plants produce a variety of chemical compounds to protect themselves from herbivorous insects. However, numerous insects have developed adaptations to tolerate and even utilize these toxins. These adaptations can include behavioral changes (feeding on or avoiding toxic plant parts), detoxification (metabolizing or excreting the toxins), barriers (preventing uptake or protecting target sites), or sequestration (storing secondary plant compounds in the body for self-defense). A widely distributed group of phytochemicals are the cardiac glycosides, which have been extensively studied in chemical ecology. These toxic steroids act in a highly specific manner by inhibiting the Na+/K+-ATPase, an essential transmembrane cation carrier that is ubiquitously expressed in animal cells. A cardiac glycoside-resistant Na+/K+-ATPase was first described in the monarch butterfly (Danaus plexippus, Lepidoptera: Danaini), which sequesters cardiac glycosides from its food plants. This led to the hypothesis that the evolution of a resistant Na+/K+-ATPase is linked to the sequestration of cardiac glycosides. It is now known that species from at least six insect orders possess Na+/K+-ATPases with reduced affinity for cardiac glycosides. This resistance is mediated by a few amino acid substitutions in the binding site, a mechanism referred to as "target site insensitivity." Interestingly, the ability to tolerate dietary cardiac glycosides does not necessarily require a resistant Na+/K+-ATPase. For example, the non-sequestering common crow (Euploea core, Lepidoptera: Danaini) can develop on cardiac glycoside-containing plants without expressing a resistant Na+/K+-ATPase. Although plant toxin sequestration has been documented in more than 275 insect species, the underlying kinetics and dynamics of toxin transport - such as the duration and mode of transport, target sites, barriers, and potential modified binding sites - remain largely unknown. Like the Danaini, milkweed bugs (Heteroptera: Lygaeinae) also have the ability to sequester cardiac glycosides. Remarkably, one species within this family, Spilostethus saxatilis, is able to sequester not only cardiac glycosides but also colchicine alkaloids from autumn crocus (Colchicum autumnale, Liliales: Colchicaceae). Colchicine binds to unpolymerized betatubulin at the interface with alpha-tubulin, thereby inhibiting microtubule formation. However, it is not known how colchicine resistance is achieved in S. saxatilis and whether there is also target site insensitivity. In my dissertation, I investigated the mechanisms behind the sequestration of cardiac glycosides in D. plexippus and colchicine in S. saxatilis within a phylogenetic framework. In D. plexippus, I was able to show that sequestration begins at the behavioral level, because unlike related species, caterpillars can increase their toxicity in the adult stage by drinking plant latex containing cardiac glycosides. I also found that a large portion of cardiac glycosides is lost during metamorphosis, suggesting that excessive toxin intake during the larval stage may be important. Furthermore, I could show that the uptake of the toxin by the intestinal epithelium is a very rapid process although probably not specific to D. plexippus, as uptake in nonsequestering lepidopterans does also occur. Regarding the sequestration of colchicine in S. saxatilis, I identified an amino acid substitution in the colchicine binding site of tubulin. Subsequently, an in vitro colchicine binding assays with crude tubulin extract of bug tissue and an in vivo feeding experiment with genetically modified Drosophila with one matching substitution in the colchicine binding site were conducted. Together both experiments suggest that this modification may play a crucial role in the sequestration of colchicine, indicating a novel natural target site resistance. My dissertation provides new insights into how insects adapt to toxic food plants and how they sequester their toxic metabolites. This contributes not only to a better understanding of how insects cope with natural plant toxins, but also to a better understanding of the physiological complexity of these adaptations.
Spatial metabolomics reveal divergent cardenolide processing in the monarch (Danaus plexippus) and the common crow butterfly (Euploea core)
(2023) Dreisbach, Domenic; Bhandari, Dhaka R.; Betz, Anja; Tenbusch, Linda; Vilcinskas, Andreas; Spengler, Bernhard; Petschenka, Georg
Although being famous for sequestering milkweed cardenolides, the mechanism of sequestration and where cardenolides are localized in caterpillars of the monarch butterfly (Danaus plexippus, Lepidoptera: Danaini) is still unknown. While monarchs tolerate cardenolides by a resistant Na+/K+‐ATPase, it is unclear how closely related species such as the nonsequestering common crow butterfly (Euploea core, Lepidoptera: Danaini) cope with these toxins. Using novel atmospheric‐pressure scanning microprobe matrix‐assisted laser/desorption ionization mass spectrometry imaging, we compared the distribution of cardenolides in caterpillars of D. plexippus and E. core. Specifically, we tested at which physiological scale quantitative differences between both species are mediated and how cardenolides distribute across body tissues. Whereas D. plexippus sequestered most cardenolides from milkweed (Asclepias curassavica), no cardenolides were found in the tissues of E. core. Remarkably, quantitative differences already manifest in the gut lumen: while monarchs retain and accumulate cardenolides above plant concentrations, the toxins are degraded in the gut lumen of crows. We visualized cardenolide transport over the monarch midgut epithelium and identified integument cells as the final site of storage where defences might be perceived by predators. Our study provides molecular insight into cardenolide sequestration and highlights the great potential of mass spectrometry imaging for understanding the kinetics of multiple compounds including endogenous metabolites, plant toxins, or insecticides in insects.