Escalation in coevolution: Characterization of novel plant toxins, how specialist herbivores cope, and tests of phylogenetic intensification in plant defense potency

以植物为食的昆虫被认为对它们所吃的植物组织中的化学防御施加了自然选择。这里正在进行的研究是在乳草植物上进行的，这种植物含有一种高度有效的毒素--拉布福林，这种毒素在几个物种中已经进化到高浓度。当高表达时，拉布福明抑制帝王蝶和大马利筋虫的一种必要的运输酶(钠钾泵)。这些专门的害虫通过生物化学将拉布福林改造成毒性较低的化合物，并将这些产物隔离以用于自身防御。君主和种虫高度耐受的钠泵和生化修饰表明，拉布拉福明处于马利筋和其草食动物之间相互进化的中心。这项工作旨在通过生理和进化分析来揭示这种相互作用的机制。这项研究也为理解植物和害虫之间的自然和农业相互作用提供了一个模型。最后，马利筋-君主制度很适合公众参与，因为它具有标志性的地位，受到保护关注的人口不断减少，并在学校课程中使用。作为该项目的一部分，研究人员正在为K-12学校开发模块，并通过演示和博客参与公共宣传。在以几种特殊的昆虫食草动物谱系为食的乳草植物中，共同进化的相互作用已经得到了很好的研究，并阐明了新的机制。研究人员最近发现了一种含有高水平拉布福明的乳草，这是一种剧毒的卡宾内酯，形式不同寻常(含有一个含氮(N)和硫的环(S))，这很可能是一种升级的昂贵防御形式。目前的工作试图在马利筋的系统发育中研究更多的含N的隆起内酯，并试图破译它们在独立草食动物谱系中的生理影响和解毒作用。植物-食草动物谱系中不断进化的进攻-防御机制可能会揭示共同进化的共性。三个目标将涉及共同进化的两个方面：1)确定两种专门的乳草昆虫--帝王毛虫和种子昆虫--加工、解毒和储存含N的香豆内酯防御系统的机制。这将使用分离的化合物进行喂养试验、生理测试和行为研究，既使用专家，也使用CRISPR编辑的果蝇，这些果蝇的基因替代了甘露糖耐受性。2)利用数量遗传学，结合饲喂和化学分析，评估在含有和不含含N的隆起内酯的乳草植株上进行解毒和封存的成本和效益。遗传相关性将揭示与差异隔离相关的生活史成本和特征。3)研究整个系统发育过程中黄连属植物含氮防御进化的模式，检验有关氮限制、防御分配以及卡地那内酯浓度和效力之间的权衡的假设。一种假说是，后来分叉的乳草进化出了更有效但浓度较低的卡登内酯，这一假说由理论预测，并由系统发育模式提出，但从未经过严格的检验。总体而言，这些目标将通过表征新的植物防御和专业草食动物的逆适应的生理生态，解毒和隔离毒素的遗传变异，以及宏观进化规模的植物防御进化，来促进对共同进化机制的理解。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Insects that feed on plants are thought to impose natural selection for chemical defenses in the plant tissues they eat. The research being conducted here is on milkweed plants, which contains a highly potent toxin, labriformin, which has evolved to high concentrations in in several species. When highly expressed, labriformin inhibits an essential transport enzyme (the sodium-potassium pump) of the monarch butterfly and the large milkweed bug. These specialized insect pests biochemically modify labriformin to less toxic compounds and sequester these-products for their own defense. The monarch and seed bug’s highly tolerant sodium pump and biochemical modifications point to labriformin being at the center of reciprocal evolution between milkweed and its herbivores. This work serves to unravel the mechanisms of the interaction through physiological and evolutionary analyses. The research also serves as a model for understanding both natural and agricultural interactions between plants and pests. Finally, the milkweed - monarch system lends itself well to public engagement because of its iconic stature, declining populations of conservation concern, and use in school curricula. As part of this project, the researchers are developing modules for K-12 schools and engaging in public outreach through presentations and blogs. Among milkweed plants which are fed upon by several specialized lineages of insect herbivores, coevolutionary interactions have been well-studied and novel mechanisms have been elucidated. Researchers recently discovered a milkweed that contains high levels of labriformin, a highly toxic cardenolide which is unusual in form (containing a ring with nitrogen (N) and sulfur (S)), which could well be an escalated form of costly defense. The current work seeks to investigate additional N-containing cardenolides across the milkweed phylogeny and seek to decipher their physiological impacts and detoxification in independent herbivores lineages. Repeatedly evolving offense-defense mechanisms in plant-herbivore lineages will likely reveal generalities of coevolution. Three objectives will address both sides of coevolution: 1) Determine the mechanisms by which N-containing cardenolide defenses are processed, detoxified, and stored by two specialist milkweed insects, monarch caterpillars and seed bugs. This will employ feeding trials with isolated compounds, physiological assays, and studies of behavior, using both specialists as well as CRISPR-edited Drosophila that have genetic substitutions for tolerating cardenolides. 2) Assess the costs and benefits of detoxification and sequestration using quantitative genetics coupled with feeding and chemical assays of monarchs and seed bugs on milkweed plants with and without N-containing cardenolides. Genetic correlations will reveal life-history costs and traits associated with differential sequestration. 3) Examine the pattern of N-containing defense evolution among milkweeds across the phylogeny, testing hypotheses about N-limitation, defense allocation, and trade-offs between cardenolide concentration and potency. One hypothesis, that later diverging milkweeds have evolved more potent but less concentrated cardenolides, was predicted by theory and suggested by a phylogenetic pattern, but never rigorously tested. Overall these objectives will advance understanding of coevolutionary mechanisms by characterizing novel plant defenses and the physiological ecology of counter-adaptation in specialist herbivores, genetic variation in detoxification and sequestration of toxins, and plant defense evolution at the macroevolutionary scale.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.