Historic exposure to herbivores, not constitutive traits, explains plant tolerance to herbivory in the case of two Medicago species (Fabaceae).

Mechanisms that allow plants to survive and reproduce after herbivory are considered to play a key role in plant evolution. In this study, we evaluated how tolerance varies in species with different historic exposure to herbivores considering ontogeny. We exposed the range-restricted species Medicago citrina and its closely related and widespread species M. arborea to one and two herbivory simulations (80 % aerial biomass loss). Physiological and growth parameters related to tolerance capacity were assessed to evaluate constitutive values (without herbivory) and induced tolerance after damage. Constitutive traits were not always related to greater tolerance, and each species compensated for herbivory through different traits. Herbivory damage only led to mortality in M. citrina; adults exhibited root biomass loss and increased oxidative stress after damage, but also compensated aerial biomass. Despite seedlings showed a lower death percentage than adults after herbivory in M. citrina, they showed less capacity to recover control values than adults. Moderate tolerance to M. arborea herbivory and low tolerance to M. citrina is found. Thus, although the constitutive characteristics are maintained in the lineage, the tolerance of plants decreases in M. citrina. That represents how plants respond to the lack of pressure from herbivores in their habitat.

Transcriptomic and metabolomic analyses identify a role for chlorophyll catabolism and phytoalexin during Medicago nonhost resistance against Asian soybean rust.

Asian soybean rust (ASR) caused by Phakopsora pachyrhizi is a devastating foliar disease affecting soybean production worldwide. Understanding nonhost resistance against ASR may provide an avenue to engineer soybean to confer durable resistance against ASR. We characterized a Medicago truncatula-ASR pathosystem to study molecular mechanisms of nonhost resistance. Although urediniospores formed appressoria and penetrated into epidermal cells of M. truncatula, P. pachyrhizi failed to sporulate. Transcriptomic analysis revealed the induction of phenylpropanoid, flavonoid and isoflavonoid metabolic pathway genes involved in the production of phytoalexin medicarpin in M. truncatula upon infection with P. pachyrhizi. Furthermore, genes involved in chlorophyll catabolism were induced during nonhost resistance. We further characterized one of the chlorophyll catabolism genes, Stay-green (SGR), and demonstrated that the M. truncatula sgr mutant and alfalfa SGR-RNAi lines showed hypersensitive-response-like enhanced cell death upon inoculation with P. pachyrhizi. Consistent with transcriptomic analysis, metabolomic analysis also revealed the accumulation of medicarpin and its intermediate metabolites. In vitro assay showed that medicarpin inhibited urediniospore germination and differentiation. In addition, several triterpenoid saponin glycosides accumulated in M. truncatula upon inoculation with P. pachyrhizi. In summary, using multi-omic approaches, we identified a correlation between phytoalexin production and M. truncatula defense responses against ASR.

Specific domains of plant defensins differentially disrupt colony initiation, cell fusion and calcium homeostasis in Neurospora crassa.

MsDef1 and MtDef4 from Medicago spp. are small cysteine-rich defensins with potent antifungal activity against a broad range of filamentous fungi. Each defensin has a hallmark γ-core motif (GXCX(3-9) C), which contains major determinants of its antifungal activity. In this study, the antifungal activities of MsDef1, MtDef4, and peptides derived from their γ-core motifs, were characterized during colony initiation in the fungal model, Neurospora crassa. These defensins and their cognate peptides inhibited conidial germination and accompanying cell fusion with different potencies. The inhibitory effects of MsDef1 were strongly mediated by the plasma membrane localized sphingolipid glucosylceramide. Cell fusion was selectively inhibited by the hexapeptide RGFRRR derived from the γ-core motif of MtDef4. Fluorescent labelling of this hexapeptide showed that it strongly bound to the germ tube plasma membrane/cell wall. Using N. crassa expressing the Ca(2+) reporter aequorin, MsDef1, MtDef4 and their cognate peptides were each shown to perturb Ca(2+) homeostasis in specific and distinct ways, and the disruptive effects of MsDef1 on Ca(2+) were mediated by glucosylceramide. Together, our results demonstrate that MsDef1 and MtDef4 differ markedly in their antifungal properties and specific domains within their γ-core motifs play important roles in their different modes of antifungal action.