From root to shoot: quantifying nematode tolerance in Arabidopsis thaliana by high-throughput phenotyping of plant development.

Nematode migration, feeding site formation, withdrawal of plant assimilates, and activation of plant defence responses have a significant impact on plant growth and development. Plants display intraspecific variation in tolerance limits for root-feeding nematodes. Although disease tolerance has been recognized as a distinct trait in biotic interactions of mainly crops, we lack mechanistic insights. Progress is hampered by difficulties in quantification and laborious screening methods. We turned to the model plant Arabidopsis thaliana, since it offers extensive resources to study the molecular and cellular mechanisms underlying nematode-plant interactions. Through imaging of tolerance-related parameters, the green canopy area was identified as an accessible and robust measure for assessing damage due to cyst nematode infection. Subsequently, a high-throughput phenotyping platform simultaneously measuring the green canopy area growth of 960 A. thaliana plants was developed. This platform can accurately measure cyst nematode and root-knot nematode tolerance limits in A. thaliana through classical modelling approaches. Furthermore, real-time monitoring provided data for a novel view of tolerance, identifying a compensatory growth response. These findings show that our phenotyping platform will enable a new mechanistic understanding of tolerance to below-ground biotic stress.

The root-knot nematode effector MiMSP32 targets host 12-oxophytodienoate reductase 2 to regulate plant susceptibility.

To establish persistent infections in host plants, herbivorous invaders, such as root-knot nematodes, must rely on effectors for suppressing damage-induced jasmonate-dependent host defenses. However, at present, the effector mechanisms targeting the biosynthesis of biologically active jasmonates to avoid adverse host responses are unknown. Using yeast two-hybrid, in planta co-immunoprecipitation, and mutant analyses, we identified 12-oxophytodienoate reductase 2 (OPR2) as an important host target of the stylet-secreted effector MiMSP32 of the root-knot nematode Meloidogyne incognita. MiMSP32 has no informative sequence similarities with other functionally annotated genes but was selected for the discovery of novel effector mechanisms based on evidence of positive, diversifying selection. OPR2 catalyzes the conversion of a derivative of 12-oxophytodienoate to jasmonic acid (JA) and operates parallel to 12-oxophytodienoate reductase 3 (OPR3), which controls the main pathway in the biosynthesis of jasmonates. We show that MiMSP32 targets OPR2 to promote parasitism of M. incognita in host plants independent of OPR3-mediated JA biosynthesis. Artificially manipulating the conversion of the 12-oxophytodienoate by OPRs increases susceptibility to multiple unrelated plant invaders. Our study is the first to shed light on a novel effector mechanism targeting this process to regulate the susceptibility of host plants.

Comparative Transcriptome Analysis Reveals the Specific Activation of Defense Pathways Against Globodera pallida in Gpa2 Resistant Potato Roots.

Cyst nematodes are considered a dominant threat to yield for a wide range of major food crops. Current control strategies are mainly dependent on crop rotation and the use of resistant cultivars. Various crops exhibit single dominant resistance (R) genes that are able to activate effective host-specific resistance to certain cyst nematode species and/or populations. An example is the potato R gene Gpa2, which confers resistance against the potato cyst nematode (PCN), Globodera pallida population D383. Activation of Gpa2 results in a delayed resistance response, which is characterized by a layer of necrotic cells formed around the developing nematode feeding structure. However, knowledge about the Gpa2-induced defense pathways is still lacking. Here, we uncover the transcriptional changes and gene expression network induced upon Gpa2 activation in potato roots infected with G. pallida. To this end, in vitro-grown Gpa2-resistant potato roots were infected with the avirulent population D383 and virulent population Rookmaker. Infected root segments were harvested at 3 and 6 dpi and sent for RNA sequencing. Comparative transcriptomics revealed a total of 1,743 differentially expressed genes (DEGs) upon nematode infection, of which 559 DEGs were specifically regulated in response to D383 infection. D383-specific DEGs associated with Gpa2-mediated defense mainly relates to calcium-binding activity, salicylic acid (SA) biosynthesis, and systemic acquired resistance (SAR). These data reveal that cyst nematode resistance in potato roots depends on conserved downstream signaling pathways involved in plant immunity, which are also known to contribute to R genes-mediated resistance against other pathogens with different lifestyles.

The TIR-NB-LRR pair DSC1 and WRKY19 contributes to basal immunity of Arabidopsis to the root-knot nematode Meloidogyne incognita.

BACKGROUND: Root-knot nematodes transform vascular host cells into permanent feeding structures to withdraw nutrients from the host plant. Ecotypes of Arabidopsis thaliana can display large quantitative variation in susceptibility to the root-knot nematode Meloidogyne incognita, which is thought to be independent of dominant major resistance genes. However, in an earlier genome-wide association study of the interaction between Arabidopsis and M. incognita we identified a quantitative trait locus harboring homologs of dominant resistance genes but with minor effect on susceptibility to the M. incognita population tested. RESULTS: Here, we report on the characterization of two of these genes encoding the TIR-NB-LRR immune receptor DSC1 (DOMINANT SUPPRESSOR OF Camta 3 NUMBER 1) and the TIR-NB-LRR-WRKY-MAPx protein WRKY19 in nematode-infected Arabidopsis roots. Nematode infection studies and whole transcriptome analyses using the Arabidopsis mutants showed that DSC1 and WRKY19 co-regulate susceptibility of Arabidopsis to M. incognita. CONCLUSION: Given the head-to-head orientation of DSC1 and WRKY19 in the Arabidopsis genome our data suggests that both genes may function as a TIR-NB-LRR immune receptor pair. Unlike other TIR-NB-LRR pairs involved in dominant disease resistance in plants, DSC1 and WRKY19 most likely regulate basal levels of immunity to root-knot nematodes.

Genetic architecture of plant stress resistance: multi-trait genome-wide association mapping.

Plants are exposed to combinations of various biotic and abiotic stresses, but stress responses are usually investigated for single stresses only. Here, we investigated the genetic architecture underlying plant responses to 11 single stresses and several of their combinations by phenotyping 350 Arabidopsis thaliana accessions. A set of 214 000 single nucleotide polymorphisms (SNPs) was screened for marker-trait associations in genome-wide association (GWA) analyses using tailored multi-trait mixed models. Stress responses that share phytohormonal signaling pathways also share genetic architecture underlying these responses. After removing the effects of general robustness, for the 30 most significant SNPs, average quantitative trait locus (QTL) effect sizes were larger for dual stresses than for single stresses. Plants appear to deploy broad-spectrum defensive mechanisms influencing multiple traits in response to combined stresses. Association analyses identified QTLs with contrasting and with similar responses to biotic vs abiotic stresses, and below-ground vs above-ground stresses. Our approach allowed for an unprecedented comprehensive genetic analysis of how plants deal with a wide spectrum of stress conditions.

Apoplastic venom allergen-like proteins of cyst nematodes modulate the activation of basal plant innate immunity by cell surface receptors.

Despite causing considerable damage to host tissue during the onset of parasitism, nematodes establish remarkably persistent infections in both animals and plants. It is thought that an elaborate repertoire of effector proteins in nematode secretions suppresses damage-triggered immune responses of the host. However, the nature and mode of action of most immunomodulatory compounds in nematode secretions are not well understood. Here, we show that venom allergen-like proteins of plant-parasitic nematodes selectively suppress host immunity mediated by surface-localized immune receptors. Venom allergen-like proteins are uniquely conserved in secretions of all animal- and plant-parasitic nematodes studied to date, but their role during the onset of parasitism has thus far remained elusive. Knocking-down the expression of the venom allergen-like protein Gr-VAP1 severely hampered the infectivity of the potato cyst nematode Globodera rostochiensis. By contrast, heterologous expression of Gr-VAP1 and two other venom allergen-like proteins from the beet cyst nematode Heterodera schachtii in plants resulted in the loss of basal immunity to multiple unrelated pathogens. The modulation of basal immunity by ectopic venom allergen-like proteins in Arabidopsis thaliana involved extracellular protease-based host defenses and non-photochemical quenching in chloroplasts. Non-photochemical quenching regulates the initiation of the defense-related programmed cell death, the onset of which was commonly suppressed by venom allergen-like proteins from G. rostochiensis, H. schachtii, and the root-knot nematode Meloidogyne incognita. Surprisingly, these venom allergen-like proteins only affected the programmed cell death mediated by surface-localized immune receptors. Furthermore, the delivery of venom allergen-like proteins into host tissue coincides with the enzymatic breakdown of plant cell walls by migratory nematodes. We, therefore, conclude that parasitic nematodes most likely utilize venom allergen-like proteins to suppress the activation of defenses by immunogenic breakdown products in damaged host tissue.