Divergent regulation of auxin responsive genes in root-knot and cyst nematodes feeding sites formed in Arabidopsis.

Cysts (CNs) and root-knot nematodes (RKNs) induce specialized feeding cells, syncytia, and giant cells (GCs), respectively, within plant roots. The plant tissues around the GCs usually by respond forming a root swelling called a gall that contains the GCs. The ontogenesis of feeding cells is different. GC formation is a process of new organogenesis from vascular cells, which are still not well characterized, that differentiate into GCs. In contrast, syncytia formation involves the fusion of adjacent cells that have already differentiated. Nonetheless, both feeding sites show an auxin maximum pertinent to feeding site formation. However, data on the molecular divergences and similarities between the formation of both feeding sites regarding auxin-responsive genes are still scarce. We studied genes from the auxin transduction pathways that are crucial during gall and lateral root (LR) development in the CN interaction by using promoter-reporter (GUS/LUC)transgenic lines, as well as loss of function lines of Arabidopsis. The promoters pGATA23 and several deletions of pmiR390a were active in syncytia, as were in galls, but pAHP6 or putative up-stream regulators as ARF5/7/19 were not active in syncytia. Additionally, none of these genes seemed to play a key role during cyst nematode establishment in Arabidopsis, as the infection rates in loss of function lines did not show significant differences compared to control Col-0 plants. Furthermore, the presence of only canonical AuxRe elements in their proximal promoter regions is highly correlated with their activation in galls/GCs (AHP6, LBD16), but those promoters active in syncytia (miR390, GATA23) carry AuxRe overlapping core cis-elements for other transcription factor families (i.e., bHLH, bZIP). Strikingly, in silico transcriptomic analysis showed very few genes upregulated by auxins common to those induced in GCs and syncytia, despite the high number of upregulated IAA responsive genes in syncytia and galls. The complex regulation of auxin transduction pathways, where different members of the auxin response factor (ARF) family may interact with other factors, and the differences in auxin sensitivity, as indicated by the lower induction of the DR5 sensor in syncytia than galls, among other factors, may explain the divergent regulation of auxin responsive genes in the two types of nematode feeding sites.

Local auxin synthesis mediated by YUCCA4 induced during root-knot nematode infection positively regulates gall growth and nematode development.

Parasites and pathogens are known to manipulate the host's endogenous signaling pathways to facilitate the infection process. In particular, plant-parasitic root-knot nematodes (RKN) are known to elicit auxin response at the infection sites, to aid the development of root galls as feeding sites for the parasites. Here we describe the role of local auxin synthesis induced during RKN infection. Exogenous application of auxin synthesis inhibitors decreased RKN gall formation rates, gall size and auxin response in galls, while auxin and auxin analogues produced the opposite effects, re-enforcing the notion that auxin positively regulates RKN gall formation. Among the auxin biosynthesis enzymes, YUCCA4 (YUC4) was found to be dramatically up-regulated during RKN infection, suggesting it may be a major contributor to the auxin accumulation during gall formation. However, yuc4-1 showed only very transient decrease in gall auxin levels and did not show significant changes in RKN infection rates, implying the loss of YUC4 is likely compensated by other auxin sources. Nevertheless, yuc4-1 plants produced significantly smaller galls with fewer mature females and egg masses, confirming that auxin synthesized by YUC4 is required for proper gall formation and RKN development within. Interestingly, YUC4 promoter was also activated during cyst nematode infection. These lines of evidence imply auxin biosynthesis from multiple sources, one of them being YUC4, is induced upon plant endoparasitic nematode invasion and likely contribute to their infections. The coordination of these different auxins adds another layer of complexity of hormonal regulations during plant parasitic nematode interaction.

A potential role of salicylic acid in the evolutionary behavior of Trichoderma as a plant pathogen: from Marchantia polymorpha to Arabidopsis thaliana.

MAIN CONCLUSION: Recognition of the interaction of Trichoderma during the evolution of land plants plays a potential key role in the development of the salicylic acid defense pathway and the establishment of a mutualistic relationship. Marchantia polymorpha is a common liverwort considered in recent years as a model plant for evolutionary studies on plant-microorganism interactions. Despite the lack of research, remarkable results have been reported regarding the understanding of metabolic and evolutionary processes of beneficial and/or harmful interactions, owing to a better understanding of the origin and evolution of different plant defense pathways. In this study, we have carried out work on the direct and indirect interactions (exudates and volatiles) of M. polymorpha with different species of the fungal genus Trichoderma. These interactions showed different outcomes, including resistance or even growth promotion and disease. We have analyzed the level of tissue colonization and defense-related gene expression. Furthermore, we have used the pteridophyte Dryopteris affinis and the angiosperm Arabidopsis thaliana, as subsequent steps in plant evolution, together with the plant pathogen Rhizoctonia solani as a control of plant pathogenicity. Trichoderma virens, T. brevicompactum and T. hamatum are pathogens of M. polymorpha, while exudates of T. asperellum are harmful to the plant. The analysis of the expression of several defense genes in M. polymorpha and A. thaliana showed that there is a correlation of the transcriptional activation of SA-related genes with resistance or susceptibility of M. polymorpha to Trichoderma. Moreover, exogenous SA provides resistance to the virulent Trichoderma species. This beneficial fungus may have had an evolutionary period of interaction with plants in which it behaved as a plant pathogen until plants developed a defense system to limit its colonization through a defense response mediated by SA.

The DNA methylation landscape of the root-knot nematode-induced pseudo-organ, the gall, in Arabidopsis, is dynamic, contrasting over time, and critically important for successful parasitism.

Root-knot nematodes (RKNs) induce giant cells (GCs) within galls which are characterized by large-scale gene repression at early stages. However, the epigenetic mechanism(s) underlying gene silencing is (are) still poorly characterized. DNA methylation in Arabidopsis galls induced by Meloidogyne javanica was studied at crucial infection stages (3 d post-infection (dpi) and 14 dpi) using enzymatic, cytological, and sequencing approaches. DNA methyltransferase mutants (met1, cmt2, cmt3, cmt2/3, drm1/2, ddc) and a DNA demethylase mutant (ros1), were analyzed for RKN resistance/tolerance, and galls were characterized by confocal microscopy and RNA-seq. Early galls were hypermethylated, and the GCs were found to be the major contributors to this hypermethylation, consistent with the very high degree of gene repression they exhibit. By contrast, medium/late galls showed no global increase in DNA methylation compared to uninfected roots, but exhibited large-scale redistribution of differentially methylated regions (DMRs). In line with these findings, it was also shown that DNA methylation and demethylation mutants showed impaired nematode reproduction and gall/GC-development. Moreover, siRNAs that were exclusively present in early galls accumulated at hypermethylated DMRs, overlapping mostly with retrotransposons in the CHG/CG contexts that might be involved in their repression, contributing to their stability/genome integrity. Promoter/gene methylation correlated with differentially expressed genes encoding proteins with basic cell functions. Both mechanisms are consistent with reprogramming host tissues for gall/GC formation. In conclusion, RNA-directed DNA methylation (RdDM; DRM2/1) pathways, maintenance methyltransferases (MET1/CMT3) and demethylation (ROS1) appear to be prominent mechanisms driving a dynamic regulation of the epigenetic landscape during RKN infection.

Biological Control of Plant-Parasitic Nematodes by Filamentous Fungi Inducers of Resistance: Trichoderma, Mycorrhizal and Endophytic Fungi.

Plant-parasitic-nematodes represent a major threat to the agricultural production of different crops worldwide. Due to the high toxicity of chemical nematicides, it is necessary to develop new control strategies against nematodes. In this respect, filamentous fungi can be an interesting biocontrol alternative. The genus Trichoderma, mycorrhizal and endophytic fungi are the main groups of filamentous fungi studied and used as biological control agents (BCAs) against nematodes as resistance inducers. They are able to reduce the damage caused by plant-parasitic nematodes directly by parasitism, antibiosis, paralysis and by the production of lytic enzymes. But they also minimize harm by space and resource-competition, by providing higher nutrient and water uptake to the plant, or by modifying the root morphology, and/or rhizosphere interactions, that constitutes an advantage for the plant-growth. Besides, filamentous fungi are able to induce resistance against nematodes by activating hormone-mediated (salicylic and jasmonic acid, strigolactones among others) plant-defense mechanisms. Additionally, the alteration of the transport of chemical defense components through the plant or the synthesis of plant secondary metabolites and different enzymes can also contribute to enhancing plant defenses. Therefore, the use of filamentous fungi of the mentioned groups as BCAs is a promising durable biocontrol strategy in agriculture against plant-parasitic nematodes.