Summary: DNA methylation is antagonistically controlled by DNA methyltransferases and DNA demethylases. The level of DNA methylation controls plant gene expression on a global level. We have examined impacts of global changes in DNA methylation on the Arabidopsis immune system. A range of hypo‐methylated mutants displayed enhanced resistance to the biotrophic pathogen Hyaloperonospora arabidopsidis (Hpa), whereas two hyper‐methylated mutants were more susceptible to this pathogen. Subsequent characterization of the hypo‐methylated nrpe1 mutant, which is impaired in RNA‐directed DNA methylation, and the hyper‐methylated ros1 mutant, which is affected in DNA demethylation, revealed that their opposite resistance phenotypes are associated with changes in cell wall defence and salicylic acid (SA)‐dependent gene expression. Against infection by the necrotrophic pathogen Plectosphaerella cucumerina, nrpe1 showed enhanced susceptibility, which was associated with repressed sensitivity of jasmonic acid (JA)‐inducible gene expression. Conversely, ros1 displayed enhanced resistance to necrotrophic pathogens, which was not associated with increased responsiveness of JA‐inducible gene expression. Although nrpe1 and ros1 were unaffected in systemic acquired resistance to Hpa, they failed to develop transgenerational acquired resistance against this pathogen. Global transcriptome analysis of nrpe1 and ros1 at multiple time‐points after Hpa infection revealed that 49% of the pathogenesis‐related transcriptome is influenced by NRPE1‐ and ROS1‐controlled DNA methylation. Of the 166 defence‐related genes displaying augmented induction in nrpe1 and repressed induction in ros1, only 25 genes were associated with a nearby transposable element and NRPE1‐ and/or ROS1‐controlled DNA methylation. Accordingly, we propose that the majority of NRPE1‐ and ROS1‐dependent defence genes are regulated in trans by DNA methylation.

RNA–directed DNA methylation (RdDM) is an epigenetic control mechanism driven by small interfering RNAs (siRNAs) that influence gene function. In plants, little is known of the involvement of the RdDM pathway in regulating traits related to immune responses. In a genetic screen designed to reveal factors regulating immunity in Arabidopsis thaliana, we identified NRPD2 as the OVEREXPRESSOR OF CATIONIC PEROXIDASE 1 (OCP1). NRPD2 encodes the second largest subunit of the plant-specific RNA Polymerases IV and V (Pol IV and Pol V), which are crucial for the RdDM pathway. The ocp1 and nrpd2 mutants showed increases in disease susceptibility when confronted with the necrotrophic fungal pathogens Botrytis cinerea and Plectosphaerella cucumerina. Studies were extended to other mutants affected in different steps of the RdDM pathway, such as nrpd1, nrpe1, ago4, drd1, rdr2, and drm1drm2 mutants. Our results indicate that all the mutants studied, with the exception of nrpd1, phenocopy the nrpd2 mutants; and they suggest that, while Pol V complex is required for plant immunity, Pol IV appears dispensable. Moreover, Pol V defective mutants, but not Pol IV mutants, show enhanced disease resistance towards the bacterial pathogen Pseudomonas syringae DC3000. Interestingly, salicylic acid (SA)–mediated defenses effective against PsDC3000 are enhanced in Pol V defective mutants, whereas jasmonic acid (JA)–mediated defenses that protect against fungi are reduced. Chromatin immunoprecipitation analysis revealed that, through differential histone modifications, SA–related defense genes are poised for enhanced activation in Pol V defective mutants and provide clues for understanding the regulation of gene priming during defense. Our results highlight the importance of epigenetic control as an additional layer of complexity in the regulation of plant immunity and point towards multiple components of the RdDM pathway being involved in plant immunity based on genetic evidence, but whether this is a direct or indirect effect on disease-related genes is unclear. The influence of epigenetic regulation in controlling the adaptive responses of living organisms to changes in the environment is becoming a common theme in biology. RNA–directed DNA methylation (RdDM) is an epigenetic control mechanism driven by a subset of noncoding small interfering RNAs (siRNAs) that influence gene function without changing DNA sequence by inducing de novo methylation of cytosines, or by modification of histones, at their target genomic regions. The implication and roles of the RdDM mechanism in the orchestration of plant immune responses still remains to be characterized. A recent study in the model plant Arabidopsis showed that ARGONAUTE4, one of the characteristic components of the RdDM pathway, was required for plant immunity against bacterial pathogens. Here, in a genetic screen aiming to identify cellular factors integral in regulating immunity in Arabidopsis, we further identified that the RNA polymerases V, another crucial component of the RdDM pathway, is pivotal for plant immunity against fungal pathogens. Similarly, we identified that additional components of the RdDM pathway, but surprisingly not RNA polymerase IV, are similarly required for plant immunity. Based on genetic evidence, our results highlight the importance of RdDM as an additional layer of complexity in the regulation of plant immune responses.

As primary producers, plants are under constant pressure to defend themselves against potentially deadly pathogens and herbivores. In this review, we describe short- and long-term strategies that enable plants to cope with these stresses. Apart from internal immunological strategies that involve physiological and (epi)genetic modifications at the cellular level, plants also employ external strategies that rely on recruitment of beneficial organisms. We discuss these strategies along a gradient of increasing timescales, ranging from rapid immune responses that are initiated within seconds to (epi)genetic adaptations that occur over multiple plant generations. We cover the latest insights into the mechanistic and evolutionary underpinnings of these strategies and present explanatory models. Finally, we discuss how knowledge from short-lived model species can be translated to economically and ecologically important perennials to exploit adaptive plant strategies and mitigate future impacts of pests and diseases in an increasingly interconnected and changing world.

β-Aminobutyric acid (BABA) is a small-molecule activator of plant disease resistance. Binding of (R)-BABA to IBI1—an aspartyl-tRNA synthetase (AspRS)—primes noncanonical defense pathways, resulting in broad-spectrum disease resistance. Ligand binding also blocks AspRS activity, leading to accumulation of aspartic acid and uncharged tRNA, which activate a second plant stress response. Specific chemicals can prime the plant immune system for augmented defense. β-aminobutyric acid (BABA) is a priming agent that provides broad-spectrum disease protection. However, BABA also suppresses plant growth when applied in high doses, which has hampered its application as a crop defense activator. Here we describe a mutant of Arabidopsis thaliana that is impaired in BABA-induced disease immunity (ibi1) but is hypersensitive to BABA-induced growth repression. IBI1 encodes an aspartyl-tRNA synthetase. Enantiomer-specific binding of the R enantiomer of BABA to IBI1 primed the protein for noncanonical defense signaling in the cytoplasm after pathogen attack. This priming was associated with aspartic acid accumulation and tRNA-induced phosphorylation of translation initiation factor eIF2α. However, mutation of eIF2α-phosphorylating GCN2 kinase did not affect BABA-induced immunity but relieved BABA-induced growth repression. Hence, BABA-activated IBI1 controls plant immunity and growth via separate pathways. Our results open new opportunities to separate broad-spectrum disease resistance from the associated costs on plant growth.

In higher eukaryotes, induced resistance associates with acquisition of a priming state of the cells for a more effective activation of innate immunity; however, the nature of the components for mounting this type of immunological memory is not well known. We identified an extracellular subtilase from Arabidopsis, SBT3.3, the overexpression of which enhances innate immune responses while the loss of function compromises them. SBT3.3 expression initiates a durable autoinduction mechanism that promotes chromatin remodeling and activates a salicylic acid(SA)-dependent mechanism of priming of defense genes for amplified response. Moreover, SBT3.3 expression-sensitized plants for enhanced expression of the OXI1 kinase gene and activation of MAP kinases following pathogen attack, providing additional clues for the regulation of immune priming by SBT3.3. Conversely, in sbt3.3 mutant plants pathogen-mediated induction of SA-related defense gene expression is drastically reduced and activation of MAP kinases inhibited. Moreover, chromatin remodeling of defense-related genes normally associated with activation of an immune priming response appear inhibited in sbt3.3 plants, further indicating the importance of the extracellular SBT3.3 subtilase in the establishment of immune priming. Our results also point to an epigenetic control in the regulation of plant immunity, since SBT3.3 is up-regulated and priming activated when epigenetic control is impeded. SBT3.3 represents a new regulator of primed immunity. Following a first encounter with a pathogen, higher eukaryotes develop enhanced resistance to subsequent infections by a broad spectrum of pathogens. This type of induced resistance (IR) exhibits memory characteristics after the first encounter with a pathogen —training effect— and appears evolutionarily conserved. IR response components must reside in a plant's capacity to reprogram gene expression. Among the mechanisms involved in immune-related transcriptional reprogramming, the importance of chromatin remodelling is emerging. Recent studies indicated a causal link between priming and chromatin remodelling, pointing to a histone memory for information storage in the plant immune response. These results emphasized the importance of epigenetic control as an additional layer of complexity in plant immunity. However, the nature of the components for activating this type of immunological memory remains elusive. Here, in a search aiming to identify cellular factors integral in regulating immunity in Arabidopsis, we found that the SBT3.3 gene, encoding an extracellular subtilase enzyme, is pivotal for establishing plant immune priming. Moreover, based on molecular and genetic evidences, our results indicate that SBT3.3 expression is under epigenetic control thus highlighting the importance of this mechanism of gene regulation in the control of plant immunity and IR.