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Summary: Arabidopsis Col‐0 RPP2A and RPP2B confer recognition of Arabidopsis downy mildew (Hyaloperonospora arabidopsidis [Hpa]) isolate Cala2, but the identity of the recognized ATR2Cala2 effector was unknown.To reveal ATR2Cala2, an F2 population was generated from a cross between Hpa‐Cala2 and Hpa‐Noks1. We identified ATR2Cala2 as a non‐canonical RxLR‐type effector that carries a signal peptide, a dEER motif, and WY domains but no RxLR motif. Recognition of ATR2Cala2 and its effector function were verified by biolistic bombardment, ectopic expression and Hpa infection.ATR2Cala2 is recognized in accession Col‐0 but not in Ler‐0 in which RPP2A and RPP2B are absent. In ATR2Emoy2 and ATR2Noks1 alleles, a frameshift results in an early stop codon. RPP2A and RPP2B are essential for the recognition of ATR2Cala2. Stable and transient expression of ATR2Cala2 under 35S promoter in Arabidopsis and Nicotiana benthamiana enhances disease susceptibility.Two additional Col‐0 TIR‐NLR (TNL) genes (RPP2C and RPP2D) adjacent to RPP2A and RPP2B are quantitatively required for full resistance to Hpa‐Cala2. We compared RPP2 haplotypes in multiple Arabidopsis accessions and showed that all four genes are present in all ATR2Cala2‐recognizing accessions. [ABSTRACT FROM AUTHOR]

SUMMARY: The transition from vegetative to reproductive development (floral transition) is a costly process in annual plants requiring increased investments in metabolic resources. The Arabidopsis thaliana (Arabidopsis) PHD finger protein EDM2 and RRM domain proteins EDM3 and IBM2 are known to form chromatin‐associated complexes controlling transcript processing. We are reporting that distinct splice isoforms of EDM3 and IBM2 cooperate in the coordination of the floral transition with basal immune responses. These cooperating splice isoforms, termed EDM3L and IBM2L, control the intensity of basal immunity and, via a separate pathway, the timing of the floral transition. During the developmental phase prior to the floral transition expression of EDM3L and IBM2L strongly and gradually increases, while these isoforms simultaneously down‐regulate expression of the floral suppressor gene FLC and promote the transition to reproductive growth. At the same time these accumulating EDM3 and IBM2 splice isoforms gradually suppress basal immunity against the virulent Noco2 isolate of the pathogenic oomycete Hyaloperonospora arabidopsidis and down‐regulate expression of a set of defense‐associated genes and immune receptor genes. We are providing clear evidence for a functional link between the floral transition and basal immunity in the annual plant Arabidopsis. Coordination of these two biological processes, which compete for metabolic resources, is likely critical for plant survival and reproductive success. Significance Statement: We are providing clear evidence for a dynamic mechanism coordinating in Arabidopsis two processes competing for metabolic resources, the transition from vegetative to reproductive growth and basal immunity. As Arabidopsis development approaches the transition to flowering distinct splice‐isoforms of the RRM‐domain proteins EDM3 and IBM2 accumulate and enable the floral transition by gradually down‐regulating the floral suppressor gene FLC, while, via a separate pathway, progressively limiting the extent of basal immune responses. [ABSTRACT FROM AUTHOR]

The NLR-receptor RPP7 mediates race-specific immunity in Arabidopsis. Previous screens for enhanced downy mildew (edm) mutants identified the co-chaperone SGT1b (EDM1) and the PHD-finger protein EDM2 as critical regulators of RPP7. Here, we describe a third edm mutant compromised in RPP7 immunity, edm3. EDM3 encodes a nuclear-localized protein featuring an RNA-recognition motif. Like EDM2, EDM3 promotes histone H3 lysine 9 dimethylation (H3K9me2) at RPP7. Global profiling of H3K9me2 showed EDM3 to affect this silencing mark at a large set of loci. Importantly, both EDM3 and EDM2 co-associate in vivo with H3K9me2-marked chromatin and transcripts at a critical proximal polyadenylation site of RPP7, where they suppress proximal transcript polyadeylation/termination. Our results highlight the complexity of plant NLR gene regulation, and establish a functional and physical link between a histone mark and NLR-transcript processing.

White mold is caused by the fungal pathogen Sclerotinia sclerotiorum and leads to rapid and significant loss in plant yield. Among its many brassicaceous hosts, including Brassica napus (canola) and Arabidopsis, the response of individual tissue layers directly at the site of infection has yet to be explored. Using laser microdissection coupled with RNA sequencing, we profiled the epidermis, mesophyll, and vascular leaf tissue layers of B. napus in response to S. sclerotiorum. High-throughput tissue-specific mRNA sequencing increased the total number of detected transcripts compared with whole-leaf assessments and provided novel insight into the conserved and specific roles of ontogenetically distinct leaf tissue layers in response to infection. When subjected to pathogen infection, the epidermis, mesophyll, and vasculature activate both specific and shared gene sets. Putative defense genes identified through transcription factor network analysis were then screened for susceptibility against necrotrophic, hemi-biotrophic, and biotrophic pathogens. Arabidopsis deficient in PR5-like RECEPTOR KINASE (PR5K) mRNA levels were universally susceptible to all pathogens tested and were further characterized to identify putative interacting partners involved in the PR5K signaling pathway. Together, these data provide insight into the complexity of the plant defense response directly at the site of infection. [ABSTRACT FROM AUTHOR]

SUMMARY: Translating ribosome affinity purification (TRAP) utilizes transgenic plants expressing a ribosomal protein fused to a tag for affinity co‐purification of ribosomes and the mRNAs that they are translating. This population of actively translated mRNAs (translatome) can be interrogated by quantitative PCR or RNA sequencing. Condition‐ or cell‐specific promoters can be utilized to isolate the translatome of specific cell types, at different growth stages and/or in response to environmental variables. While advantageous for revealing differential expression, this approach may not provide sufficient sensitivity when activity of the condition/cell‐specific promoter is weak, when ribosome turnover is low in the cells of interest, or when the targeted cells are ephemeral. In these situations, expressing tagged ribosomes under the control of these specific promoters may not yield sufficient polysomes for downstream analysis. Here, we describe a new TRAP system that employs two transgenes: One is constitutively expressed and encodes a ribosomal protein fused to one fragment of a split green fluorescent protein (GFP); the second is controlled by a stimulus‐specific promoter and encodes the second GFP fragment fused to an affinity purification tag. In cells where both transgenes are active, the purification tag is attached to ribosomes by bi‐molecular folding and assembly of the split GFP fragments. This approach provides increased sensitivity and better temporal resolution because it labels pre‐existing ribosomes and does not depend on rapid ribosome turnover. We describe the optimization and key parameters of this system, and then apply it to a plant–pathogen interaction in which spatial and temporal resolution are difficult to achieve with current technologies. Significance Statement: Translating ribosome affinity purification (TRAP) has been modified to allow with increased sensitivity the isolation of RNA from sets of cells in which the activity of condition/cellspecific promoters is weak, ribosome turnover is low, or cells whose nature is ephemeral. Based on the use of a split linker constituted of the GFP driven by a pathogen‐inducible promoter, this new TRAP system enabled efficient isolation of translated RNA from pathogen‐infected leaf cells. [ABSTRACT FROM AUTHOR]

Arabidopsis Col-0 RPP2A and RPP2B confer recognition of Arabidopsis downy mildew (Hyaloperonospora arabidopsidis [Hpa]) isolate Cala2, but the identity of the recognized ATR2 Cala2 effector was unknown. To reveal ATR2 Cala2 , an F 2 population was generated from a cross between Hpa-Cala2 and Hpa-Noks1. We identified ATR2 Cala2 as a non-canonical RxLR-type effector that carries a signal peptide, a dEER motif, and WY domains but no RxLR motif. Recognition of ATR2 Cala2 and its effector function were verified by biolistic bombardment, ectopic expression and Hpa infection. ATR2 Cala2 is recognized in accession Col-0 but not in Ler-0 in which RPP2A and RPP2B are absent. In ATR2 Emoy2 and ATR2 Noks1 alleles, a frameshift results in an early stop codon. RPP2A and RPP2B are essential for the recognition of ATR2 Cala2 . Stable and transient expression of ATR2 Cala2 under 35S promoter in Arabidopsis and Nicotiana benthamiana enhances disease susceptibility. Two additional Col-0 TIR-NLR (TNL) genes (RPP2C and RPP2D) adjacent to RPP2A and RPP2B are quantitatively required for full resistance to Hpa-Cala2. We compared RPP2 haplotypes in multiple Arabidopsis accessions and showed that all four genes are present in all ATR2 Cala2 -recognizing accessions., (© 2024 The Authors. New Phytologist © 2024 New Phytologist Foundation.)

Plant genes differentially expressed during plant-pathogen interactions can be important for host immunity or can contribute to pathogen virulence. Large-scale transcript profiling studies, such as microarray- or mRNA-seq-based analyses, have revealed hundreds of genes that are differentially expressed during plant-pathogen interactions. However, transcriptional responses limited to a small number of cells at infection sites can be difficult to detect using these approaches, as they are under-represented in the whole-tissue datasets typically generated by such methods. This study examines the interactions between Arabidopsis thaliana (Arabidopsis) and the pathogenic oomycete Hyaloperonospora arabidopsidis (Hpa) by enhancer trapping to uncover novel plant genes involved in local infection responses. We screened a β-glucuronidase (GUS) reporter-based enhancer-trap population for expression patterns related to Hpa infection. Several independent lines exhibited GUS expression in leaf mesophyll cells surrounding Hpa structures, indicating a regulatory response to pathogen infection. One of these lines contained a single enhancer-trap insertion in an exon of At1g08800 (MyoB1, Myosin Binding Protein 1) and was subsequently found to exhibit reduced susceptibility to Hpa. Two additional Arabidopsis lines with T-DNA insertions in exons of MyoB1 also exhibited approximately 30% fewer spores than wild-type plants. This study demonstrates that our enhancer-trapping strategy can result in the identification of functionally relevant pathogen-responsive genes. Our results further suggest that MyoB1 either positively contributes to Hpa virulence or negatively affects host immunity against this pathogen.

Summary: The oomycete pathogen Hyaloperonospora arabidopsidis delivers diverse effector proteins into host plant cells to suppress the plant's innate immunity. In this study, we investigate the mechanism of action of a conserved RxLR effector, HaRxLL470, in suppressing plant immunity.Genomic, molecular and biochemical analyses were performed to investigate the function of HaRxLL470 and the mechanism of the interaction between HaRxLL470 and the target host protein during H. arabidopsidis infection.We report that HaRxLL470 enhances plant susceptibility to H. arabidopsidis isolate Noco2 by interacting with the host photomorphogenesis regulator protein HY5. Our results demonstrate that HY5 is not only an important component in the regulation of light signalling, but also positively regulates host plant immunity against H. arabidopsidis by transcriptional activation of defense‐related genes. We show that the interaction between HaRxLL470 and HY5 compromises the function of HY5 as a transcription factor by attenuating its DNA‐binding activity.The present study demonstrates that HY5 positively regulates host plant defense against H. arabidopsidis whereas HaRxLL470, a conserved RxLR effector across oomycete pathogens, enhances pathogenicity by interacting with HY5 and suppressing transcriptional activation of defense‐related genes. [ABSTRACT FROM AUTHOR]

Summary: Pattern‐triggered immunity (PTI) is activated in plants upon recognition by pattern recognition receptors (PRRs) of damage‐ and microbe‐associated molecular patterns (DAMPs and MAMPs) derived from plants or microorganisms, respectively. To understand better the plant mechanisms involved in the perception of carbohydrate‐based structures recognized as DAMPs/MAMPs, we have studied the ability of mixed‐linked β‐1,3/1,4‐glucans (MLGs), present in some plant and microbial cell walls, to trigger immune responses and disease resistance in plants. A range of MLG structures were tested for their capacity to induce PTI hallmarks, such as cytoplasmic Ca2+ elevations, reactive oxygen species production, phosphorylation of mitogen‐activated protein kinases and gene transcriptional reprogramming. These analyses revealed that MLG oligosaccharides are perceived by Arabidopsis thaliana and identified a trisaccharide, β‐d‐cellobiosyl‐(1,3)‐β‐d‐glucose (MLG43), as the smallest MLG structure triggering strong PTI responses. These MLG43‐mediated PTI responses are partially dependent on LysM PRRs CERK1, LYK4 and LYK5, as they were weaker in cerk1 and lyk4 lyk5 mutants than in wild‐type plants. Cross‐elicitation experiments between MLG43 and the carbohydrate MAMP chitohexaose [β‐1,4‐d‐(GlcNAc)6], which is also perceived by these LysM PRRs, indicated that the mechanism of MLG43 recognition could differ from that of chitohexaose, which is fully impaired in cerk1 and lyk4 lyk5 plants. MLG43 treatment confers enhanced disease resistance in A. thaliana to the oomycete Hyaloperonospora arabidopsidis and in tomato and pepper to different bacterial and fungal pathogens. Our data support the classification of MLGs as a group of carbohydrate‐based molecular patterns that are perceived by plants and trigger immune responses and disease resistance. Significance Statement: A better characterization of plant mechanisms involved in the perception of carbohydrate‐based structures recognized as damage‐/microbe‐associated molecular patterns is needed to understand plant immunity further. We show here that in addition to the β‐1,3‐ and β‐1,4‐glucans damage‐/microbe‐associated molecular patterns, plants are able to perceive mixed‐linked β‐1,3/1,4‐glucans that are present in the cell walls of some plant species and microbes, such as the oomycete Hyaloperonospora arabidopsidis. Mixed‐linked β‐1,3/1,4‐glucans trigger immune responses and disease resistance against different pathogens in several plant species. [ABSTRACT FROM AUTHOR]

Iron (Fe) is a poorly available mineral nutrient which affects the outcome of many cross-kingdom interactions. In Arabidopsis thaliana , Fe starvation limits infection by necrotrophic pathogens. Here, we report that Fe deficiency also reduces disease caused by the hemi-biotrophic bacterium Pseudomonas syringae and the biotrophic oomycete Hyaloperonospora arabidopsidis , indicating that Fe deficiency-induced resistance is effective against pathogens with different lifestyles. Furthermore, we show that Fe deficiency-induced resistance is not caused by withholding Fe from the pathogen but is a plant-mediated defense response that requires activity of ethylene and salicylic acid. Because rhizobacteria-induced systemic resistance (ISR) is associated with a transient up-regulation of the Fe deficiency response, we tested whether Fe deficiency-induced resistance and ISR are similarly regulated. However, Fe deficiency-induced resistance functions independently of the ISR regulators MYB72 and BGLU42, indicating that both types of induced resistance are regulated in a different manner. Mutants opt3 and frd1 , which display misregulated Fe homeostasis under Fe-sufficient conditions, show disease resistance levels comparable with those of Fe-starved wild-type plants. Our results suggest that disturbance of Fe homeostasis, through Fe starvation stress or other non-homeostatic conditions, is sufficient to prime the plant immune system for enhanced defense. [ABSTRACT FROM AUTHOR]

Our understanding of obligate biotrophic pathogens is limited by lack of knowledge concerning the molecular function of virulence factors. We established Arabidopsis host-induced gene silencing (HIGS) to explore gene functions of Hyaloperonospora arabidopsidis , including CYSTEINE-RICH PROTEIN (HaCR) 1 , a potential secreted effector gene of this obligate biotrophic pathogen. Ha CR1 HIGS resulted in H. arabidopsidis -induced local plant cell death and reduced pathogen reproduction. We functionally characterized Ha CR1 by ectopic expression in Nicotiana benthamiana. Ha CR1 was capable of inhibiting effector-triggered plant cell death. Consistent with this, Ha CR1 expression in N. benthamiana led to stronger disease symptoms caused by the hemibiotrophic oomycete pathogen Phytophthora capsici , but reduced disease symptoms caused by the necrotrophic fungal pathogen Botrytis cinerea. Expressing Ha CR1 in transgenic Arabidopsis confirmed higher susceptibility to H. arabidopsidis and to the bacterial hemibiotrophic pathogen Pseudomonas syringae. Increased H. arabidopsidis infection was in accordance with reduced PATHOGENESIS RELATED (PR) 1 induction. Expression of full-length Ha CR1 was required for its function, which was lost if the signal peptide was deleted, suggesting its site of action in the plant apoplast. This study provides phytopathological and molecular evidence for the importance of this widespread, but largely unexplored class of non-RxLR effectors in biotrophic oomycetes. [ABSTRACT FROM AUTHOR]

Transposable elements (TEs) can drive evolution by creating genetic and epigenetic variation. Although examples of adaptive TE insertions are accumulating, proof that epigenetic information carried by such "domesticated" TEs has been coopted to control host gene function is still limited. We show that COPIA-R7, a TE inserted into the Arabidopsis thaliana disease resistance gene RPP7 recruited the histone mark H3K9me2 to this locus. H3K9me2 levels at COPIA-R7 affect the choice between two alternative RPP7 polyadenylation sites in the pre-mRNA and, thereby, influence the critical balance between RPP7-coding and non-RPP7-coding transcript isoforms. Function of RPP7 is fully dependent on high levels of H3K9me2 at COPIA-R7. We present a direct in vivo demonstration for cooption of a TE-associated histone mark to the epigenetic control of pre-mRNA processing and establish a unique mechanism for regulation of plant immune surveillance gene expression. Our results functionally link a histone mark to alternative polyadenylation and the balance between distinct transcript isoforms from a single gene.

Summary: The oomycete pathogen Hyaloperonospora arabidopsidis (Hpa) causes downy mildew disease on Arabidopsis. To colonize its host, Hpa translocates effector proteins that suppress plant immunity into infected host cells. Here, we investigate the relevance of the interaction between one of these effectors, HaRxL106, and Arabidopsis RADICAL‐INDUCED CELL DEATH1 (RCD1). We use pathogen infection assays as well as molecular and biochemical analyses to test the hypothesis that HaRxL106 manipulates RCD1 to attenuate transcriptional activation of defense genes. We report that HaRxL106 suppresses transcriptional activation of salicylic acid (SA)‐induced defense genes and alters plant growth responses to light. HaRxL106‐mediated suppression of immunity is abolished in RCD1 loss‐of‐function mutants. We report that RCD1‐type proteins are phosphorylated, and we identified Mut9‐like kinases (MLKs), which function as phosphoregulatory nodes at the level of photoreceptors, as RCD1‐interacting proteins. An mlk1,3,4 triple mutant exhibits stronger SA‐induced defense marker gene expression compared with wild‐type plants, suggesting that MLKs also affect transcriptional regulation of SA signaling. Based on the combined evidence, we hypothesize that nuclear RCD1/MLK complexes act as signaling nodes that integrate information from environmental cues and pathogen sensors, and that the Arabidopsis downy mildew pathogen targets RCD1 to prevent activation of plant immunity. [ABSTRACT FROM AUTHOR]

Plants use a tightly regulated immune system to fight off various pathogens. Phospholipase D (PLD) and its product, phosphatidic acid, have been shown to influence plant immunity; however, the underlying mechanisms remain unclear. Here, we show that the Arabidopsis mutants pldα1 and pldδ, respectively, exhibited enhanced resistance and enhanced susceptibility to both well-adapted and poorly adapted powdery mildew pathogens, and a virulent oomycete pathogen, indicating that PLDα1 negatively while PLDδ positively modulates post-penetration resistance. The pldα1δ double mutant showed a similar infection phenotype to pldα1, genetically placing PLDα1 downstream of PLDδ. Detailed genetic analyses of pldδ with mutations in genes for salicylic acid (SA) synthesis (SID2) and/or signaling (EDS1 and PAD4), measurement of SA and jasmonic acid (JA) levels, and expression of their respective reporter genes indicate that PLDδ contributes to basal resistance independent of EDS1/PAD4, SA, and JA signaling. Interestingly, while PLDα1-enhanced green fluorescent protein (eGFP) was mainly found in the tonoplast before and after haustorium invasion, PLDδ-eGFP's focal accumulation to the plasma membrane around the fungal penetration site appeared to be suppressed by adapted powdery mildew. Together, our results demonstrate that PLDα1 and PLDδ oppositely modulate basal, post-penetration resistance against powdery mildew through a non-canonical mechanism that is independent of EDS1/PAD4, SA, and JA. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

The hormone salicylic acid (SA) generally induces plant defenses against biotrophic pathogens. Jasmonic acid and its oxylipin derivatives (JAs) together with ethylene (ET) are generally important hormonal regulators of induced plant defenses against necrotrophic pathogens, whereas JAs together with abscisic acid (ABA) are implicated in induced plant defenses against herbivorous insects. Hormonal crosstalk between the different plant defense pathways has often been hypothesized to be a cost-saving strategy that has evolved as a means of the plant to reduce allocation costs by repression of unnecessary defenses, thereby minimizing trade-offs between plant defense and growth. However, proof for this hypothesis has not been demonstrated yet. In this study the impact of defense hormonal crosstalk on disease resistance and fitness of Arabidopsis thaliana when under multi-species attack was investigated. Induction of SA- or JA/ABA-dependent defense responses by the biotrophic pathogen Hyaloperonospora arabidopsidis or the herbivorous insect Pieris rapae, respectively, was shown to reduce the level of induced JA/ET-dependent defense against subsequent infection with the necrotrophic pathogen Botrytis cinerea. However, despite the enhanced susceptibility to this second attacker, no additional long-term negative effects were observed on plant fitness when plants had been challenged by multiple attackers. Similarly, when plants were grown in dense competition stands to enlarge fitness effects of induced defenses, treatment with a combination of SA and MeJA did not cause additional negative effects on plant fitness in comparison to the single MeJA treatment. Together, these data support the notion that defense hormonal crosstalk in plants during multi-attacker interactions allows plants to prioritize their defenses, while limiting the fitness costs associated with induction of defenses.

Nucleotide-binding leucine-rich repeat ( NB- LRR, or NLR) receptors mediate pathogen recognition. The Arabidopsis thaliana NLR RPP1 recognizes the tandem WY-domain effector ATR1 from the oomycete Hyaloperonospora arabidopsidis through direct association with C-terminal LRRs., We isolated and characterized homologous NLR genes RPP1-EstA and RPP1-ZdrA from two Arabidopsis ecotypes, Estland (Est-1) and Zdarec (Zdr-1), responsible for recognizing a novel spectrum of ATR1 alleles., RPP1-EstA and -ZdrA encode nearly identical NLRs that are phylogenetically distinct from known immunity-activating RPP1 homologs and possess greatly expanded LRR domains. Site-directed mutagenesis and truncation analysis of ATR1 suggests that these homologs recognize a novel surface of the 2nd WY domain of ATR1, partially specified by a C-terminal region of the LRR domain. Synteny comparison with RPP1 loci involved in hybrid incompatibility suggests that these functions evolved independently., Closely related RPP1 homologs have diversified their recognition spectra through LRR expansion and sequence variation, allowing them to detect multiple surfaces of the same pathogen effector. Engineering NLR receptor specificity may require a similar combination of repeat expansion and tailored amino acid variation. [ABSTRACT FROM AUTHOR]

Experiments were conducted on the role of intra- and inter-genotypic competition in ecological processes operating at the population scale in diseased plant populations., Combinations of Arabidopsis thaliana genotypes showing variation for phenotypic traits relating to competitive ability and pathogen compatibility were infected with the oomycete Hyaloperonospora arabidopsidis and Turnip yellows virus in separate experiments. Plant fitness and competitive ability were estimated from phenotypic measurements., Pathogen-induced reduction in competitive ability for susceptible genotypes increased the competitive ability of resistant genotypes, resulting in maintenance of yield via competitive release. The two diseases had different effects on competitive interactions between plants. In experiments involving the oomycete, the highest yields were produced by mixtures of two weakly competing genotypes., The Arabidopsis model system has elucidated the ecological processes by which compensatory competitive interactions can increase the buffering capacity of plant populations under pathogen attack. Highly competitive genotypes may not maximize the productivity of the population as a whole, as they may over-yield at the expense of less competitive, more productive genotypes. The specific outcomes of competitive interactions cannot be generalized because they depend on the disease and the host genotypes. [ABSTRACT FROM AUTHOR]

Plant genes differentially expressed during plant-pathogen interactions can be important for host immunity or can contribute to pathogen virulence. Large-scale transcript profiling studies, such as microarray- or mRNA-seq-based analyses, have revealed hundreds of genes that are differentially expressed during plant-pathogen interactions. However, transcriptional responses limited to a small number of cells at infection sites can be difficult to detect using these approaches, as they are under-represented in the whole-tissue datasets typically generated by such methods. This study examines the interactions between A rabidopsis thaliana ( Arabidopsis) and the pathogenic oomycete H yaloperonospora arabidopsidis ( H pa) by enhancer trapping to uncover novel plant genes involved in local infection responses. We screened a β-glucuronidase ( GUS) reporter-based enhancer-trap population for expression patterns related to H pa infection. Several independent lines exhibited GUS expression in leaf mesophyll cells surrounding H pa structures, indicating a regulatory response to pathogen infection. One of these lines contained a single enhancer-trap insertion in an exon of At1g08800 ( MyoB1, M yosin B inding P rotein 1) and was subsequently found to exhibit reduced susceptibility to Hpa. Two additional Arabidopsis lines with T-DNA insertions in exons of MyoB1 also exhibited approximately 30% fewer spores than wild-type plants. This study demonstrates that our enhancer-trapping strategy can result in the identification of functionally relevant pathogen-responsive genes. Our results further suggest that MyoB1 either positively contributes to Hpa virulence or negatively affects host immunity against this pathogen. [ABSTRACT FROM AUTHOR]

The accurate quantification of disease severity is important for the assessment of host-pathogen interactions in laboratory or field settings. The interaction between Arabidopsis thaliana and its naturally occurring downy mildew pathogen, Hyaloperonospora arabidopsidis ( Hpa), is a widely used reference pathosystem for plant-oomycete interactions. Current methods for the assessment of disease severity in the Arabidopsis- Hpa interaction rely on measurements at the terminal stage of pathogen development; namely, visual counts of spore-producing structures or the quantification of spore production with a haemocytometer. These assays are useful, but do not offer sensitivity for the robust quantification of small changes in virulence or the accurate quantification of pathogen growth prior to the reproductive stage. Here, we describe a quantitative real-time polymerase chain reaction (q PCR) assay for the monitoring of Hpa growth in planta. The protocol is rapid, inexpensive and can robustly distinguish small changes in virulence. We used this assay to investigate the dynamics of early Hpa mycelial growth and to demonstrate the proof of concept that this assay could be used in screens for novel oomycete growth inhibitors. [ABSTRACT FROM AUTHOR]

The hormone salicylic acid (SA) generally induces plant defenses against biotrophic pathogens. Jasmonic acid (JA) and its oxylipin derivatives together with ethylene (ET) are generally important hormonal regulators of induced plant defenses against necrotrophic pathogens, whereas JAs together with abscisic acid (ABA) are implicated in induced plant defenses against herbivorous insects. Hormonal crosstalk between the different plant defense pathways has often been hypothesized to be a cost-saving strategy that has evolved as a means of the plant to reduce allocation costs by repression of unnecessary defenses, thereby minimizing trade-offs between plant defense and growth. However, proof for this hypothesis has not been demonstrated yet. In this study the impact of hormonal crosstalk on disease resistance and fitness of Arabidopsis thaliana when under multi-species attack was investigated. Induction of SA- or JA/ABA-dependent defense responses by the biotrophic pathogen Hyaloperonospora arabidopsidis or the herbivorous insect Pieris rapae, respectively, was shown to reduce the level of induced JA/ET-dependent defense against subsequent infection with the necrotrophic pathogen Botrytis cinerea. However, despite the enhanced susceptibility to this second attacker, no additional long-term negative effects were observed on plant fitness when plants had been challenged by multiple attackers. Similarly, when plants were grown in dense competition stands to enlarge fitness effects of induced defenses, treatment with a combination of SA and MeJA did not cause additional negative effects on plant fitness in comparison to the single MeJA treatment. Together, these data support the notion that hormonal crosstalk in plants during multi-attacker interactions allows plants to prioritize their defenses, while limiting the fitness costs associated with induction of defenses. [ABSTRACT FROM AUTHOR]

Importin-αs are essential adapter proteins that recruit cytoplasmic proteins destined for active nuclear import to the nuclear transport machinery. Cargo proteins interact with the importin-α armadillo repeat domain via nuclear localization sequences ( NLSs), short amino acids motifs enriched in Lys and Arg residues. Plant genomes typically encode several importin-α paralogs that can have both specific and partially redundant functions. Although some cargos are preferentially imported by a distinct importin-α it remains unknown how this specificity is generated and to what extent cargos compete for binding to nuclear transport receptors. Here we report that the effector protein HaRxL106 from the oomycete pathogen Hyaloperonospora arabidopsidis co-opts the host cell's nuclear import machinery. We use Ha Rx L106 as a probe to determine redundant and specific functions of importin-α paralogs from Arabidopsis thaliana. A crystal structure of the importin-α3/ MOS6 armadillo repeat domain suggests that five of the six Arabidopsis importin-αs expressed in rosette leaves have an almost identical NLS-binding site. Comparison of the importin-α binding affinities of Ha Rx L106 and other cargos in vitro and in plant cells suggests that relatively small affinity differences in vitro affect the rate of transport complex formation in vivo. Our results suggest that cargo affinity for importin-α, sequence variation at the importin-α NLS-binding sites and tissue-specific expression levels of importin-αs determine formation of cargo/importin-α transport complexes in plant cells. [ABSTRACT FROM AUTHOR]

Plants are highly capable of recognizing and defending themselves against invading microbes. Adapted plant pathogens secrete effector molecules to suppress the host's immune system. These molecules may be recognized by host-encoded resistance proteins, which then trigger defense in the form of the hypersensitive response ( HR) leading to programmed cell death of the host tissue at the infection site. The three proteins PEN1, PEN2 and PEN3 have been found to act as central components in cell wall-based defense against the non-adapted powdery mildew Blumeria graminis fsp. hordei ( Bgh). We found that loss of function mutations in any of the three PEN genes cause decreased hypersensitive cell death triggered by recognition of effectors from oomycete and bacterial pathogens in Arabidopsis. There were considerable additive effects of the mutations. The HR induced by recognition of AvrRpm1 was almost completely abolished in the pen2 pen3 and pen1 pen3 double mutants and the loss of cell death could be linked to indole glucosinolate breakdown products. However, the loss of the HR in pen double mutants did not affect the plants' ability to restrict bacterial growth, whereas resistance to avirulent isolates of the oomycete Hyaloperonospora arabidopsidis was strongly compromised. In contrast, the double and triple mutants demonstrated varying degrees of run-away cell death in response to Bgh. Taken together, our results indicate that the three genes PEN1, PEN2 and PEN3 extend in functionality beyond their previously recognized functions in cell wall-based defense against non-host pathogens. [ABSTRACT FROM AUTHOR]