Mimicking the Host Regulation of Salicylic Acid: A Virulence Strategy by the Clubroot Pathogen Plasmodiophora brassicae.

The plant hormone salicylic acid (SA) plays a critical role in defense against biotrophic pathogens such as Plasmodiophora brassicae, which is an obligate pathogen of crucifer species and the causal agent of clubroot disease of canola (Brassica napus). P. brassicae encodes a protein, predicted to be secreted, with very limited homology to benzoic acid (BA)/SA-methyltransferase, designated PbBSMT. PbBSMT has a SA- and an indole-3-acetic acid-binding domain, which are also present in Arabidopsis thaliana BSMT1 (AtBSMT1) and, like AtBSMT1, has been shown to methylate BA and SA. In support of the hypothesis that P. brassicae uses PbBSMT to overcome SA-mediated defenses by converting SA into inactive methyl salicylate (MeSA), here, we show that PbBSMT suppresses local defense and provide evidence that PbBSMT is much more effective than AtBSMT1 at suppressing the levels of SA and its associated effects. Basal SA levels in Arabidopsis plants that constitutively overexpress PbBSMT compared with those in Arabidopsis wild-type Col-0 (WT) were reduced approximately 80% versus only a 50% reduction in plants overexpressing AtBSMT1. PbBSMT-overexpressing plants were more susceptible to P. brassicae than WT plants; they also were partially compromised in nonhost resistance to Albugo candida. In contrast, AtBSMT1-overexpressing plants were not more susceptible than WT to either P. brassicae or A. candida. Furthermore, transgenic Arabidopsis and tobacco plants overexpressing PbBSMT exhibited increased susceptibility to virulent Pseudomonas syringae pv. tomato DC3000 (DC3000) and virulent Pseudomonas syringae pv. tabaci, respectively. Gene-mediated resistance to DC3000/AvrRpt2 and tobacco mosaic virus (TMV) was also compromised in Arabidopsis and Nicotiana tabacum 'Xanthi-nc' plants overexpressing PbBSMT, respectively. Transient expression of PbBSMT or AtBSMT1 in lower leaves of N. tabacum Xanthi-nc resulted in systemic acquired resistance (SAR)-like enhanced resistance to TMV in the distal systemic leaves. Chimeric grafting experiments revealed that, similar to SAR, the development of a PbBSMT-mediated SAR-like phenotype was also dependent on the MeSA esterase activity of NtSABP2 in the systemic leaves. Collectively, these results strongly suggest that PbBSMT is a novel effector, which is secreted by P. brassicae into its host plant to deplete pathogen-induced SA accumulation.

Transposable element-assisted evolution and adaptation to host plant within the Leptosphaeria maculans-Leptosphaeria biglobosa species complex of fungal pathogens.

BACKGROUND: Many plant-pathogenic fungi have a tendency towards genome size expansion, mostly driven by increasing content of transposable elements (TEs). Through comparative and evolutionary genomics, five members of the Leptosphaeria maculans-Leptosphaeria biglobosa species complex (class Dothideomycetes, order Pleosporales), having different host ranges and pathogenic abilities towards cruciferous plants, were studied to infer the role of TEs on genome shaping, speciation, and on the rise of better adapted pathogens. RESULTS: L. maculans 'brassicae', the most damaging species on oilseed rape, is the only member of the species complex to have a TE-invaded genome (32.5%) compared to the other members genomes (<4%). These TEs had an impact at the structural level by creating large TE-rich regions and are suspected to have been instrumental in chromosomal rearrangements possibly leading to speciation. TEs, associated with species-specific genes involved in disease process, also possibly had an incidence on evolution of pathogenicity by promoting translocations of effector genes to highly dynamic regions and thus tuning the regulation of effector gene expression in planta. CONCLUSIONS: Invasion of L. maculans 'brassicae' genome by TEs followed by bursts of TE activity allowed this species to evolve and to better adapt to its host, making this genome species a peculiarity within its own species complex as well as in the Pleosporales lineage.

Two independent approaches converge to the cloning of a new Leptosphaeria maculans avirulence effector gene, AvrLmS-Lep2.

Brassica napus (oilseed rape, canola) seedling resistance to Leptosphaeria maculans, the causal agent of blackleg (stem canker) disease, follows a gene-for-gene relationship. The avirulence genes AvrLmS and AvrLep2 were described to be perceived by the resistance genes RlmS and LepR2, respectively, present in B. napus 'Surpass 400'. Here we report cloning of AvrLmS and AvrLep2 using two independent methods. AvrLmS was cloned using combined in vitro crossing between avirulent and virulent isolates with sequencing of DNA bulks from avirulent or virulent progeny (bulked segregant sequencing). AvrLep2 was cloned using a biparental cross of avirulent and virulent L. maculans isolates and a classical map-based cloning approach. Taking these two approaches independently, we found that AvrLmS and AvrLep2 are the same gene. Complementation of virulent isolates with this gene confirmed its role in inducing resistance on Surpass 400, Topas-LepR2, and an RlmS-line. The gene, renamed AvrLmS-Lep2, encodes a small cysteine-rich protein of unknown function with an N-terminal secretory signal peptide, which is a common feature of the majority of effectors from extracellular fungal plant pathogens. The AvrLmS-Lep2/LepR2 interaction phenotype was found to vary from a typical hypersensitive response through intermediate resistance sometimes towards susceptibility, depending on the inoculation conditions. AvrLmS-Lep2 was nevertheless sufficient to significantly slow the systemic growth of the pathogen and reduce the stem lesion size on plant genotypes with LepR2, indicating the potential efficiency of this resistance to control the disease in the field.