Receptor-Like Kinases BAK1 and SOBIR1 Are Required for Necrotizing Activity of a Novel Group of Sclerotinia sclerotiorum Necrosis-Inducing Effectors.

Sclerotinia sclerotiorum is a characteristic necrotrophic plant pathogen and is dependent on the induction of host cell death for nutrient acquisition. To identify necrosis-inducing effectors, the genome of S. sclerotiorum was scanned for genes encoding small, secreted, cysteine-rich proteins. These potential effectors were tested for their ability to induce necrosis in Nicotiana benthamiana via Agrobacterium-mediated expression and for cellular localization in host cells. Six novel proteins were discovered, of which all but one required a signal peptide for export to the apoplast for necrotizing activity. Virus-induced gene silencing revealed that the five necrosis-inducing effectors with a requirement for secretion also required the plant co-receptor-like kinases Brassinosteroid Insensitive 1-Associated Receptor Kinase 1 (BAK1) and Suppressor of BAK1-Interacting Receptor-like Kinase 1 (SOBIR1) for the induction of necrosis. S. sclerotiorum necrosis-inducing effector 2 (SsNE2) represented a new class of necrosis-inducing proteins as orthologs were identified in several other phytopathogenic fungi that were also capable of inducing necrosis. Substitution of conserved cysteine residues with alanine reduced, but did not abolish, the necrotizing activity of SsNE2 and full-length protein was required for function as peptides spanning the entire protein were unable to induce necrosis. These results illustrate the importance of necrosis-inducing effectors for S. sclerotiorum virulence and the role of host extracellular receptor(s) in effector-triggered susceptibility to this pathogen.

The Brassica napus receptor-like protein RLM2 is encoded by a second allele of the LepR3/Rlm2 blackleg resistance locus.

Leucine-rich repeat receptor-like proteins (LRR-RLPs) are highly adaptable parts of the signalling apparatus for extracellular detection of plant pathogens. Resistance to blackleg disease of Brassica spp. caused by Leptosphaeria maculans is largely governed by host race-specific R-genes, including the LRR-RLP gene LepR3. The blackleg resistance gene Rlm2 was previously mapped to the same genetic interval as LepR3. In this study, the LepR3 locus of the Rlm2 Brassica napus line 'Glacier DH24287' was cloned, and B. napus transformants were analysed for recovery of the Rlm2 phenotype. Multiple B. napus, B. rapa and B. juncea lines were assessed for sequence variation at the locus. Rlm2 was found to be an allelic variant of the LepR3 LRR-RLP locus, conveying race-specific resistance to L. maculans isolates harbouring AvrLm2. Several defence-related LRR-RLPs have previously been shown to associate with the RLK SOBIR1 to facilitate defence signalling. Bimolecular fluorescence complementation (BiFC) and co-immunoprecipitation of RLM2-SOBIR1 studies revealed that RLM2 interacts with SOBIR1 of Arabidopsis thaliana when co-expressed in Nicotiana benthamiana. The interaction of RLM2 with AtSOBIR1 is suggestive of a conserved defence signalling pathway between B. napus and its close relative A. thaliana.