A new family of structurally conserved fungal effectors displays epistatic interactions with plant resistance proteins

Abstract

Recognition of a pathogen avirulence (AVR) effector protein by a cognate plant resistance (R) protein triggers a set of immune responses that render the plant resistant. Pathogens can escape this so-called Effector-Triggered Immunity (ETI) by different mechanisms including the deletion or loss-of-function mutation of the AVR gene, the incorporation of point mutations that allow recognition to be evaded while maintaining virulence function, and the acquisition of new effectors that suppress AVR recognition. The Dothideomycete Leptosphaeria maculans, causal agent of oilseed rape stem canker, is one of the few fungal pathogens where suppression of ETI by an AVR effector has been demonstrated. Indeed, AvrLm4-7 suppresses the recognition of AvrLm3 and AvrLm5-9 by the R proteins Rlm3 and Rlm9, respectively. The presence of AvrLm4-7 does not impede AvrLm3 and AvrLm5-9 expression, and the three AVR proteins do not appear to physically interact. To decipher the epistatic interaction between these L. maculans AVR effectors, we determined the crystal structure of AvrLm5-9 and obtained a 3D model of AvrLm3, based on the crystal structure of Ecp11-1, a homologous avirulence effector candidate from Fulvia fulva. Despite a lack of sequence similarity, AvrLm5-9 and AvrLm3 are structural analogues of AvrLm4-7 (structure previously characterized). Structure-informed sequence database searches identified a larger number of putative structural analogues among L. maculans effector candidates, including the avirulence effector AvrLmS-Lep2, all produced during the early stages of oilseed rape infection, as well as among effector candidates from other phytopathogenic fungi. These structural analogues were called LARS (for Leptosphaeria AviRulence-Suppressing) effectors. Remarkably, transformants of L. maculans expressing one of these structural analogues, Ecp11-1, triggered Rlm3-mediated immunity. Furthermore, this recognition could be suppressed by AvrLm4-7. These results suggest that Ecp11-1 has the same function as AvrLm3, or that the Ecp11-1 structure is sufficiently close to that of AvrLm3 to be recognized by Rlm3.Author summaryAn efficient strategy to control fungal diseases in the field is genetic control using resistant crop cultivars. Crop resistance mainly relies on gene-for-gene relationships between plant resistance (R) genes and pathogen avirulence (AVR) genes, as defined by Flor in the 1940s. However, gene-for-gene relationships diversify to increased complexity in the course of plant-pathogen co-evolution. Resistance against the plant-pathogenic fungus Leptosphaeria maculans by Brassica napus and other Brassica species relies on recognition of effector (AVR) proteins by R proteins; however, L. maculans is able to produce an effector that suppresses a subset of these specific recognitions. Using a protein structure approach, we revealed structural analogy between different effectors within L. maculans and other plant-pathogenic species in the Dothideomycetes and Sordariomycetes classes, defining a new family of effectors called LARS, and providing a clue to the mechanisms behind recognition suppression. Cross-species expression of one LARS effector from Fulvia fulva, a pathogen of tomato, in L. maculans triggers recognition by an oilseed rape resistant cultivar. These results highlight the need to integrate knowledge on effector structures to improve resistance management and to develop broad-spectrum resistances for multi-pathogen control of diseases.

The structural repertoire ofFusarium oxysporumf. sp.lycopersicieffectors revealed by experimental and computational studies

“…The MAX effector family were originally defined as AVR effectors from M. oryzae and ToxB from P. tritici-repentis [28] but pattern-based sequence searches suggest they are widely distributed amongst the Dothideomycetes [28,48]. Similarly, LARS effectors, defined in Leptosphaeria maculans and Fulvia fulva, have structural homologues predicted in at least 13 different fungal species [30]. For the newly described FOLD effectors, homologues of SIX1 (Avr3), SIX6 and SIX13 are found in many formae speciales of F. oxysporum and are also present in several fungal genera including Colletotrichum, Ustilaginoidea, Leptosphaeria, Pyrenophora and Bipolaris.…”

“…The LARS effectors represent another example of effectors that can activate and suppress resistance-gene-mediated immunity. AvrLm4-7 can prevent recognition of AvrLm3 and AvrLm9 (all LARS structural homologues [30]), by their cognate Rlm receptors [75,76]. Rlm9 encodes a wall-associated kinase [77], but the identify of Rlm4, -7 and -3 remain unknown.…”

“…To date, five fungal effector families have been defined based on experimentally-determined structural homology, including the MAX [28], RALPH [29,45,46], ToxA-like [25][26][27], LARS [30,47] and FOLD effectors, defined here. Effectors that fall within many of these structural families are shared across distantly related fungal species.…”

Apoplastic effector candidates of a foliar forest pathogen trigger cell death in host and non-host plants

“…Recent studies showed that some effector protein families share structural similarity rather than sequence similarity with effectors from different fungi 27,28 . Thus the DsCEs were assessed for possible structural similarities to characterised proteins using HHpred.…”

Computational structural genomics unravels common folds and predicted functions in the secretome of fungal phytopathogenMagnaporthe oryzae

“…Avirulent effectors Avr1-CO39, Avr-Pia and AvrPiz-t from M. oryzae and ToxB from the wheat pathogen Pyrenophora tritici-repentis are unrelated by their sequences; however, they share a common β sandwich fold and are together classified as MAX (Magnaporthe Avrs and ToxB-like) effectors (16). Similarly, in phytopathogenic Leptosphaeria maculans, AvrLm3, AvrLm4-7 and AvrLm5-9 are structural analogs belonging to LARS (Leptosphaeria avirulence-suppressing) effectors (17,18). In powdery mildews, such as Blumeria graminis, RALPH (RNase-like proteins expressed in haustoria) effectors, that commonly adopt the structural folds of RNase despite highly divergent sequences, represent a significant portion of the effectorome (19)(20)(21).…”